The Dementias Early Diagnosis and Evaluation
The Dementias Early Diagnosis and Evaluation
The Dementias
Early Diagnosis and Evaluation
edited by
Karl Herholz
Wolfson Molecular Imaging Centre
University of Manchester
Manchester, U.K.
Daniela Perani
Vita-Salute San Raffaele University
San Raffaele Scientific Institute
Milan, Italy
Chris Morris
Wolfson Unit of Clinical Pharmacology
University of Newcastle upon Tyne
Newcastle upon Tyne, U.K.
Our motivation to compile this book stems from our exposure to a number of
issues that are difficult to reconcile. Most people probably welcome the increase
in longevity that is observed in many countries today, but this also brings with it a
substantial increase in dementia with prevalence and incidence rising steeply
beyond 70 years of age. It is widely acknowledged that this imposes severe
stress on families and caregivers and risks the failure of social welfare systems.
Dementia may therefore become one of the most difficult challenges for societies
in this century. Yet, we are far away from a medical solution. Even if a drug that
could slow or halt the progression of dementing diseases was made available
tomorrow, it probably would have only limited effect since too much brain
function has been lost to degenerative brain disease once the clinical symptoms of
dementia become clearly evident. Restoration of normal cognition will therefore
remain an elusive goal. Thus, there is the danger that any available drug would
either be given too late to prevent dementia and its consequences, or it would be
given indiscriminately to people with nonspecific complaints or even without any
symptoms. Since only a small proportion of these individuals are likely to
develop dementia within the next few years, precious medical resources would be
wasted. Early and accurate diagnosis of dementing diseases before the onset
of dementia therefore becomes of paramount importance if we are to avoid such
a situation and use preventative treatment strategies efficiently once they become
available.
Over the past few years, many new methods and techniques have been
developed to improve early diagnosis. Most of these require special training and
expert knowledge for application and an understanding of their potential and
limitations. In this book, active researchers who are experts in their respective
fields present and evaluate these methods to provide a guide to their use in early
iii
iv Preface
dementia diagnosis and assessment. It is, however, quite clear that no single
method can cover all aspects that are relevant for early diagnosis. With this
overview we hope to provide comprehensive information to clinicians,
researchers, and the pharmaceutical industry about which methods are the most
effective ones for screening, early diagnosis of symptomatic subjects, and
distinguishing between different dementing diseases. While we cannot yet provide
definitive answers to all these questions, we hope that the evaluation will provide
guidance for further research.
Clinical evaluation and care are the basis of dementia diagnosis and
treatment, and we therefore put the chapter on clinical issues at the beginning.
Neuropsychology provides methods for the objective and quantitative assessment
of symptoms and behavioral and cognitive deficits, and therefore this naturally
takes its place close to clinical evaluation. Next comes cerebrospinal fluid and
blood biomarkers that hold promise in complementing symptom-based diagnosis
by adding biochemical information on the underlying disease process. Chapters
on genetics and novel molecular targets provide the latest information on risk
factors and molecular mechanisms with their perspectives for early diagnosis.
After that we enter the wide arena of in vivo imaging methods, which have
undergone rapid development and bewildering diversification in recent years,
ranging from structural imaging with computed tomography and magnetic
resonance imaging to functional and molecular imaging with functional magnetic
resonance imaging, magnetic resonance spectroscopy, single-photon emission
computed tomography, and positron emission tomography. Most of these are
ready to be applied in the clinical arena, but quite often their actual value and
efficacy is still under dispute. Authors therefore explain the techniques and
provide information on diagnostic sensitivity and specificity when available. The
methods chapters are then concluded with a view from neuropathology, which
still provides the most detailed methods for tissue examination and therefore
sets the diagnostic gold standard, although the significance of post-mortem
studies in early dementia is limited because of the lack of clinical follow-up data,
for obvious reasons. We did not want to leave our readers with just a compilation
of methods and data but felt that an overview and final evaluation, although
necessarily somewhat subjective, should conclude the book. We also address
some of the wider implications of improved early diagnosis that need considera-
tion when we enter this extremely important clinical field, which may hold
dangers, but hopefully many surprises as well.
Karl Herholz
Daniela Perani
Chris Morris
Acknowledgments
We are very grateful to the authors, who are all busy researchers but dedicated
a significant amount of their time to the book. Their contribution cannot be
overestimated. This interdisciplinary project would probably not have been
possible without previous scientific collaboration of the editors and some of the
authors, which was supported by the European Commission in their framework
programs 4 to 6. Last but not least, we thank the publisher’s editorial and
production team, especially Geoff Greenwood and Dana Bigelow, for their
continuous encouragement and support. The publication of this book was
generously supported by an educational grant from GE Healthcare.
v
Contents
Preface . . . . iii
Acknowledgments . . . . v
Contributors . . . . xiii
vii
viii Contents
Index . . . . 443
Contributors
xiii
xiv Contributors
Norman L. Foster
Imaging and Research, Center for Alzheimer’s Care, and Department
of Neurology and The Brain Institute, University of Utah, Salt Lake City,
Utah, U.S.A.
INTRODUCTION
The diagnosis and management of dementia will be a major challenge for health
care systems in the coming decades. As the world’s population ages, the
prevalence of dementia will grow dramatically (1). Alzheimer’s disease (AD),
the most common cause of dementia, accounts for much of the increasing
prevalence of dementia with age (2). However, many other causes of dementia,
such as stroke and Parkinson’s disease, also increase dramatically as people
become older. The care of those with dementia causes great burdens on caregivers
and society. The number of individuals with dementia will grow most rapidly in the
developing world, but dementing diseases will have their greatest social impact in
Japan, Europe, and North America, where their proportion of the population will
be highest. Prevention and improved diagnosis and treatment of cognitive
impairment and dementia offer the only hope to alter this grim equation.
Fortunately, there has been an explosion of information about neuro-
degenerative diseases, and dementia has moved from being a backwater to the
vanguard of neuroscience research. New insights and conceptual breakthroughs
have led to improved detection and evaluation of dementia, and there are many
more new approaches now in development. There is every reason to expect these
advances will continue and be translated into clinical practice. Awareness and
application of these recent developments provide great opportunities for
physicians to help patients and society.
1
2 Foster
DEMENTIA EVALUATION
The Role of Dementia Evaluation
Patients and families have many reasons for seeking a physician’s advice when
memory loss or cognitive impairments appear (Table 1). The hope of treatment that
improves symptoms is a major motivation, but other important concerns also
should be addressed. Physicians too often focus solely on treatment and fail to
recognize the great value patients and families place on accurate diagnosis. There
is comfort in understanding the cause of symptoms and in knowing that everything
that should be done has been done. An accurate and specific diagnosis also
empowers physicians. Once the cause of dementia is determined, the physicians
can use their specialized knowledge of the disease to guide management and
coordinate care. It becomes possible to develop a rational and evidence-based care
plan appropriate for that disease. Family members also may be seeking help for
themselves. An authoritative dementia evaluation can resolve conflicting opinions
among family members and clarify their role in care. With a detailed management
plan and prognostic information, patients, families, and physicians can plan for the
future together.
Physicians often are reluctant to evaluate dementia because it opens a
“Pandora’s box” of concerns they aren’t prepared to address (3). Many physicians
lack the experience and confidence to perform an evaluation. Even if an evaluation
is performed, physicians may be reluctant to inform patients and families of their
diagnosis (4). Information about prognosis also may be withheld, even though it
greatly benefits patients and their families (5). Usually patients and families aren’t
satisfied until they learn the specific cause of cognitive impairment; generalities,
platitudes, and nostrums aren’t sufficient. In one study, caregivers reported that a
correct diagnosis of AD was provided in only 38% of initial physician
consultations, causing them to seek care from someone else (6). Most recognize
the limits of medicine, and appreciate a definitive diagnosis, even when few drug
treatments can be offered. Patients with suspected neurological disease have less
distress about their symptoms and less anxiety when they receive a definitive
diagnosis (7). Physicians therefore should use whatever effective means are
available to accurately determine the cause of dementia.
epidemiological studies). Criteria for these conditions are not consistent from
report to report, and their use should be scrutinized carefully. The prevalence of
cognitive impairment insufficient to cause functional impairment appears to be
greater than the prevalence of dementia, but more studies are needed (12).
Despite the difficulties of definition, recognizing these conditions has more than
heuristic value. They represent a group of patients with a significantly increased
risk of developing dementia over subsequent years and are an appropriate target
for new therapies (13).
It may be helpful to distinguish different kinds of cognitive impairment
insufficient to be classified as dementia. Since different dementing diseases have
predominant deficits in different cognitive domains, it is logical to assume that
their initial symptoms also differ. For example, isolated memory impairment
would be expected to precede the development of dementia in AD, where memory
loss usually is the most prominent symptom. Likewise, progressive aphasia may
precede frontotemporal dementia (FTD) (14). Thus individuals with amnestic
MCI and predominant or isolated memory loss are believed to be more likely to
progress to dementia caused by AD, and those with non-amnestic MCI may
precede other forms of dementia (15). It also may be useful to classify patients by
presumed etiology, since this can have prognostic significance (16,17).
Deriving reliable criteria to identify prodromal dementia based purely upon
clinical grounds turns out to be more difficult than it might first appear. Not only
must multiple areas of cognition be considered, but boundaries of both normal
cognition for age and impaired cognition sufficient to represent dementia also must
be defined. Detailed neuropsychological testing provides age-appropriate norms to
guide clinical judgment and explicit operational criteria for clinical trials. MCI
differs from CIND by requiring a voiced complaint of impairment. An expressed
memory complaint is important because it is a strong predictor of AD and a face
valid criterion for a change in ability (18). Individuals categorized as having MCI
are more likely than those with CIND to progress to unequivocal dementia (13,19).
In a diverse clinical population where diagnosis and management must be
provided to everyone, the early recognition of prodromal dementia must rely on
physician judgment and interpretation, rather than solely on neuropsychological
test results or strict application of criteria.
Table 4 Characteristic Presentation and Course of Mild Cognitive Impairment and the Most Common Dementing Diseases
Disease Usual features Diagnostic guidelines
Mild cognitive impairment Insidious onset of progressive decline in cognitive abilities ADCS criteria (13); International Workshop
insufficient to cause impairment in everyday activities; criteria (20)
must be associated with a complaint; several subtypes
have been proposed: the amnestic form in which
memory loss is predominant appears to be most likely to
progress to Alzheimer’s disease
Alzheimer’s disease Insidious onset of gradually progressive memory loss; NINCDS-ADRDA
memory loss remains the most prominent feature as criteria (21)
aphasia, apraxia, impaired judgment, and behavior
change develop; seizures and incontinence late in the
disease
Vascular dementia Sudden onset, stepwise progression of dementia associated NINDS-AIREN criteria (22); Modified
with ischemic stroke; usually with focal motor and Hachinski Ischemic Score (23)
sensory deficits, although these deficits may resolve;
cognitive deficits variable and correspond to the location
of strokes; frontal lobe impairment may be most obvious
with subcortical stroke; seizures and incontinence may
occur early in the course
Foster
Dementia with Lewy bodies Insidious onset of gradually progressive dementia with Consensus second workshop criteria (24)
parkinsonism (either within a year of developing
Parkinson’s disease or alternatively the development of
parkinsonism concurrent with dementia or soon there-
after); spontaneous visual hallucinations, unexplained
fluctuations in symptom severity; visuo-spatial deficits
The Clinical Issues
that computed tomography (CT) and magnetic resonance imaging (MRI) aid the
recognition of many neurological illnesses causing dementia (Table 8). Although
many of these diseases often cause focal abnormalities, the neurological
examination alone may be insensitive for early detection or when patients are
unable to adequately participate in a detailed assessment.
Simply identifying an abnormal laboratory test or brain scan is insufficient;
one must interpret the results and their clinical significance. It is important to
realize that “Occam’s razor,” or the search for a single unifying diagnosis to
explain symptoms and signs, is not always appropriate when evaluating older
patients (29). Since it is common for a medical illness to exacerbate an
underlying dementing illness or for dementing diseases to co-exist and interact,
the principle of parsimony of diagnosis may lead one astray (30,31). It is more
appropriate to consider the possibility that multiple pathologies are causing
cognitive decline.
It can be very difficult to be sure when two or more conditions are
contributing to a patient’s dementia, yet this has major implications for deciding
on treatment and assessing response. Serial examinations may help disentangle
symptoms and reveal which of several conditions is the predominant cause of the
dementia. For instance, in a patient with mixed dementia due to AD and stroke,
gradual progression of dementia without further strokes on MRI suggests that AD
has caused the worsening of symptoms. By contrast, if there is sudden worsening
Table 8 Causes of Dementia that Can Be Detected Early with Structural Brain Imaging
Primary and metastatic brain tumors
Ischemic stroke
Intracerebral hemorrhage
Subarachnoid hemorrhage
Subdural hematoma
Multiple sclerosis
Brain abscess
Progressive multifocal leukoencephalopathy
Traumatic brain injury
Normal pressure hydrocephalus
Creutzfeldt–Jakob disease (diffusion-weighted and FLAIR MRI only)
Abbreviation: FLAIR MRI, fluid attenuated inversion recovery magnetic resonance imaging.
12 Foster
of dementia associated with new focal deficits, stroke is implicated as the cause of
worsening and is likely to be more important. Unfortunately, awaiting serial
observations entails delaying diagnosis and appropriate interventions.
In most cases, the cause of dementia currently can be determined with a
high degree of accuracy, but not complete certainty. Consensus diagnostic
criteria are available for several dementing diseases and can help, but they cannot
address all clinical situations. Physicians must approach diagnosis with
conviction and humility, recognizing the limits of current knowledge. The
cause of cognitive impairment always should be considered open to review and
possible revision until it is confirmed by pathologic examination (usually
impossible during life) or the identification of a causative genetic mutation
(relevant only in early-onset familial dementia). It is appropriate for physicians to
recommend that families always obtain a postmortem examination to confirm
clinical diagnoses, particularly because the results may have substantial
implications for other family members.
status epilepticus. However, despite such extensive testing, it still may be difficult
to determine the cause of rapidly progressive dementia.
It is particularly important to identify Creutzfeldt–Jakob disease (CJD) so
proper precautions can be used to prevent the potential iatrogenic trans-
mission (51). Definitively identifying CJD is difficult, but several clinical
findings and laboratory tests often are abnormal in CJD and can aid diagnosis.
Sequential EEGs may demonstrate the evolution of typical periodic discharges.
MRI can reveal focal abnormalities in the cerebral cortex or striatum with
diffusion-weighted or FLAIR sequences (52). High levels of the 14-3-3 protein
usually can be detected in the spinal fluid (53). Clinical criteria for CJD have been
developed incorporating these features (54).
Despite such clues, the diagnosis of CJD can remain elusive. Although
diagnosis can often be made easily when typical features have evolved, early
definitive identification of CJD is not easy, and a substantial group of patients
have atypical features. CJD can have a surprising variety of clinical
presentations. There can be initial memory loss that suggests AD, focal cognitive
deficits that suggest a focal lesion, or a gait disorder that suggests a
neurodegenerative disease. Some of these clinical phenotypes can be explained
by variations in prion protein glycotype and its interaction with prion genotype,
but this information currently is available only after a brain biopsy or
autopsy (55). New variant CJD associated with bovine spongiform encephalo-
pathy also has atypical features (56). Current laboratory testing for CJD is also
fallible and cannot be relied upon when symptoms are atypical. While a spinal
fluid 14-3-3 protein can be helpful if it is elevated, it is not always abnormal in
CJD, and it can be elevated in several other disorders associated with rapid brain
destruction (57). Brain biopsy is sometimes the only way to avoid the serious
consequences of misdiagnosis or treatment delay.
Rapidly progressive dementia illustrates how clinical methods and
diagnostic technology sometimes must be reconciled. The serious implications
of a rapidly progressive dementia must be weighed against the costs and real risks
of brain biopsy to the patient, surgical team, and pathology staff. The quandary of
The Clinical Issues 21
frontal association cortex. Aphasia also is not unique to FTD, and has long been
considered a cardinal feature of AD as indicated in diagnostic criteria for
AD (21). Diagnostic criteria suggest FTD can be distinguished from AD by the
relatively early onset of behavioral and language symptoms, but this requires
considerable subjective medical judgment. It also may be helpful to identify
behavioral and language symptoms often seen in FTD but uncommon in AD,
such as disinhibition, apathy, perseverative behavior, stereotypy of speech,
echolalia, and mutism. However, their reliability and specificity for FTD are
uncertain, and improvements in clinical criteria for FTD are still needed (65).
Nevertheless, such differences in type, predominance, and evolution of
symptoms are the basis for behavioral inventories meant to aid clinical
diagnosis (43). Structural brain imaging may help distinguish AD and FTD by
contrasting the severity of atrophy in involved brain regions (66). Probably the
most frequent pathologic abnormality in FTD is nonspecific neuronal loss and
gliosis with or without vacuolization. However, FTD sometimes is caused by
distinctive abnormalities in tau, ubiquitin, or neurofilament protein (67,68).
Eventually these should permit the development of biomarkers that identify
histopathological subtypes of FTD.
DLB is another important AD mimic that now appears to be the second most
common cause of dementia in the United States. It accounted for approximately
20% of all dementia cases in recent autopsy series (69). Although DLB always has
pathologic changes characteristic of Parkinson’s disease, it may not have
noticeable motor signs or other distinctive features (70). DLB has the same
biochemical signature of dopamine deficiency typical of Parkinson’s disease and
also has characteristic deposits of ubiquitin in cortical Lewy bodies; it is realistic to
expect new diagnostic methods could exploit these features and aid in diagnosis.
cognitive symptom onset (24). The criteria indicate that when spontaneous motor
symptoms precede the onset of dementia, the cause is Parkinson’s disease with
dementia. If motor symptoms and dementia occur simultaneously early in the
illness, the clinical diagnosis is DLB. However, if motor symptoms develop late
in a progressive dementia, the cause is AD. Additional criteria address other
symptoms and amplify these categories, but symptom onset remains critical to
classification. Unfortunately, rules of symptom onset are arbitrary and require
subjective interpretation of historical information that sometimes is limited or
unreliable. These clinical criteria understandably have only modest ability to
predict pathology (71). New technology to replicate biochemical and anatomic
information used by pathologists for classification would aid diagnosis and
development of more specific treatments enormously.
The challenges of clinical diagnosis of parkinsonism and dementia are
understandable. Because continued use of drugs may be clinically necessary, it
may be impossible to determine the onset of motor symptoms or whether they are
spontaneous, as required by criteria. Extrapyramidal symptoms also are
frequently seen in AD, although they usually begin later than in DLB. In one
study that monitored motor symptoms prospectively, over half of patients with
moderately severe AD develop new parkinsonian signs over two years (72).
Consequently, many disorders causing parkinsonism also are AD mimics. For
example, FTD may cause parkinsonism. This is particularly relevant in FTD with
parkinsonism linked to chromosome 17 (FTDP-17), an early-onset, autosomal
dominant disorder. In some families with FTDP-17, parkinsonism even may be
the presenting symptom, and much more evident than dementia (73).
The consideration of additional symptoms considered in criteria for DLB
may not resolve the difficulty with clinical diagnosis. For example, consensus
criteria use visual hallucinations and fluctuations to distinguish DLB. However,
medications and AD may cause visual hallucinations (74). Family reports of
fluctuations can be difficult to interpret, and subjective clinical judgment is
required to know when fluctuations are significant.
Pathologic studies also suggest why clinical diagnosis is so challenging.
Cerebral Lewy bodies are accompanied by pathological changes of AD about
two-thirds of the time (75). Thus some have called this “Lewy body variant
24 Foster
of AD” (76). Much additional work will be needed to clarify these difficult
clinical issues.
aids in identifying the specific cause of dementia, it plays an important role in the
selection of initial treatment. Perhaps less well recognized is how the results of
diagnostic brain imaging can be incorporated into subsequent treatment
decisions. A specific diagnosis helps to predict treatment outcomes. When the
response to appropriate disease specific therapy is unexpected, the accuracy of
the clinical diagnosis should be questioned. For example, an explanation other
than depression should be sought when severe apathy fails to improve with an
anti-depressant. In this situation, FTD should be considered. Likewise, the motor
symptoms of idiopathic Parkinson’s disease almost always respond to treatment
with dopaminergic agents. If they have never been beneficial, the diagnosis of
Parkinson’s disease with dementia is difficult to sustain, and another dementing
disorder is more likely. In diagnostic reassessments like these, additional brain
imaging methods can be considered or a repeat study that may reveal new or
evolving abnormalities can help provide critical confirmatory information.
Since definitive diagnostic test results are usually lacking in dementing
disorders, selection of treatment also is not definitive. Diagnostic confidence
inevitably modifies physician judgments about treatment. The results of brain
imaging can alter diagnostic confidence and influence therapy, particularly as the
effects of initial treatments are monitored. For example, treatment with
cholinesterase inhibitors might be aggressively pursued when clinical and
imaging features are typical of AD, but discontinued if side effects develop when
clinical and imaging findings are discordant for this diagnosis. Likewise, a
therapeutic trial of drugs used to treat AD sometimes may be warranted when the
diagnosis is uncertain. However, they might be avoided or discontinued if
imaging finds predominant frontal abnormalities, since cholinesterase inhibitors
provide little benefit and may worsen symptoms in FTD (85).
There is also great interest in using brain imaging to evaluate new drug
therapies. There is evidence that longitudinal changes in MRI and glucose
metabolism measured by positron emission tomography may permit more rapid
and cost-effective clinical trials in dementia (86,87). However, validation of an
imaging surrogate biomarker for disease progression is a major undertaking.
Further investigations are needed before it is clear whether imaging results are
reliable and can be appropriately interpreted in complex clinical trials. For
example, one recent randomized trial of immunotherapy for AD found surprising
imaging results that appear to be discordant with clinical outcomes (88). In the
future, it may be possible with molecular imaging to design therapies for
dementing diseases based upon individual biochemical profiles and disease
staging, just as it is now starting to be used in clinical oncology (89,90).
SUMMARY
The evaluation of dementia is challenging, but worthwhile. Our ability to
determine the cause of dementia has improved, but much remains to be done.
There are many opportunities to refine our current methods at critical and difficult
26 Foster
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The Clinical Issues 31
Daniela Perani
Departments of Neuroscience and Nuclear Medicine, Vita-Salute San
Raffaele University and San Raffaele Scientific Institute, Milan, Italy
INTRODUCTION
Dementia is a syndrome characterized by the reduction of cognitive abilities
(most often memory) to the point that impairment in daily activities occurs.
Clinical assessment, supported by the results of neuropsychological tests, is at
present the only in vivo non-invasive screening and diagnostic tool for Alzheimer’s
disease (AD) and other disorders associated with dementia. Dementia diagnosis,
indeed, is based exclusively on clinical, behavioral, and neuropsychological
findings according to DSM-IV (1). The same principle applies to the diagnosis of
the most common causes of dementia: AD (2), vascular dementia (VaD) (3), and
frontotemporal dementia (FTD) (4). According to NINCDS-ADRDA criteria,
neuropsychological testing is required to confirm the presence of dementia,
established on clinical grounds.
The recent development of symptomatic pharmacological treatment for AD
and the possible introduction of new therapies in the near future make the
assessment of dementia at its different stages a great scientific and public health
33
34 Perani
NEUROPSYCHOLOGICAL ASSESSMENT
An ideal neuropsychological test should be psychometrically adequate. It should
thus be reliable, valid, and have a high sensitivity and high specificity.
Reliability refers to the consistency of the information provided by the test,
if administered to the same subject by two different examiners, or by the same
examiner on two different occasions. An important issue, in particular for follow-
up and therapeutic studies, is the availability of alternate forms, to avoid test-retest
effects in the case of repeated examinations of the same subjects. The concept of
validity is complex and has multiple facets. Validity refers to the function to be
assessed in absolute terms (construct validity), or in relation to other tests, which
are considered to be valid from the former point of view (concurrent validity).
Moreover, diagnostic validity refers to the ability to correctly classify affected and
unaffected subjects. The proportion of patients who are actually classified as
demented by a test reflects its sensitivity. The sensitivity of a test decreases as a
function of the proportion of cases of dementia, which the test is unable to classify
(false negatives). The drawback of increasing the sensitivity of a test is the parallel
increase of false positives, i.e., patients erroneously classified as demented.
Therefore, a second index is crucial, i.e., specificity. Specificity reflects the
proportion of non-demented subjects, which are correctly classified by the test. At
present, the majority of diagnostic tests for dementia show very good sensitivity
and specificity in the moderate stage of the disease, but not in the early stage. The
Mini-Mental State Examination (MMSE) (5), for example, shows a sensitivity of
87% and a specificity of 92% for scores of 23–24 (6). Another issue is the need for
tests with high sensitivity and specificity, not only in the very early, even
“preclinical,” and moderate stages of the disease, but also during the late stages
(severe dementia).
Neuropsychological Screening and Advanced Tests 35
Clinical Assessment
The clinical evaluation of mental function is a part of the neurological and
psychiatric examination. It consists of a series of “paper and pencil” tests that are
aimed at investigating the main cognitive areas with only limited supportive
material. The patient’s performances are not measured quantitatively, but on the
basis of the clinical experience of the examiner. There are several published
guides to the neuropsychological examination at the “patient’s bedside” (7–9).
This kind of cognitive evaluation is the only level of assessment that is required
by most diagnostic criteria, such as DSM-IV. It is, however, recommended to
complete the assessment with brief quantitative screening tests. The clinical assess-
ment can be standardized using rating scales, such as the Global Deterioration Scale
(GDS) (10) and Clinical Dementia Rating (CDR) (11), which also include an
assessment of functional status.
Medical history even in mildly affected patients should always include
information from both patient and caregiver. In particular, the assessment of
awareness of cognitive dysfunction might help in the distinction between AD and
MCI. While most MCI patients tend to overestimate cognitive deficits when
compared to their caregiver’s assessment, AD patients in the early stages of
disease underestimate cognitive dysfunction (12). Anosognosia can thus be
regarded as a characteristic symptom at a stage of very mild AD, but not MCI.
Intelligence Tests
Tests that have been developed for the standardized evaluation of intelligence
level in normal subjects have been traditionally used to assess cognitive decline
due to neurological dysfunction. The intelligence quotient (IQ) provided, for
example, by the Wechsler Adult Intelligence Scale (WAIS) battery has been used
to diagnose and assess dementia. The IQ evaluates the multidimensionality of
mental ability and provides a global score that characterizes an individual’s
performance as a whole, as well as a measure of aggregate intelligence that is
composed of elements or abilities that are qualitatively differentiable (13). An
early observation is that discrepancies between some verbal and performance
36 Perani
tasks are evident in normal aging and accentuate in early dementia. Wechsler
himself noticed that some subtests tend to “Hold” (Vocabulary, Information,
Object Assembly, and Picture Completion) opposed to “Don’t Hold” tests (Digit
Span, Similarities, Digit Symbol, and Block Design). On this basis, he proposed a
deterioration quotient (Hold–Don’t Hold/Hold), which has been used as an index
of dementia.
The length of the administration procedures, as well as the development of
tests that are directly related to the cognitive impairments and that are specific to
dementia, has progressively reduced the diffusion of the diagnostic application of
intelligence tests. However, it should be noted that the use of the WAIS subtests
with an emphasis on the quality of the responses rather than on the quantity is a
source of interesting supplementary information for the clinician (14,15). There
are abbreviated versions of the WAIS (16), which continue to be used in AD
patients (17).
Screening Tests
These include a number of tests that can be administered in a limited amount of
time and provide a brief examination of multiple aspects of cognition. Adequate
psychometric information is available for some of these measures, such as
the MMSE.
Other Tests
It must be stressed that even though these screening tests are frequently employed
in clinical practice for the detection and diagnosis of dementia and AD, most of
them lack sensitivity and specificity. Even if the MMSE is the most widely used
by frontline physicians, difficulties with the MMSE in detecting early dementia
have been reported, making it lengthy and potentially cumbersome to use (27).
The main goal for future research in this domain is to provide clinicians with
simple and brief tests, based on solid theoretical and experimental data, with high
sensitivity and specificity.
The Blessed-Roth test, which includes an Information, Memory, and
Orientation test (28), is largely used in dementia assessment.
The Clock Drawing test, widely appreciated for its simplicity and ease of
administration, rendering it a popular tool in both clinical and research practices, is
a brief measure used to detect cognitive decline associated with a variety of
neurobehavioral disorders. It correlates highly with the MMSE and other measures
of global cognitive decline. However, a challenging issue has been identifying the
neuropsychological and neuroanatomic substrates underlying impaired
38 Perani
Neuropsychological Batteries
Several batteries have been designed to test dementia. They comprise
standardized, structured interviews, tests, and examinations for diagnosing
common mental disorders in later life, with special reference to the dementias.
Their principal aim is to incorporate in a single instrument all the
information required to make an accurate clinical diagnosis of dementia.
dementia and for the detection of persons that are likely to develop dementia in
the future.
There is a general agreement (59,60) that measures of delayed recall are the
most effective for the early diagnosis of AD compared to measures of immediate
recall. A non-progressive decrease in delay recall is a frequent finding in normal
aging, but it is also very common in AD, in such a way to render it unusable for
the evaluation of disease progression. The most used tests are learning of a word
list and the “story recall test” logical memory (LM), based on the evaluation of
the encoding of a written piece immediately after its reading by the examiner and
after ten minutes delay with different interferences. The California Verbal
Learning Test (CVLT) (61), for the evaluation of short- and long-term memory in
dementia using local normative values, is widely applied as part of batteries in
multicenter studies (62–64). A deficit in delay recall is typical in the initial phase
of dementia. The performances are indeed severely affected in early mild
dementia. In the non-verbal domain, the recall of Rey’s figure and Corsi’s
supraspan learning are among the most popular measures.
The theoretical principle of “encoding specificity” underlines the strict
dependence between the encoding and retrieval conditions. Indeed, an effective
retrieval requires specific encoding, because “what is perceived determines what
is stored, and what is stored determines what retrieval cues are effective in
providing access to what is stored” (65). It is well known that the encoding
specificity effect is lacking or is very weak even in the early stage of AD (54).
Most tests currently used for the diagnosis of dementia do not take into account
the relationship between encoding and retrieval. One exception is represented by
the Free and Cued Selective Reminding Test (FCSRT) (66). The FCSRT shows
high sensitivity and specificity for the diagnosis of early dementia (57,67).
However, as pointed out by Buschke et al. the FCSRT has two major limitations
(54). First, the power of the test is limited by ceiling effects. These ceiling effects
make important bias in the comparison of the effect of encoding specificity for
aged with dementia and aged without dementia. Buschke and co-workers
developed a test, the Double Memory Test (DMT), to remedy the limitations of
the FCSRT. The DMT includes two memory tests, one with and one without
coordinated acquisition and retrieval. The Category Cued Recall (CCR) memory
test optimizes encoding specificity by using the same category cues for
acquisition as well as retrieval. Buschke and co-workers compared the DMT
with two standard memory tests, Paired Associated (PA) and LM (68), in the
diagnosis of early dementia. They found that the CCR had much higher
sensitivity (93%) and specificity (99%) than PA (68%, 91%) and LM (48%,
92%). CCR had the greatest advantage in the mildest cases. These results show
that tests based on encoding specificity are much more powerful for the diagnosis
of early dementia than traditional learning tests. For the screening of dementia,
Buschke and co-workers have recently devised a test, the Memory Screen Test,
which is a brief, four-item delay free and cued recall memory test (69). This test
Neuropsychological Screening and Advanced Tests 43
showed good sensitivity and specificity for the screening of dementia (70),
providing efficient, reliable, and valid screening for AD and other dementias.
Attention
Attention is also a non-unitary function. Tests that evaluate vigilance and
selective focused attention include the verbal and non-verbal span and letter
cancellation tests. Selective divided attention is measured by the Brown-
Peterson, Trail B, and Stroop tests. The test most widely used to evaluate
selective attention abilities is the “cancellation” test. In this test, of which many
variants exist, the subjects are required to detect as quickly as possible a given
series of stimuli, for example a letter or a digit, among a great number of
distracters. Another widely used test of selective attention is the Stroop test (71).
Here the subjects are exposed to ambiguous stimuli, namely names of colors
written in an incongruous ink color (for example the word BLUE written in red).
In this task, the subjects are required to name as fast as possible the color of the
ink in which each word is written and to ignore the word itself.
Executive Functions
The most widely used tests in clinical practice are verbal fluency, Trail Making
Test (TMT), and Wisconsin Card Sorting Test (WCST). In the verbal fluency
tasks, the subject is required to retrieve as fast as possible words belonging to a
specific category (e.g., animals) or beginning with a given letter. Another widely
used test of executive functions and mental flexibility is the TMT (72). This test is
given in two parts, A and B. The subject must first draw lines to connect
consecutively numbered and lettered circles on one work sheet (Part A), and then
connect the same number of consecutively numbered and lettered circles on
another worksheet by alternating between the two sequences (Part B). The WCST
is probably the most widely used test to assess executive functions, in particular
“abstract behavior” and “shift of set” (73). In this test, the subject is given a pack
of 60 cards on which are printed one to four symbols, triangle, star, cross, or
circle, in red, green, yellow, or blue. The patient’s task is to place them by one
under four-stimulus cards according to a principle that the patient must deduce
from the pattern of the examiner responses to the patient’s placement of the cards.
Language
Language impairments are a prominent feature of AD (74) and FTD. The
mechanisms underlying the language deficits found in persons with AD are often
attributed to the direct result of the cognitive deficits, particularly those related to
memory. Impaired naming is the major change in language functioning in early
AD, and reflects the impairment of semantic memory. Word-finding difficulties
tend to be prominent, and their assessment plays a role in the differential
diagnosis of the dementias, especially AD (75,76). Naming tests such as the
Boston Naming Test (77) are therefore commonly used; as mentioned above it is
also part of the CERAD battery. Normative data for normal aging have recently
44 Perani
Praxis
Another cognitive deficit encountered in all stages of dementia is apraxia. An
evaluation battery proposed by De Renzi and Lucchelli has been used in AD (86).
The authors showed that both ideational apraxia and, to a lesser extent, ideomotor
apraxia (IMA) are present in AD. Apraxia is a crucial feature of corticobasal
degeneration. Spatial orientation and limb praxis are usually spared in FTD (87).
In AD, all aspects of visuoconstruction, including copying, drawing, and
construction in 3D, are impaired. The immediate reproduction of Rey’s Complex
Figure (88) and the Constructional Apraxia test (89) test visuospatial competence
as well as planning and perceptual organization. The WAIS-R block design task is
impaired in AD (90). Another possible source of constructional difficulties is
impaired executive function and global or local attention processes.
Visuospatial Abilities
In AD there are severe impairments of spatial abilities (91). AD patients have
difficulties in scanning the visual field in order to locate objects, and may present
with a deficit in spatial exploration and Balint’s syndrome (gaze apraxia, optic
ataxia, and simultagnosia) (92). These impairments lead to deficits in everyday
life including problems in writing, drawing, driving, following lines of text when
reading, and telling the time from a watch. Some patients, though rarely, may
present with spatial hemineglect. There is a disruption of personal and
extrapersonal orientation: inability to follow unfamiliar route, but also becoming
lost in familiar surroundings. Tests of the ability to undertake the “road map test”
in relation to one’s own body position showed a prevalence deficit in AD (93).
Neuropsychological Screening and Advanced Tests 45
The two aspects of spatial cognition that are most frequently evaluated in AD
are mental rotation and judgement of line orientation. Tests of detection of object
rotation or object rotation to solve object matching problems (94,95) show
significant impairment. The TMT B (96), described above, is also used to measure
the visuomotor planning. Street’s test (89) also measures visual and perceptual
abilities.
Neuropsychiatric Inventory
The Neuropsychiatric Inventory (NPI) scale was developed in order to gather
data on the presence and nature of psychopathological disorders in patients with
cerebral pathologies (100), and addresses particularly patients with AD or other
dementias. It is based on caregiver’s information obtained by a questionnaire and
provides measurements of frequency and severity of behavioral disturbances. The
scale relies on the caregiver and evaluates twelve common psychiatric disorders,
from delusion, hallucinations, agitation, and depression to anxiety, disinhibition,
and apathy. Each single item is correlated to further sub-items that provide more
detailed information. Administration time is about 20 minutes. NPI is a valid
instrument for behavioral evaluation in AD, VaD, and FTD and may also help in
the differential diagnosis of dementia (101).
Assessment of Depression
A particular concern is the evaluation of depression. This is usually achieved with
the application of rating scales, such as the Geriatric Depression Scale, but also the
Hamilton, Beck, and Cornell scales. The latter properly addresses the evaluation
of depressive symptoms in dementia. It is the only scale validated in patient
populations, even at moderate-severe stages. It consists of a series of standardized
items obtained by patient and caregiver’s interviews. It is also based on the
patient’s own observations and does not require direct answers by the patient to a
standardized questionnaire. There are 19 items with graduate scale points from 0
(absence of symptom) to 2 (severe symptom).
FUNCTIONAL SCALES
It should be underlined that all diagnostic criteria require that the cognitive
deficits, assessed clinically and quantified by neuropsychological tests, have an
impact on the activities of daily living. Several functional scales are used with the
aim of evaluating working, social, and relational capacity in patients with
dementia (105). These scales present with a series of specific problems, which is
mainly concerned with the limited information about their validity and
sensitivity. A functional impairment depends also on occupational status and
personal attitudes, and it is additionally dependant on the subjective evaluation
and clinical interview (1). Nevertheless, standardized examinations are largely
used because they can give answers when it is necessary to compare different
phases of the disease, thereby measuring disease progression. They are also
useful for determining the impact of rehabilitative and pharmacological
treatments (106).
More precisely, a functional evaluation is aimed at measuring the
individual capacities in concrete activities and in social roles (107).
dependent or completely dependent when he/she can exert control over all of
these functions, needs assistance or is completely dependent on nurse or
caregiver help. The ADL are widely used but are influenced by the cognitive
status, by the social and frequent usage of the subject, and also by the physical
conditions, such as for the BADL (106). A complete autonomy reflects the
capacity for the subject to live independently at home. In dementia there is a rapid
loss of IADL, and a certain sparing of the BADL, but in the more advanced
stages, all the activities are lost. A major problem, however, mainly concerns the
limited sensitivity in the early phase of dementia, in particular in individuals with
large social and relational interests. To solve this problem the Advanced
Activities of Daily Living (AADL) have been introduced. These represent
complex and demanding aspects of daily living such as travelling, hobbies, and
recreational and social activities (107). The AADL can reveal behavioral
modifications that, while not indicative of a specific cognitive impairment,
indicate the likelihood of incipient dementia.
A serious problem is the floor effect, since the lowest level of these
scales includes individuals with very different degrees of disability. The use
of the Bedford Alzheimer Assessment Nursing Scale, for severe disability,
allows a precise evaluation of the functional level in each individual, with the
possibility of monitoring also the effect of rehabilitative and therapeutic
approaches (109).
These indirect functional evaluations, all based on information provided by
the patient or very often by the caregiver, have some limits linked to variables
that might influence a correct report on the real capacity of the patients. To
overcome this problem, there are more direct evaluations based on objective
observation of functional capacity of the patient to perform a series of
standardized tasks that mimic the basic daily activities. Among these the
Physical Performance Test and the Direct Assessment Functional Scale formerly
used in the elderly have been applied in the evaluation of patients with dementia
(110,111). Finally, it is useful in monitoring of crucial survival abilities, such as
walking capacity, equilibrium, and feeding by standardized instruments (Tinetti
Scale) (112).
the elderly depressed subjects were less impaired than were the mild AD subjects.
Finally, although performance of the moderate AD and Wernicke’s aphasic
subjects did not differ in terms of total CADL score, the performance of these
groups’ subtests was markedly different.
Early Diagnosis
It is well accepted that the natural course of dementia from a condition of full
health to an established dementia status may last several years. At the beginning
the patient might have a deficit limited to memory or to attention (not both),
without any disorder of instrumental and daily activities. The cognitive
impairment then proceeds to a degree that allows the diagnosis of dementia. In
the transitional state between normal aging and mild dementia is fitted the
term MCI.
The term MCI is an operational diagnostic term used to describe subjects at
risk of developing AD or in the pre-clinical stage of the disease (115). It is
reserved for subjects whose impairment is objectively demonstrable but is not
pronounced in more than one domain of cognition and does not seriously affect
activities of daily living. Neuropsychological assessment in MCI subjects shows
impairment in memory tests, especially delayed recall, a possible index of an
Neuropsychological Screening and Advanced Tests 49
subgroups of converters showed low IQ scores and lower verbal memory scores
than non-converters (125). In MCI and very early AD, impairment on tests of
episodic memory is generally the most predictive measure (57,115), but deficits
in semantic memory [semantic knowledge of famous people is also lost in very
early AD (126,127)], attention, and mental speed also predict conversion to AD
(15,128,129). Longitudinal studies revealed the role of a spatial learning test from
the CANTAB computer battery, the PA Learning (PAL) test, which is sensitive in
detecting the earliest stage of AD and is also able to distinguish patients with
depression (130). In the early phase of AD several extensive cognitive batteries
have been designed showing considerable promise (131).
Vascular Dementia
VaD is the second most common type of dementia; however, it is increasingly
being recognised that VaD is actually a heterogeneous syndrome and that several
vascular pathologies can lead to cognitive deterioration (132). In contrast to the
striking deficits produced by cortical infarcts, lesions of the subcortical white
matter are mainly associated with a non-specific slowing of behavior.
Cerebrovascular disease also plays an important role in forms of cognitive
decline other than dementia, and as such, it appears to be no less prevalent in old
age than AD. In addition, AD and VaD represent the extremes of the clinical
spectrum of dementia conditions that includes also various combinations of
cerebrovascular and degenerative pathology. It is thus evident how classical
diagnostic criteria are often of reduced clinical value (133). The relationship
between cerebrovascular lesions, as shown by neuroimaging, and focal
neuropsychological signs is difficult to define during clinical follow-up. There
is a common belief that the cognitive impairment should be ascribed to mixed
vascular and AD types of dementia (134). The prevalence of this kind of
dementia is high in the elderly population. A large multi-center study for the
treatment of CVD and AD showed a slower rate of progression of cognitive
impairment in “pure” VaD in comparison to AD (135) using the ADAS-cog
assessment in a two-year follow-up study.
A specific neuropsychological profile, characterized by a prominent
impairment of executive and visuospatial functions associated with a relative
sparing of episodic memory, has been suggested to be typical of subcortical
VaD (136).
Neuropsychological Screening and Advanced Tests 51
measure of dementia severity, Tower of London Test, WCST for frontal function,
and Raven’s Colored Progressive Matrices (148) for abstract reasoning. Also,
there are the CVLT (61), Story Recall, and Word Learning (149) tests to assess
verbal recall and verbal learning, the recall of Rey’s Complex Figure (88) for
visual long-term memory, and the Digit Span forward and reverse test (150) to
evaluate STM. The pattern of memory decline differs from that of AD: memory
performances benefit from cues and from the provision of multiple-choice
alternative responses. A deficit of retrieval strategies more than a storage problem
has been suggested (84). The TMT A is used to test attention/concentration of the
patients, and the TMT B, which also measures visuomotor planning, is used for
the assessment of divided attention (96). The immediate reproduction of Rey’s
Complex Figure (88) and the Constructional Apraxia test for visuospatial
competence, as well as planning and perceptual organization and the Street’s test
(89) for visual and perceptual abilities, are also used. Language ability should be
tested not only for the classic naming, repetition, reading, and writing
competences, but also for phonological, semantic, and grammatical aspects, as
well as language comprehension (e.g., Token Test) (89). With this aim, the
BADA Italian standardized aphasia protocol (151) has been used in Italy. In FTD
there is an economy of language output, hypophonia, and loss of prosody.
Perseveration, echolalia, and stereotypies occur in the course of the disease.
Mutism usually ensues in the final stages. Patients have no perceptuospatial
difficulty, and skills are preserved even in advanced disease. There is no spatial
disorientation, which is an important distinguishing feature from AD. Due to
reduced cooperation and concentration deficits, it might be difficult to administer
the whole neuropsychological battery to some patients.
Primary progressive aphasia usually presents with insidious and gradual
progression of anomia or word-comprehension impairment. Intact premorbid
language and absence of a “specific” cause, stroke in particular, are also
hallmarks. A non-fluent aphasia is typical. Acalculia and IMA may be present, but
there is an absence of significant forgetfulness and apathy for the initial two years.
ADL impairment, when present, should be ascribed to language deficits.
Semantic dementia presents with anomia, impairment in word comprehen-
sion, and loss of knowledge on semantic tasks (152). A fluent aphasia of the
transcortical type is a hallmark. There might be also reading difficulties (surface
dyslexia) with inability to read irregularly spelled words, but intact syntactic
functions. The Pyramids and Palms test can be used to evaluate the impairment of
semantic knowledge (153).
In a series of comparative studies, patients with the clinical diagnosis of
probabile AD and FTD showed significantly different neuropsychological profiles.
In an object and action naming test, FTD patients were particularly impaired in the
latter (154); during word generation on a phonemic cue (f, p, l as initial letters) or a
semantic cue (animal, fruit names), FTD patients showed major impairment with
the phonemic fluency, whereas AD patients were impaired in semantic fluency.
Neuropsychological Screening and Advanced Tests 53
Severe Dementia
The great increase of the prevalence of age-associated dementias, particularly AD,
has become a major health problem. The majority of cases evolve towards a stage
of marked severity, which can last many years. Severe dementia is accompanied by
loss of autonomy and impairment in activity of daily living ADL, and severe
Neuropsychological Screening and Advanced Tests 55
the spatial perception score. This study confirmed that impairment in visual
perceptual tests requiring access to semantic and lexical knowledge is present in
the earliest phase of AD, whereas visuospatial and constructional impairments
became evident only later. The authors suggest that this pattern of progression
may represent the clinical correlate of increasing pathological involvement of
posterior associative cortex.
There is also a characteristic cognitive profile and course of dementia in
FTD. Nonetheless, cognitive test performance does not clearly distinguish FTD
from AD in the early phase. A cross-sectional and longitudinal study was
specifically instituted to address the cognitive characteristics of FTD, with the
aim of determining which features distinguish FTD from AD. A large series of
patients were included, some with a pathologically verified diagnosis of Pick’s
disease. Information regarding the initial symptoms of dementia were obtained
from each patient’s caregiver, global dementia severity was estimated by the
BDS and the ADL scales, and specific cognitive domains were assessed by
administering tests of memory, language, visuospatial, and reasoning abilities
and selective attention. Among initial symptoms reported by caregivers,
personality change and language impairment were significantly more common
in FTD than AD. At initial cognitive testing, deficits in memory were common in
both groups, but more prevalent in AD. Patients with FTD had greater
impairments on the ADL Scale. During the course of illness, patients with FTD
declined significantly faster than those with AD on language tests and on global
measures of dementia severity (187). These results in this large co-hort of
patients with dementia underline the importance of determining cognitive
profiles at the beginning and during the course of the disease, for the
differential diagnosis.
The same is supported by another study that suggests a selection of
neuropsychological tests are sensitive in the differential diagnosis of AD (155).
Neuropsychological performance (including measures of language, semantic
memory, visual and spatial perception, and executive functions) was compared in
two groups of patients with clinical diagnosis of probable AD or FTD. The aim was
to identify a specific cognitive profile for FTD to be used as a sensitive short
evaluation for the differential diagnosis with AD. Both groups were severely
impaired in most tasks, but interestingly also in “frontal lobe” tests that have been
suggested to play an important role in the differential diagnosis. Noteworthy,
significant differences were found for a minority of tests that assessed oral praxis,
visuospatial perception, and verbal fluency. The authors suggest a shortened testing
procedure based on four tests (Rey Complex Figure recall, phonemic fluency, cube
analysis, and oral apraxia test), together with other measures sensitive to frontal
impairment as a useful tool in the differential diagnosis between AD and FTD.
New research approaches in the neuropsychological evaluation of dementia
might derive from neuroimaging studies. Normal subjects executing specific
cognitive tasks during functional magnetic resonance imaging (fMRI) or positron
emission tomography (PET) acquisitions have shown the activation of complex
Neuropsychological Screening and Advanced Tests 59
neural systems. These studies provide evidence for the neural correlates of
cognition that go far beyond classical neuropsychological correlates in brain-
damaged patients. For example, they suggest that whereas the left temporal
neocortex plays a crucial role in all tasks involving lexical-semantic processing,
some regions of the left prefrontal convexity are selectively recruited during verb
processing. Repetitive transcranial magnetic stimulation (rTMS) also showed
different neural correlates for noun and verb processing in the human brain (188).
In fact, a shortening of naming latency for actions was observed only after
stimulation of left prefrontal cortex. On this basis, noun and verb processing in
different dementia types were assessed (154). Object and action naming was
tested in probable AD patients with mild to moderate dementia and in a group of
FTD patients. AD and FTD patients were impaired in naming compared with
control subjects; action naming being more severely impaired. However, the
discrepancy between object and action naming was significantly greater in FTD
than in AD patients, independent of the severity of dementia or of overall
language impairment. The latter finding is compatible with the hypothesis that
the frontal lobe plays a crucial role in action naming. The authors suggest a
relatively selective impairment in action naming as a characteristic neuropsy-
chological feature of FTD.
New interesting research approaches aimed at identifying specific
personality and behavioral profiles in dementia have recently emerged. In
FTD, diagnostic criteria agree that alterations in personality and social conduct
are a central clinical feature of the disease (4,137). However, quantitative
instruments have not been used to systematically measure these changes.
Similarly, specific social deficits in the FTD syndrome have not been isolated to
either the frontal or temporal variants of the disease. A promising approach is the
study of “theory of mind,” a key aspect of social cognition related to the
prefrontal cortex, which has been reported to be affected in the frontal variant of
FTD (189). One source of confusion may be that clinicians and researchers have
focused on discrete behavioral changes (138,142,190), since they do not have the
tools to observe a patient’s social processing on a higher level. Since personality
becomes fixed by early adulthood and remains constant throughout old age,
significant changes in personality during adulthood typically have a neuropatho-
logic etiology (191). Because the primary brain areas affected in FTD are the
frontal lobes, in particular the orbitofrontal and frontal medial cortex, the anterior
temporal lobes, the amygdala, and ventral striatum (63), the observed personality
changes ostensibly arise from a disruption of these structures. In contrast, early in
the disease, social functioning remains preserved in AD, most likely because of
the relative sparing of these anterior structures (192,193). It therefore seems
important to include investigations of how particular social deficits correlate with
neuropsychological functioning and areas of pathology on functional brain scans.
It will become possible to further characterize a patient’s personality change as a
function of the timing of the disease progression. A recent paper suggests the
Interpersonal Adjectives Scales, since they were able to differentiate frontal and
60 Perani
temporal variants of FTD from patients with AD, who remained within the
normal range on all scores, on the basis of both degree and direction of
personality change (194,195). On a similar basis, the Frontal Behavioral
Inventory Scale is a useful tool for the assessment of behavioral changes in
FTD patients.
The identification of clinical subtypes of probable AD, such as the so-called
posterior cortical atrophy variant, raises the issue of a more focused
neuropsychological assessment. Posterior cortical atrophy presents with prevalent
visuospatial deficits, thus evaluation of visuoperceptual abilities and also praxis is
necessary. Two batteries have been proposed so far, the Visual Object and Space
Perception Battery and the Birmingham Object Recognition Battery. They are
both time-demanding, and therefore considering the patient’s compliance, a
useful evaluation can be achieved by a using a shorter task administration, such as
Benton’s Line Judgment Orientation and Benton’s Face Test.
CONCLUSIONS
Not long ago, cognitive aging was viewed as a general deterioration of cognition
(reflected by a decreasing IQ) and dementia as an acceleration of this process.
Currently, we know that cognitive decline occurs in various forms, marked by
considerable differences in the nature and order of development of behavioral
symptoms. This knowledge is an important asset in the early detection,
management, and treatment of dementia. The neuropsychological assessment
of dementia has a central role and is concerned with all disease stages. The
current implicit assumption that all patients with AD tend to evolve and decline in
a similar fashion needs to be critically re-examined. Neuropsychological tests
allow one to determine various patients’ profiles. Future research should
determine the possible predictive value of these profiles, which has important
implications for therapeutic trials. Clinically, neuropsychological instruments are
crucial for the diagnosis, prognosis, and monitoring of pharmacological
treatments. In the field of research, its role is even more clear-cut: a correct
clinical diagnosis is mandatory in instrumental and pharmacological trials.
However, the neuropsychological approach to dementia certainly has limitations
and is at its proper place only in a multidisciplinary context. The concerted effort
of all branches of neuroscience is required for patient care, as well as research on
cognitive decline in aging individuals and in neurological diseases.
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3
Biomarkers in Blood and
Cerebrospinal Fluid
INTRODUCTION
Alzheimer’s disease (AD) is the most common neurodegenerative disorder and
afflicts about 10% of the population over 60. Biomarker research in neuro-
degenerative disorders using blood and cerebrospinal fluid (CSF) is focused on
specific targets: early detection, differential diagnosis (classification), tracking of
disease progression, and evaluation of therapeutic strategies. With the currently
approved antidementia drugs and with the development of novel disease
modifying therapeutic strategies (secondary prevention), it is of particular
clinical interest to establish early diagnostic and preclinical prognostic
biomarkers of AD.
Criteria for a useful biomarker were proposed by an international consensus
group on molecular and biochemical markers of AD in 1998 (1). According to
these guidelines, a biomarker for AD should detect important aspects of the
fundamental neuropathology and be validated in neuropathologically confirmed
cases. Its sensitivity for detecting AD should exceed 85% and its specificity in
differentiating between AD and other dementias should be at least 75%. Ideally, a
biomarker should also be reliable, reproducible, non-invasive, simple to perform,
and inexpensive.
73
74 Hampel and Buerger
Ab in Cerebrospinal Fluid
Initial reports on Ab in CSF as a biomarker for AD were disappointing, with
results ranging form a slight decrease in AD, with a large overlap, to no change
(7–9). In these studies, total Ab in CSF was examined. More recent studies have
shown two major C-terminal variants of Ab, with either 40 (Ab40) or 42 (Ab42)
amino acids. Different ELISA tests have been developed for the measurement of
Ab42 in CSF and plasma (10,11). There are commercially available ELISA-kits
for the measurement of Ab42 and Ab40 in CSF (e.g., The Genetics, Schlieren,
Switzerland; Innogenetics, Gent, Belgium). Several studies reported that Ab42
concentrations are decreased in CSF of AD patients (12–17). The specificity to
distinguish patients with AD from controls has varied from 42% to 88%, and the
Biomarkers in Blood and Cerebrospinal Fluid 75
sensitivity has varied from 72% to 100% in these studies. CSF levels for Ab42
seem to be unrelated to age. Based on recent data, a cutoff-level of O500 pg/ml
has been suggested to discriminate AD from normal aging (18). Further, it has
been reported that Ab42 levels were lower in AD patients with the APOE 34
allele than those without APOE 34 allele (19). The sensitivity of Ab42
measurement was 83.6% for AD patients carrying an APOE 34 allele, whereas in
AD patients without APOE 34 allele it was 54.2%. Decreased CSF Ab42 levels
are not specific for AD since studies showed that half of the patients with vascular
dementia (VaD) had decreased CSF Ab42 (16).
It has been hypothesized that a decrease of Ab42 in CSF indicates an early
stage of AD before clinically overt dementia is detectable. A significant decrease
of CSF Ab42 in MCI subjects compared to controls has been shown, but this
study had no follow-up measure (20). In a further study investigating MCI
patients, Ab42 levels did not differ significantly from age-matched normal
controls (21). Control subjects in this study, however, were subjects with memory
complaints without neuropsychological impairment. Since it has been reported
that individuals with memory complaints have a higher risk of dementia (22–24),
the control group in this study might represent a subgroup with subclinical AD
pathology reflected in altered CSF Ab42 levels. More recently, it has been shown
that Ab42 protein may be an indicator of early identification of AD in MCI
subjects when taking potential confounding factors into account such as age,
severity of cognitive decline, time of observation, APOE E 34 (APOE 34) carrier
status, and gender (25).
Studies correlating CSF Ab42 protein concentrations with cognitive
performance in AD have been contradictory. Cross-sectionally, the concentration
of Ab42 protein and cognitive measures were either inversely correlated (26–28)
or no significant correlation was found (12,15,16,29). In a longitudinal study, a
decrease in CSF Ab42 protein has been found within a three year follow-up (30).
A highly significant correlation between low CSF concentrations at baseline and
one year serial follow-up was demonstrated. In a separate study, no correlation
was found between CSF levels and duration or severity of AD (12). The potential
value of Ab42 during the course of AD progression should be further evaluated.
Several studies have been performed on the potential of CSF Ab42 to
differentiate AD from other neurodegenerative disorders. Compared to control
subjects with other neurological conditions, a slight decrease has been described
in non-AD dementias (31). Normal or decreased levels of Ab42 were reported in
Parkinson’s disease (PD) (11,32). In Lewy body dementia, a disorder also
characterized by the presence of senile plaques, low levels of Ab42 protein have
been detected. The range of Ab42 protein concentrations found in Lewy body
dementia overlaps with Ab42 concentrations found in AD patients (20,33–35).
Furthermore, low CSF Ab42 protein is found in a relatively large percentage of
patients with frontotemporal dementia (FTD) and VaD (16,36). In summary, CSF
Ab42 does not seem to significantly support the differential diagnosis of AD.
76 Hampel and Buerger
A satisfying explanation for the decrease of the Ab42 level in CSF to about
40–50% of control levels in AD (14) is still lacking. One suggested mechanism is
that it is caused by the deposition of Ab42 in plaques, with lower amounts of Ab
being free to diffuse into CSF (11). This explanation is also supported by the
finding of a strong correlation between low Ab42 in ventricular CSF and high
numbers of plaques in the neocortex and hippocampus (37). However,
subsequent studies also found a marked reduction in CSF Ab42 in disorders
without b-amyloid plaques, such as CJD (38), amyotrophic lateral sclerosis (39),
and multiple system atrophy (40). This points to other mechanisms, such as
disturbances in Ab formation and breakdown in AD. Interestingly, one study
reported a significant increase of Ab42 protein in patients with early stage of AD
followed by a steady decrease concentration (10). This might be explained by
different sets of antibodies and protocols used, as well as by differences in the
stage of the disease when the patients were included.
Several CSF studies have shown that concentrations of another amyloid
b peptide species, Ab40, were similar in AD and controls (11,17,41,42). In one
study (43), however, a decrease in CSF Ab40 values was found with significant
overlap between the groups. Another study reported that Ab40 is deposited later
in the disease and is prominent in vascular amyloid deposits (44).
In addition to Ab42 and Ab40 in CSF alone, the ratio of Ab42 to Ab40 has
been investigated. The ratio of Ab42 to Ab40 is suggested to be superior to the
concentration of Ab42 alone in discriminating patients with AD from normal
controls (42). Furthermore, a recent study showed that for the neurochemical
diagnosis of AD, the number of correctly classified patients turned out to be
slightly higher compared to all groups (non-Alzheimer dementia and control
subjects) when the Ab peptide ratio was used instead of the Ab42 concentration
alone (correct discrimination of AD and controls increases from 86.7% to 94%
when Ab42 is replaced with Ab peptide ratio); however, this effect failed to reach
significance (17).
Ab in Plasma
Some studies suggest that plasma Ab40 and Ab42 levels were two- to threefold
higher in patients with familial AD and with presenilin mutations than in subjects
with sporadic AD and controls (45). Further, it has been reported that Ab levels are
approximately 100-fold lower in plasma than CSF (45). Some studies
demonstrated that plasma Ab40 and Ab42 levels are similar in AD and control
groups (46,47). However, others have shown that plasma Ab40 levels are
increased in AD patients with APOE 34 allele compared to those without this allele
and age-matched controls (46). Because of the potential overlap between the AD
patients and controls, measurement of plasma Ab40 levels is not useful as a
diagnostic tool to distinguish patients with sporadic AD from elderly non-
demented (ND) controls (46,47). A longitudinal study of unrelated individuals
reported that those who subsequently developed AD had higher plasma Ab42
levels at entry than those who did not develop dementia (48). These results indicate
Biomarkers in Blood and Cerebrospinal Fluid 77
that elevated plasma Ab42 levels may be detected several years before onset of
symptoms, supporting the role of extracellular Ab42 in the pathogenesis of AD.
However, although blood is easy to obtain, it is still unclear if there are
systemic changes specific for AD, and to what extent changes in blood
composition reflect pathological changes seen in the brain since plasma Ab42
levels showed no difference between AD and controls, whereas data with Ab40
are controversial (with either an increase in AD or no change). One longitudinal
study has shown that elevated plasma Ab42 levels occur before the onset of MCI
in some individuals (48).
In summary, Ab1–42 comes close to fulfilling the criteria for a useful AD
diagnostic test as recently summarized by an expert review (49). It is, however, of
limited value in differentiating AD from other primary dementias. Further studies
will be required to evaluate the whether the ratio of Ab42 to Ab40 is superior to
the concentration of Ab42 alone (17).
TAU PROTEINS
Neurofibrillary tangles are one of the major pathological hallmarks of AD. They
consist of paired helical filaments (PHF), derived from abnormally hyper-
phosphorylated microtubule-associated protein tau (59). Physiologically, tau is
located in the neuronal axons and a component of the cytoskeleton and
intracellular transport systems. Due to alternative splicing of tau mRNA, there
are six isoforms ranging in size from 352 to 441 amino acids, with molecular
weights ranging from 50 to 65 kDa (60), encoded by a single gene consisting of
16 exons on chromosome 17q21. Furthermore, at least 30 phosphorylation sites,
either threonine or serine, exist on tau extracted from human brain (61). Due to
hyperphosphorylation, tau loses its ability to bind to the microtubules and to
stimulate their assembly, and shows an increased tendency to aggregate (62).
Total tau (t-tau) and truncated forms of monomeric and phosphorylated tau are
released and can be measured in the CSF.
The first promising report on CSF t-tau as a biomarker for AD was
published in 1993. An ELISA with a polyclonal reporter antibody was used (63).
After this, ELISA methods based on monoclonal antibodies have been developed
that detect all isoforms of tau independent of phosphorylation sites (64,65) to
measure total and phosphorylated CSF tau protein (66–68).
Total Tau
T-tau, a general marker of neuronal destruction, has been intensively studied in
more than 30 studies on 2000 AD patients and 1000 age-matched elderly controls
over the last 5–10 years (69). The most consistent finding is a statistically
significant increase in CSF t-tau protein in AD. The mean level of CSF t-tau
protein concentration is about 300% higher in AD compared to elderly controls.
Across the reviewed studies, sensitivity and specificity levels varied due to the
different control groups and statistical methods used. Specificity levels were
between 65% and 86% and sensitivity between 40% and 86% (14). In several
studies, a significant elevation was also found in patients with very early
dementia (31,70,71). Overall, in mild dementia, the potential of CSF t-tau protein
to discriminate between AD and normal aging is high, with a mean sensitivity of
75% and a specificity of 85% (69). An age-associated increase of t-tau protein has
been shown in ND subjects (72). Therefore, the effect of age should be considered
when t-tau protein levels are diagnostically employed. Age-dependent reference
values for t-tau protein have already been established: for subjects between
21–50 years old at !300 pg/ml, between 51–70 years old at !450 pg/ml, and
between 70–93 years old at !500 pg/ml (18).
MCI is a major risk factor for AD. Ten to fifteen percent of patients with
MCI have been reported to convert to AD in a year (73). In patients suffering
from MCI who converted to AD during follow-up, elevated t-tau levels at
baseline were found in a relatively high number of individuals (20,74). Memory
impaired subjects who later developed AD could be discriminated by high
Biomarkers in Blood and Cerebrospinal Fluid 79
CSF t-tau from those who did not progress with 90% sensitivity and 100%
specificity (75). Longitudinally, elevated CSF levels of t-tau in MCI subjects
were found and still remained elevated after conversion to clinical AD. Another
study showed that 88% of patients with MCI had elevated t-tau concentrations
and/or low CSF Ab1–42 levels at baseline (13). Thus, elevated CSF t-tau in MCI
may have the potential to predict AD, a finding that was supported by a recent
study (2). Cross-sectional studies correlating CSF t-tau concentrations with
cognitive status in AD have shown a correlation between elevation of t-tau and
cognitive decline (26,27,76), though others have found no systematic effect
(16,77–80). Longitudinal studies of t-tau in mildly to moderately demented AD
patients showed no statistically significant correlation with progression during
follow-up (12,81,82). CSF t-tau remained elevated for up to two years in mild to
moderate AD. Initial and follow-up levels of t-tau correlated strongly, suggesting
a stable rate of neurodegeneration during this time period. It could be
hypothesized that CSF t-tau will decrease over time if treatment of AD achieves
disease-modification and neuroprotection (83).
An increase of CSF t-tau has also been found in a proportion of cases with
other dementia disorders. In VaD, FTD, and dementia with Lewy bodies (DLB),
elevated CSF t-tau has been found (20,33,69,74,78,84–88). Other studies,
however, found normal levels compared to controls in these disorders
(14,16,36,81,89,90). The potential of CSF t-tau, however, is limited in its ability
to discriminate AD from other relevant dementia disorders. At a sensitivity level
of 81%, CSF t-tau reached a specificity level of only 57% in distinguishing AD
from other dementias (16,91). Therefore, t-tau has not been suggested as a marker
for the differential diagnosis of AD.
T-tau rather reflects non-specific processes of axonal damage and neuronal
degeneration. This notion is further supported by an increase in CSF t-tau in
disorders with extensive and/or rapid neuronal degeneration such as CJD (92,93).
A highly significant increase of 580% was documented in CJD compared to AD
patients. At a cut-off level of 2130 pg/ml, t-tau yielded a sensitivity of 93% and
a specificity of 100% between AD and CJD (94). An elevation of CSF t-tau,
correlating with clinical severity, has been shown in normal pressure
hydrocephalus (95). Moreover, a marked transient increase of CSF t-tau has
been demonstrated after acute stroke. The transient increase of CSF t-tau
correlated with infarct size measured by cranial computed tomography (96).
Elevated levels of CSF t-tau in patients with diffuse axonal damage after
traumatic brain injury have been found, thus decreasing with clinical
improvement (68).
The differential diagnoses of AD, alcoholic dementia, PD, progressive
supranuclear palsy, corticobasal degeneration, and other psychiatric disorders
show normal CSF t-tau (20,36,64,97,98), with elevated CSF t-tau concentrations
only occasionally being reported (14,36,74,99,100).
In geriatric major depression (MD), an important psychiatric condition
in the differential diagnosis of AD, CSF t-tau has also been investigated.
80 Hampel and Buerger
Phosphorylated Tau
Promising efforts are under way to establish phosphorylated tau (p-tau) in CSF as a
putative biological marker for AD. Several ELISA methods have been developed
specifically detecting phosphorylation of tau protein at different epitopes, such
as threonine 181 and 231 (p-tau181C231) (66), threonine 181 (p-tau181) (67),
threonine 231 and serine 235 (p-tau231C235) (102), serine 199 (p-tau199)
(102), threonine 231 (p-tau231) (103), and serine 396 and 404 (p-tau396C404) (104).
Although there is no doubt that tau phosphorylation differs in AD, it is hard
to speculate why this should be the case. There have been few studies of p-tau199
and p-tau181 in the human brain. With the exception of one study (105) all that is
known about these sites is that they are phosphorylated in advanced AD
neuropathological changes (106). Furthermore, it is well established that p-tau231
appears early in the pathological development of the disease, even before the
formation of PHF in neurons of the hippocampus (105,107). Phosphorylation at
both threonine 181 and serine 199 occurs later, and these are only found to any
appreciable extent in intracellular tangles (105).
One question, yet to be resolved, is which site of abnormal phosphorylation
is most useful for the differentiation between AD and other disease groups.
A comparative study examining the different tests (p-tau181 vs. p-tau231 vs.
p-tau199) in the same subjects and controls have shown that, applied as single
markers, p-tau231 and p-tau181 reached specificity levels O75% between AD and
the combined non-AD group when sensitivity was set at R85%. With respect to
these data and with respect to the fact that the majority of data on CSF p-tau in
AD is available for p-tau181 and p-tau231, studies on p-tau181 and p-tau231 will
now be described in more detail. Moreover, there is a commercially available
assay for p-tau181, and a commercial assay for p-tau231 will soon become
available.
CSF p-tau231 distinguished between AD-patients and subjects with other
neurological disorders (OND) with a sensitivity of 85% and a specificity of 97%
showing elevated CSF p-tau231 in AD (103). In this first study describing the
assay, a total of 39 CSF samples were prepared and analyzed from individuals
with AD, a number of different dementias, other neurological conditions, and
controls. The data suggest that the assay can distinguish AD from other
dementias (103). In a subsequent study on differential diagnosis of AD using CSF
p-tau231 in an independent sample, a sensitivity level of 90.2% and a specificity
Biomarkers in Blood and Cerebrospinal Fluid 81
APOLIPOPROTEIN E
Apolipoprotein E (APOE, gene; ApoE, protein) is the major apolipoprotein in the
central nervous system where it is involved in the mobilisation and redistribution
of cholesterol, necessary for the maintenance of myelin and neuronal membranes
during development and following injury (115). However, APOE is of special
interest in AD research since the presence of the APOE 34 allele has been
suggested to be a major risk factor for the development of late-onset AD (116).
APOE polymorphism influences levels of amyloid deposits in the brain of AD
patients and elderly ND controls (117).
Previous studies on CSF APOE in AD patients and controls, however, have
shown conflicting results. While in some investigations elevated CSF APOE
levels have been found (118,119), others have reported reduced APOE
concentrations in CSF (120). Interestingly, in one study APOE levels in CSF
were reduced in AD patients without the APOE 34 allele, but increased in AD
patients and APOE 3 4 allele carriers (120). However, another study reported
decreased CSF APOE concentrations in APOE 3 4 allele carriers (118,121);
Lindh and colleagues also reported increased levels of APOE in CSF in patients
with MCI and other dementia disorders than AD compared to healthy controls
(118). To investigate whether APOE levels in CSF differ with various ages at
onset of AD, one study examined APOE in early- and late-onset AD, showing
significantly lower APOE levels in early-onset AD patients and higher in the late-
onset group compared to controls (122).
Concerning serum ApoE in AD patients in comparison to controls, studies
are still inconclusive. Some studies report decreased serum ApoE levels (123),
whereas others have observed similar or elevated serum ApoE levels between AD
and controls (119,124).
Taken together, levels of ApoE in CSF and serum seem not to fulfill the
criteria for a useful biomarker of AD.
Biomarkers in Blood and Cerebrospinal Fluid 83
ISOPROSTANES
Several studies have suggested a role for oxidative damage in the pathogenesis of
different neurodegenerative diseases, especially AD and amyotrophic lateral
sclerosis (ALS) (125–127). Oxidative damage to CNS tissue prominently manifests
as lipid peroxidation (LPO).
Isoprostanes are prostaglandin isomers that are producted exclusively from
free-radical-catalyzed peroxidation of arachidonic acid (128). Isoprostanes are
biochemically stable end-products of LPO that are released by phospholipases,
circulate in plasma, and are excreted in urine (129). Therefore, analysis of LPO by
measuring isoprostane levels has been performed in brain, CSF, serum, and urine
(130–133). Special attention has been focussed on isomers of the F2-isoprostanes
(F2a-iPs), especially 8,12-iso-iPF2a-VI. In different studies, it has been reported
that 8,12-iso-iPF2-VI levels are elevated in urine, blood, and CSF of AD patients
and that these values correlate with memory impairment, CSF tau levels, and the
number of APOE 4 alleles (130–132,134). The results, however, for blood and
urine, in contrast to CSF, have been conflicting since other studies found no
elevated F2-isoprostanes levels (135,136). In addition to AD, in early
Huntington’s disease, meningoencephalitis, and stroke, elevated levels of
isoprostanes were also found (137,138), while in ALS, PD, and schizophrenia
no increase of isoprostanes has been observed (139,140). Furthermore, CSF
F2-isoprostanes concentrations in AD patients are significantly correlated with
global indices of brain degeneration such as decreasing brain weight and degree of
cerebral cortical atrophy, but not with APOE genotype or the tissue density of
neuritic plaques or neurofibrillary tangles (140).
In summary, although additional studies are needed to confirm and extend
these findings in larger cohorts of MCI and AD patients, F2-isoprostanes seem to
be an interesting marker in AD, not only in CSF, but also in serum and urine.
a1-ANTICHYMOTRYPSIN
It has been suggested that a number of molecules associated with inflammation
are involved in the pathogenesis of AD. Antichymotrypsin (ACT), one of the
serine proteinase inhibitors, plays an important role in inflammation.
Interestingly, elevated levels of ACT were found in the brains of patients with
AD and were also described as one of the components of senile plaques (141).
ACT has been investigated in CSF and serum in several studies, but the
results have been controversial. While in some studies no difference in ACT
levels between AD and controls have been found (142), others have described
higher levels of ACT in AD than controls (143–145). A recently performed study
found that ACT levels correlate with the severity of dementia (146). Further
studies are essential to confirm these findings. In addition, investigations on MCI
are needed since ACT has not been studied in MCI so far.
84 Hampel and Buerger
t-tau protein and sIL-6RC components may add more certainty to the diagnosis of
AD (147). The reported method, however, needs to be extended to an
independent group of AD patients, other neurodegenerative conditions, and
control subjects to assess the true diagnostic applicability. Interpretation of the
relationship between CSF and brain levels of the IL-6RC at present remain
speculative and require studies based on simultaneous measurement of
corresponding CSF and brain samples.
C-REACTIVE PROTEIN
C-reactive protein (CRP) is a pentraxin acute phase reactant synthesized in the
liver. Its plasma level can increase up to 1000-fold during the acute-phase
response (149). CRP is not normally found in the brain, but previous studies have
demonstrated the presence of CRP in senile plaques and neurofibrillary tangles in
the brains of AD patients (150,151). It is up-regulated in AD brains compared to
samples from ND individuals (152). In a follow-up study, examining over 1000
cases, it has been observed that men in the upper three quartiles for serum high-
sensitivity CRP had a 3-fold, significantly increased risk for all dementias
combined, AD, and VaD (153). Remarkably, the serum samples that were
investigated had been taken and stored 20–25 years earlier, long before onset of
dementia symptoms in any subject. In contrast, another recent study of only 11
AD and 11 ND patients found comparable levels of CRP in AD and ND patients.
In summary, CRP might play a causal role, or merely be a marker of
inflammatory processes. Whether the association proves to be direct or indirect,
CRP measurements may turn out to be an important adjunct for global risk
assessment of dementia (153).
C1Q
C1q, a subcomponent of C1, the first component of the classical complement
pathway, is associated with neuritic plaques and with neurons in the hippocampus
of AD brain.
There is evidence that C1q, in addition to its normal production in the liver,
is also synthesized in the brain by pyramidal neurons and glial cells (154–156).
b-pleated, fibrillar Ab and, more recently, tau-containing neurofibrillary tangles
have been found to directly and fully activate the classical complement pathway
in vitro in an antibody-independent fashion, (157,158). Of the many different
inflammatory mediators that enhance Ab aggregation, C1q is one of the most
potent (159).
In several studies, abundantly elevated C1q levels have been observed in
the brains of AD patients compared to unaffected brains (160,161). In CSF,
significantly lower levels of C1q have been observed in AD compared to control
patients, thus correlating with cognitive deficits (162). In summary, these results
86 Hampel and Buerger
HOMOCYSTEINE
Homocysteine is a precursor of methionine and cysteine. Folate and vitamin B12
are needed for the conversion of homocysteine to methionine, and vitamin B6 is
essential for the conversion of homocysteine to cysteine (164). Deficiencies of
folate or vitamin B12, and vitamin B6, result in increased levels of homocysteine.
There are a variety of assays to measure homocysteine levels, including immuno-
assays and high pressure liquid chromatography (HPLC)-based methods, which
have similar performance and high precision (165,166).
Hyperhomocysteinemia is considered a potentially important risk factor
for heart disease, carotid atherosclerosis, and stroke (167–170). Atherosclerosis
and stroke, in turn, increase the risk for AD (171,172). However, in AD, VaD,
and cognitive deterioration, increased levels of homocysteine in serum in
combination with decreased vitamin B12 or folic acid are considered as
potential risk factors for the development of cognitive impairment (173). A
significant increase in total homocysteine levels were demonstrated in a large
case control study of 164 patients with AD compared to 108 controls (174).
Total homocysteine levels were stable over a three-year follow-up period, and
homocysteine levels did not correlate with duration of symptoms. A recent
longitudinal study has shown an association between hyperhomocysteinemia
and a higher risk of AD (175). In this study, plasma homocysteine levels greater
than 14 umol/L almost doubled the risk of AD (175). In another study it has
been demonstrated that, independent of low folate levels, higher levels of
homocysteine were associated with cognitive decline in a large group of older
subjects (176). In contrast, no relation between increased homocysteine
concentrations and cognitive decline was observed in a large study with 702
individuals with a mean follow-up duration of 2.7 years (177). A recent MRI
study suggested that increased homocysteine was a risk factor for cerebro-
vascular disease independent of AD (178). This leads to the question of whether
homocysteine is directly linked to mechanisms of AD or indirectly, via
cerebrovascular disease.
Besides homocysteinemia, low vitamin B12 and folate levels were also
found in AD. Since supplementation with B group vitamins lowers homocysteine
levels by up to 30% (174), dietary interventions are now being tried in current
studies to explore the benefit on cognitive outcome (4). Furthermore, in the US,
fortification of cereal grain products with folic acid was mandated in the late 1990s,
and there has already been a decrease in mean values of homocysteine in older
individuals. This will make it more difficult to evaluate whether further
supplementation has benefits in dementia (4).
Studies of folate supplementation in depression, one of several neuropsy-
chiatric diseases also associated with low folate serum levels, have shown benefits
Biomarkers in Blood and Cerebrospinal Fluid 87
even when folate status was not low (179). A potential explanation might be that
serum folate concentrations do not reflect concentrations in the CNS or that folate
requirements are increased in neuropsychiatric diseases (180).
In summary, elevated homocysteine levels in plasma seem to be a strong,
independent risk factor for the development of dementia and AD. Further large-
scale studies are currently in progress.
3-NITROTYROSINE
In the presence of reactive oxygen species and nitric oxide, 3-nitrotyrosine (3NT)
may be formed in constituent proteins in the brain. The predominant pathway
appears to involve nitric oxide in the presence of superoxide ion to form
peroxynitrite. Peroxynitrite in turn reacts with tyrosine residues in proteins or
with free tyrosine to form 3NT (201). 3NT can be found in CSF in specific
proteins, e.g., superoxide dismutase (202), or can be found as free 3NT. With
normal aging, 3NT concentrations in CSF increase modestly from about 0.75 nM
at 40 years to about 2 nM at 80 years (203).
In the brains of patients with AD, regionally specific increases in 3NT have
been found in the neocortex and cerebellum (204). In the CSF of patients with
AD, the concentration of 3NT is reported (203) to be 11.4 nM, or about sixfold
higher than age-matched controls. In this study, concentrations of 3NT in CSF
were inversely correlated with MMSE scores, but did not correlate with duration
of disease.
Analysis of 3NT is relatively straightforward when done by HDL with
electrochemical detection (HPLC/ED) (201). CSF analysis requires only 400 ul,
and acid precipitation of proteins is sufficient for the analysis of free 3NT
(201,203). The stability of 3NT in plasma proteins, however, is not well
established, and thus the significance of 3NT concentrations in this compartment
is less clear (201,205). Increased concentrations of 3NT in CSF, along with
changes in isoprostanes, and 8OHDG as outlined in this review, are consistent
with the suggestion that oxidative stress may play an important role in the
Biomarkers in Blood and Cerebrospinal Fluid 89
pathogenesis of AD. It is not yet clear whether the increase in reactive oxygen
species with oxidative stress is proximal or distal in the pathophysiological
cascade leading to cell death. As with a number of proposed biomarkers for AD,
longitudinal studies are necessary to determine if changes in 3NT in CSF increase
monotonically with disease progression, or might have a more complex
relationship with disease severity. Even in the absence of a linear relationship
with disease severity, measurement of 3NT and other markers of oxidative stress
may provide an indirect marker of drug efficacy in subacute clinical trials using
putative disease-modifying agents.
and APOE 34 carrier status. Ab42 antibody levels were correlated with gender
only in AD, with a higher level occurring in women. When Ab42 antibody
sensitivity (specificity) was set at O80%, specificity (sensitivity) was below 50%
in correctly allocating patients and healthy controls. These data indicate a
potentially pathophysiological decrease of serum Ab42-antibodies in AD. Ab42-
antibodies in the serum alone, however, appear not to be useful as a diagnostic
marker of AD.
So far, very little is known about Ab-antibodies with respect to their
function, induction, specificity, and role in disease processes. However,
considering data from recent investigations in transgenic AD mouse models,
there is evidence for a therapeutic impact of Ab-antibodies. A reduction of
cerebral plaque load and cognitive impairment was observed after active and
passive immunization resulting in increased production or administration of
Ab-antibodies in these animal models (214–220). Unfortunately, the phase II
clinical trial of the Ab vaccination approach had to be withdrawn because of
signs consistent with meningo-encephalitis in an increasing number of patients
(221). Setbacks in active immunization are shifting the focus to passive
administration of antibodies. Dodel and colleagues detected naturally occurring
Ab-antibodies in commercially available immunoglobulin G products. In a
clinical approach they investigated patients with different neurological diseases
treated with intravenous immunoglobulin (IVIG) preparations. A significant
decrease of total Ab and Ab1–42 in CSF compared to baseline values was
observed after treatment. In serum, a significant increase of total Ab without
change in the Ab1–42 level was observed (222). These data are in agreement with
observations in transgenic mouse models (220,223). A clinical trial involving the
administration of IVIG to five patients with AD with simultaneous measurements
of CSF and serum Ab levels has shown a decrease of total CSF Ab levels by 30%
following IVIG. Total Ab increased in the serum by 230%. No significant change
was found in CSF/ serum Ab42 levels. ADAS-cog was improved by 3.7 points
(G2.9), though MMSE scores remained essentially unchanged. Although the
sample size of this pilot study is too small to draw a clear conclusion, the study
provides evidence for a more detailed investigation of IVIG for the treatment of
AD (224). Due to the very preliminary nature of the findings, the question
of whether soluble Ab-antibodies in serum and CSF has potential value as a
biomarker of biological disease activity or a potential surrogate endpoint for
clinical trials cannot be finally answered (225).
DISCUSSION
Results and overall direction of change, respectively, that have been reported so
far on the biomarkers as discussed in this chapter are given in Table 1. The
majority of studies presented here refer to Ab42, t-tau, and p-tau in the CSF and
yield the most convincing evidence for these core biomarkers to be useful either
Biomarkers in Blood and Cerebrospinal Fluid 91
Cerebrospinal fluid
Ab42 Y Y4 Y Y Y Y
Ab40 4Y 4 4 4 4 4
APP Y4 [ 4 4 4
t-tau [ [ [4 [4 [4 [[[
p-tau [[ [ ([) 4 ([) 4
ApoE [Y4 [ [4 [4 [
Isoprostanes [ 4 [
ACT [4 4
Interleukin-6 Y4
receptor complex
C1q Y
Oxysterols and [ [
cholesterol
3-Nitrotyrosine [
14-3-3 Protein [ [ [[[
Serum
Ab42 4[ Y[
Ab40 4[ Y[4
APP
t-tau
p-tau
ApoE Y4 4
Isoprostanes 4[ 4[
ACT 4[
Interleukin-6 Y
receptor complex
CRP [ [
C1q
Homocysteine 4[ 4[
Oxysterols and Y[ Y[ 4
cholesterol
Overall direction of change of biomarkers in CSF and serum of patients with Alzheimer’s disease, mild
cognitive impairment, VaD, frontotemporal dementia, LBD, and Creutzfeldt–Jakob disease as far as
data are available. Blanks indicate that the issue has not been addressed so far. Please note that the
database for this overview, i.e., the number of studies investigating a particular biomarker, varies
considerably between markers.
Key: [, increased; 4, not different; Y, decreased in comparison to non-demented controls, respectively.
Abbreviations: VaD, vascular dementia; DLB, dementia with Lewy bodies.
92 Hampel and Buerger
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INTRODUCTION
There has been a major expansion in recent years on research into the genetic
causes of dementia, and a simple search on the PubMed database including the
terms “Dementia” and “Genetics” in March 2005 identified more than 13,000
references from 1964. Following the description of Alzheimer’s first case, the
first suggestion that genetic factors may play a role in Alzheimer’s disease (AD)
were reported in a paper of Lua (1920) and subsequently in a case report by
Flugel (1922), both speculating a possible genetic inheritance of the disease.
Since then, from 1930 to 1990, more than 50 pedigrees with a familial form of
AD have been described, suggesting that familial aggregation is a common
feature of AD. This observation has been successively confirmed by several
epidemiological studies (1,2). The analysis of these collected families and the
possible genotype-phenotype correlations, together with the outstanding progress
of molecular biological techniques, have made the field of dementia genetics in
the past 15 years an area of intense research and fruitful discoveries. This
information assembled in the past few years has led to the principal finding that
analysis of the genetic mechanisms underlying familial clustering of the disease
is likely to be directly relevant to the pathogenesis of the common and apparently
sporadic forms of AD. In addition, today’s knowledge of genetic factors is
currently evolving and leading to the progressive definition of a putative cascade
109
110 Sorbi et al.
Alzheimer’s Disease
AD is a genetically complex and heterogeneous neurodegenerative disorder
characterized by memory loss that leads to progressive and irreversible cognitive
decline until death.
The neuropathology of AD is characterized by widespread neuronal
degeneration, the abundant presence of neuritic plaques, containing beta-
amyloid, and neurofibrillary tangles, mainly composed of tau aggregates,
dystrophic cortical neurites, and amyloid microangiopathy. Senile plaques are
located in the extracellular space of the brain and are mainly formed by a highly
hydrophobic peptide that contains a 42 amino acid residues (A beta 1–42),
produced from its precursor amyloid precursor protein (APP) through proteolytic
processing (see Chapter 11).
Since 1991, the results of genetic studies have led to the identification of
gene mutations and polymorphisms that can either cause AD or substantially
increase the risk of developing the disease. Mutations in three genes (Table 1),
APP, located on chromosome 21, and presenilin-1 (PSEN1) and presenilin-2
(PSEN2), located on chromosomes 14 and 1 respectively, result in familial AD
(FAD). The FAD cases account for approximately 5% to 10% of all early onset
AD cases, and mutations in PSEN1 are the most frequent. Although more than
140 different PSEN1 mutations are described in more than 200 families with
different ethnic origins, only 10 different mutations among 13 families have been
reported for the PSEN2 gene, and only about 20 families with APP mutations are
known (3,49).
The APP mutations account for a very small proportion (2–3%) of all
published cases of FAD and 5–7% of reported cases of early onset FAD. The
pathogenicity of these mutations has been strongly supported by the fact that they
are virtually 100% penetrant in FAD kindreds where they occur in affected or
at-risk individuals, but are absent in age-matched controls. Swedish and Flemish
mutations at codon 670/671 and at codon 692 are rare; mutations at codon 717
have been described in about 20 families worldwide from different ethnic origins,
including Anglo-Saxon, Italian, and Japanese subjects (4–6). Mutations in APP
have been shown to affect the release of Ab in transfected cells and patient
fibroblasts. Transgenic mice expressing the APP V717F mutation produce
numerous Ab deposits in the form of classical senile plaques, and the brains of
these animals exhibit other neuropathological features of AD including neuronal
and synaptic loss and gliosis (6,7).While the APP codon 717 mutations are
associated with overproduction of Ab1–42, the Swedish double missense mutant
leads to an increase in total Ab secretion. The Swedish mutant involves the
substitution of the two N-terminal amino acids of the Ab domain, presumably
rendering APP more susceptible to b-secretase activity. Therefore, FAD
mutations in APP appear to affect both Ab release and the intracellular
trafficking of APP.
Presenilins
Mutations in PSEN1 (8) and in the related PSEN2 (9,10) are found to be causative
in about 50% of kindreds with FAD. The coding region of PSEN1 is derived from
10 exons (numbered 3–12). So far, mutations have been found in six of the 10
coding exons, with exons 5 and 8 accounting for 65% of the mutations. As some
mutations result in a later onset age, it cannot yet be excluded that mutations in
PSEN1 may also result in late onset forms (O65 years) of the disease.
The FAD mutations in PSEN1 are missense mutations causing single
amino acid changes or, rarely, exonic deletions and appear to be 100% penetrant
and are best classified as autosomal dominant “causative” gene defects. In
contrast, only ten PSEN2 mutations have been identified, suggesting that
mutations on PSEN2 gene are a rare but possible cause of disease and that FAD
mutations are considerably more frequent in PSEN1 than in PSEN2.
In 1995 a large Italian AD kindred was identified with a mutation in PSEN2
consisting of a methionine to valine substitution at residue 239 (9).This mutation
is characterized by some peculiarities of the clinical and neuropathologic
phenotype compared to sporadic AD. In the autopsy analysis (11) of two
probands, in addition to neurofibrillary changes and Ab deposits, ectopic neurons
in the subcortical white matter containing neurofibrillary tangles were observed.
Furthermore, an unusually high number of ghost tangles in the cerebral cortex
were found. The major peculiarity of this family was the clinical onset with
epileptic seizures before the dementia (Fig. 1).
112 Sorbi et al.
Figure 2 Amyloid precursor protein (APP) cleavage and a- and b-secretase pathways and
production of b-amyloid peptide. Abbreviations: AICD, APP intracellular domain; CTF,
C-terminal fragment; sAPP, soluble APP; TM, transmembrane. Source: From Ref. 15.
major expression of BACE2 is localized outside of the brain and inhibition studies
using antisense tools or protease overexpression does not lead to variation in
Ab production.
Gamma-secretase catalyzes the intramembrane proteolysis of APP, and this
process is closely linked to the development of Ab1–42 peptide. This protease
cleaves APP to produce C83 and C99 and releases the APP intracellular domain
(AICD) into the cytoplasm, which can bind to cytoplasmic adaptor proteins and
translocate to the nucleus where it may regulate gene expression. The gamma-
secretase activity has several substrates in addition to APP, perhaps the most
important of which is Notch, a family of cell-surface receptors which are
essential for correct embryonic development. Recent findings suggest that
gamma-secretase is a complex of at least four integral membrane proteins:
presenilin, nicastrin, Aph-1, and Pen-2 (21), with the presenilins being the active
site of gamma-secretase.
Sequence analysis of presenilins demonstrates two intramembrane
aspartate residues, closely associated with the hydrophobic region that undergoes
endoproteolysis leading to the formation of the two active presenilin
heterodimers. Mutation of the two aspartate residues results in the blockade of
gamma-secretase cleavage of the C99 peptide, leading to a reduction of Abeta
production with simultaneous accumulation of C83 and C99 fragments and
simultaneous abolition of presenilin endoproteolytic processing (22). In addition
there is a close intracellular relationship between APP and PSEN1, which
coprecipitate together in small amounts to form complexes and are located in the
same intracellular vesicle compartments.
The evidence presented above supports the notion that a complex flow of
events, known as “amyloid cascade theory,” triggered by as yet unknown stimuli
or promoted by a single genetic defect, increases Ab42 production. A chronic
imbalance between the production and clearance of fibrillogenic Ab peptide with
a tendency to misfold and aggregate progressively reduces the efficacy of
synaptic transmission, promotes a microglial and astrocytic activation, and
disrupts neuronal membrane homeostasis and potentiates oxidative stress to the
cell. Recent findings suggest that the toxicity of Ab does not lie in the insoluble
fibrils that accumulate but rather in the soluble oligomeric intermediates (23).
Soluble Ab oligomers are found in human AD cerebrospinal fluid, and their total
amount correlates with the severity of the disease better than senile plaques (24).
Evidence suggests that the soluble oligomers can directly compromise synaptic
function and that senile plaques may function as reservoirs of fibrous polymers
that are in equilibrium with the diffusible species (25).
Because APP is axonally transported and processed in presynaptic
terminals, synapses are sites where oligomers of Ab may accumulate in high
amounts. Soluble oligomers of Ab42 inhibit long term potentiation in the
hippocampus of rodents, which may suggest that these oligomers are responsible
for memory impairment in AD patients. In transgenic mouse models of AD,
synaptic dysfunction and memory impairment can occur in the absence of any
Genetics: Facts and Perspectives 115
DEMENTIA
and may be one indication that a mutation in a particular gene may be present and
require further investigation (29,31–35). A patient’s and family’s knowledge of a
family history of dementia, however, brings with it certain anxieties, and
appropriate specialist counselling measures should be instituted prior to, and
during, any investigations (36,37). If an individual is, however, presented with
Genetics: Facts and Perspectives 117
early onset dementia suggesting AD, genetic analysis can be instituted if access
to facilities and appropriate technical support is present. In the first instance, only
the proband should be tested for the most likely mutations which will entail
sequencing the PSEN1 gene, followed by exons 16 and 17 of APP, and then, if
available, all coding exons of PSEN2, since such mutations of the latter are rare.
Given the fact that in many families with suspected AD, there is no evidence of
mutations in APP, PSEN1, and PSEN2 (29), it should be noted that families may
face a degree of uncertainty over the findings of any testing, and therefore
following testing, appropriate support should be made readily available (37).
If mutations in APP, PSEN1, or PSEN2 are identified, then confirmation of
diagnosis can be made, appropriate care and treatment of the patient provided,
support and counselling given to the family (36,38), and the possibility of
predictive testing undertaken. Predictive testing, however, has certain issues
associated with it (38) and should not be undertaken lightly due to the impact
upon life choices for the individual, and should only be undertaken in those
families where a mutation has been identified in an affected individual. Once
again, extensive counselling and support should be provided. In those found to be
at risk of developing AD because of the presence of a defined mutation,
additional support could be given in the form of longitudinal neuropsychiatric
assessment to determine if there are signs of cognitive impairment. Longitudinal
magnetic resonance imaging (MRI) studies (39,40) and possibly positron
emission tomography (PET) scanning may predict cognitive changes and give the
possibility of disease slowing treatments including cholinesterase inhibitors (41)
or vaccination against Ab, if it is possible given the side effects (42), then further
supportive measures can be undertaken.
of AD, whereas the E2 allele appears to have a protective effect. The risk for AD
conferred by APOE E4 allele increases in a dose-dependent fashion, being eight
times more likely for carriers of a double dose of E4 allele compared to APOE E3
carriers. Further epidemiological studies and genotype-phenotype correlations
have suggested that E3/E4 and E4/E4 genotypes are more frequent in female
patients with AD, with a compelling dose effect, and also that particular ethnic
populations (i.e., African Americans) show a higher frequency of the APOE E4
allele, and therefore have an increased risk of developing AD. In addition, several
studies, although controversial, have shown a role for ApoE in the mechanistic
processes that lead to brain injury and ultimately to dementia. ApoE may
contribute to amyloid and tau deposition in AD, binding to amyloid in a isoform
specific manner enhancing aggregation, and in a similar way, interfering with tau
binding to microtubules. APOE E4 is strongly associated with increased neuritic
plaques, and the presence of this allele increases the odds ratio for cerebral
amyloid angiopathy. In addition, APOE E4 carriers have shown reduced glucose
metabolism and choline acetyltransferase activity in selected brain regions,
demonstrate an accelerated cognitive decline, and are poor responders to
memory-enhancing drugs prescribed for AD.
Several studies have shown that the E4 allele is also overrepresented in
mild cognitive impairment (MCI), and an increased frequency of the allele is a
strong predictor of clinical progression from MCI to AD. Although the E4 allele
alone does not imply conversion to AD in MCI, the combination of genetic
assessment with functional brain imaging is now seen as a promising preclinical
AD detection strategy (see Chapter 8). Recent PET studies have demonstrated an
association between the E4 allele and an abnormal reduction of glucose
metabolism in normal individuals carrying the E4 genotype in the same brain
regions as found in AD patients. It remains unknown if this is a predictor of future
cognitive decline. In addition, there is evidence that the E4 allele leads to greater
longitudinal metabolic decline in healthy elderly persons converting to MCI.
Nonetheless, no study has been carried out to assess the impact of the E4 allele on
brain physiology in the conversion from MCI to AD (44).
Several studies have addressed the utility of APOE genotyping in the
diagnosis of AD, and the suggestion has been made that the presence of the E4
allele is indicative of AD in a patient presenting with dementia (45). The presence
of an APOE E4 allele is, however, neither sufficient or specific to support a
clinical diagnosis of AD on the basis of genetic testing alone (46), and currently it
offers little significant gain over currently used diagnostic tools. For prediction of
dementia development, there is almost universal agreement that APOE
genotyping should not be offered or used (47,48), and therefore it remains a
tool for academic use only.
APOE E4 allele frequency has also been associated with a number of
neurological diseases, mainly degenerative, in which the major pathogenic
step is represented by proteinaceous aggregates in the brain. This has suggested
a putative role of ApoE in the clearance and internalization of small peptides that
Genetics: Facts and Perspectives 119
APOE 19q32.2 C
ACE 17q23 C/K
ACT 14q32.1 C/K
BACE 1 11q23 C/K
BChE 3q26.1–q26.2 C/K
BH 17q11.1–q11.2 C/K
CATD 11p15.5 C/K
CST3 20p11.2 C/K
CTNNA3 10q21 C/K
5-HTT 17q11.1–q12 C/K
IDE 10q23–q25 C/K
LRP 1 12q13.1–13.3 C/K
NCSTN 1q23 C/K
NOS3 7q35 C/K
PEN2 19q13 C/K
PLAU 10q22 C/K
PS1 promoter 14q24 C/K
TGF-b1 19q13.1–q13.3 C/K
UBQLN1 9q21.2–q21.3 C/K
VLDL-R 9pter–p23 C/K
a2M 12p13.3–12.3 C/K
C/K indicates the positive and negative association studies. For specific indications on association
studies of a specific gene, visit the “Alzgene” database (http://www.alzforum.org/res/com/gen/alz-
gene/default.asp).
Source: From Ref. 3.
Perspectives
The knowledge of genes involved in the amyloid cascade has moved towards the
development of specific targets for possible therapeutic approach. There is a great
interest in identifying small molecular inhibitors of the beta and gamma
secretases, although problems have arisen with these strategies because these
proteases also process other critical substrates (e.g., gamma secretase processes
Notch). Possible selective inhibitors for APP include nonsteroidal anti-
inflammatory drugs and cholesterol-lowering drugs (20).
An alternative approach is to prevent aggregation of the Ab peptide and/or
promote its clearance. In this field, early work in APP transgenic mice
demonstrates that immunization with Ab significantly attenuates AD-like
pathology and ameliorates memory deficits (68,69). Although the mechanism
by which brain Ab is reduced following vaccination remains controversial, recent
preliminary reports indicate that Ab immunization may lower senile plaque load
and stabilize behavioral decline in humans with AD (70,71). However, the
findings of an adverse effect involving meningoencephalitis in approximately 6%
of active Ab-vaccinated patients in early clinical trials conducted in 300 patients
(42) raise questions about the safety of this approach. The development of
efficacious and safe immunogens with careful antigen and antibody selection, as
well as of new vaccination techniques for immuntherapy of AD, can be expected
in the future to maximize efficacy and minimize serious adverse events.
An alternative active approach that may carry a lower risk of adverse events is
mucosal (nasal or oral) vaccination. This method has been tested in APP
Genetics: Facts and Perspectives 123
transgenic mice and has led to significant decreases in plaque burden, Ab levels,
neuritic dystrophy, and gliosis (72).
For the identification of AD genes it would be useful to combine the genetic
map information with gene expression profiling data, which in itself provides a
powerful instrument to rapidly identify a set of genes differently expressed
between normal and diseased tissues (73). Only the integration of the information
from genetic maps (location of genes) and genomic maps based on gene
expression and indicating specific genes may lead to the rapid identification of
causal genes. The combination of these two strategies may prove to be useful for
AD, and this is already evidenced by preliminary studies (74).
Another important aspect that should be considered in order to gain an
understanding of the genetic basis of complex disorders is gene–gene interaction.
With the exception of APOE, which is universally recognized to have a high risk
associated with AD, the general understanding is that several genes, each with a
moderate effect, are involved in the aetiology of a complex disease such as AD.
Therefore it has been hypothesized that other possible candidate genes, having a
small effect on the risk of AD, may not be detectable in all association studies due
to power or sample heterogeneity issues, but that they may show a considerable
effect in conjunction with other candidate genes. Much larger case-control
samples are required for gene–gene interaction studies as stratification of the data
reduces the power considerably, and this represents a limitation. With the
exception of a few published cases-controls studies that have utilized more than
1000 subjects, most genetic studies use a smaller sample, raising questions about
the meaningfulness of the association data rather than providing promising future
aims. Only significant results from appropriately powered studies could be
considered for replication in other samples from different populations.
Frontotemporal Dementias
Frontotemporal dementias (FTD) are complex clinical entities, characterized
neuropathologically by glial and neuronal tau deposition with widespread
neuronal loss and gliosis (see Chapter 11), with a marked clinical and genetic
heterogeneity. The nosological classification is still under debate, even though
Consensus Criteria have been published (75). Common clinical features include
presenile age of onset (35 to 75 years age range) and psychotic symptoms at onset
with late memory impairment. Often extrapyramidal signs or motor neuron
involvement is present.
The disorder is quite rare, with a prevalence of 10–15 per 100,000
individuals aged less than 65 years, and representing about 15–20% of all the
presenile dementing syndromes. At least three clinical syndromes have been
described in the past: a frontal variant of FTD, with pronounced changes in
behavior and personality, semantic dementia, and primary progressive
(nonfluent) aphasia. These latter forms of the disease are purely sporadic, but
124 Sorbi et al.
approximately 40% of all FTD cases are familial, with a large clinical variability
between and within kinships with FTD.
Linkage analysis studies, trying to homogenize different families with
diverse clinical patterns, have led, more or less simultaneously worldwide, to the
identification of a significant locus on chromosome 17, in the same region as the
tau protein gene (TAU). Initial sequencing of TAU in chromosome 17-linked
families showed negative results. An early report of an intronic mutation in 1998
led to the subsequent identification of TAU defects as the cause of FTD in some
families. The reason for this initial delay could be related to the structure of TAU,
which is composed of 15 exons, of which 11 are expressed in the adult brain.
Exons 2, 3, and 10 are subject to alternative splicing, producing six different
isoforms of the protein. These six isoforms are characterized by the presence of
two diverse functional domains, the first being composed of variable NH2-
terminal inserts that interact with plasma membranes and the second by three or
four COOH-terminal microtubule binding repeats that ensure binding with
tubulin and promote microtubule assembly (Fig. 4).
The first molecular variation found in TAU was an intronic mutation (77).
Further identification of novel mutations clustering in exon 10 and nearby in
intron 10, where the mutations affects tau splicing resulting in an altered 3/4
repeat ratio, provided direct evidence that intronic mutations could alter gene
structure with loss of protein function. In fact, alterations in splice regulation
caused by these mutations may disrupt a putative stem-loop structure or impair
the binding of regulatory proteins to the repeat region (78). Such a different
conformation of mutated tau could promote excessive deposition of the protein
(also in absence of hyperphosphorylation) due to microtubule malfunction, and
slowing intraneuronal transport of substrates. Thus, precipitation of tau
aggregates and subsequent NFT formation could be interpreted as the
consequence of this process.
To date more than 100 families with FTD have been identified with over 30
different mutations in TAU (78). Nucleotide changes include missense mutations,
deletions, and splice site mutations. A direct correlation between the site of
mutation and a specific cellular pathology has been predicted by early studies,
suggesting that, for example, tau aggregates predominantly composed by
filaments with four repeats were more common in patients with exon or intron 10
mutations. Today, with the discovery of novel pathogenic mutations, this is no
longer confirmed, since mutations not clustering in the exon-intron 10 region
could not necessarily lead to a specific neuronal or glial pathology or lead to a
selective deposition of a particular isoform, (i.e., aberrant insoluble tau filaments
lacking isoforms with a specific number of NH2-terminal inserts).
Most missense mutations in the microtubule-binding region reduce the
ability of tau to interact with microtubules interfering with correct microtubule
assembly. The primary effect of these mutations may be equivalent to a partial
loss of function, with resultant microtubule destabilisation and deleterious effects
on cellular processes, such as rapid axonal transport.
Genetics: Facts and Perspectives 125
Recently a novel genetic mutation, E46K, has been found in the alpha synuclein
gene (SNCA) in a Basque family with autosomal dominant parkinsonism and the
typical pathological hallmarks of DLB (91), suggesting that genetic causes of
DLB share common mechanisms with genetic forms of PD (92,93). In the
recently described family with triplication of SNCA (94), some family members
presented with a clinical picture that could be considered to fulfil criteria for
DLB. However multiplication of SCNA has been investigated in pathologically
confirmed patients with DLB, without positive results (95). Beta synuclein is a
member of the synuclein family, and is highly homologous to alpha synuclein,
both being involved in synaptic function. In a recent report, Ohtake and
Limprasert suggest that mutations in beta-synuclein gene may predispose to
DLB (96). The recent identification of the dardarin gene (LRRK2) in families
with parkinsonism, frequently with dementia or a DLB presentation (97,98),
suggests the possibility that this gene may be important in some DLB/PD cases.
Understanding the pathophysiology of DLB appears important for
developing novel therapeutic strategies. Neuroprotection appears interesting
since there is evidence in DLB of significant neuronal dysfunction but no striking
cortical neuronal degeneration (see Chapter 11). Therefore, the consistent
evidence that DLB is more treatable than other neurodegenerative disorders with
available pharmacological management, make important the accurate identifi-
cation of patients with DLB and their differentiation from other common types
of dementia.
SUMMARY
An understanding of the genetics of dementia and AD has provided vital clues to
the underlying biology that leads to this disorder with the identification of APP,
PSEN1, and PSEN2. Genetic analysis, whilst useful in certain circumstances,
cannot currently be used for the majority of cases, as the APOE gene, the only
robustly identified gene for LOAD, is neither necessary nor sufficient for the
development of this disorder.
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5
Approaches to the Identification of Novel
Diagnostic and Therapeutic Targets
in Dementia
Kate E. Wilson
Institute for Ageing and Health, University of Newcastle, Newcastle
General Hospital, Newcastle upon Tyne, Tyne and Wear, U.K.
Chris Morris
Wolfson Unit of Clinical Pharmacology, Chemical Hazards and Poisons
Division–Newcastle, Health Protection Agency, and School of Neurology,
Neurobiology, and Psychiatry, The Medical School, University of Newcastle
upon Tyne, Newcastle upon Tyne, Tyne and Wear, U.K.
INTRODUCTION
With the increasing prevalence of dementia in the elderly and the recognition that
the syndrome itself is a heterogeneous disorder, there is a pressing need to
identify not only effective treatments, but also diagnostic aids that will selectively
and specifically define the different forms of dementia. Population-based studies
backed by neuropathological assessment are few; however, they show that
Alzheimer’s disease (AD), while still the major form of dementia, accounts for
about 60% of cases, with vascular dementia, dementia with Lewy bodies, frontal
lobe syndromes, and mixed pathology disorders also contributing to the overall
problem (1). Combining diagnostics and therapeutics will provide the most
significant benefits for patients, but with different syndromes making up the total
picture, it is vital to ensure that the diagnosis is correct. A major problem with
dementia diagnosis currently is the fact that gold standard diagnosis can only
come postmortem (see Chapter 11). Clinical diagnosis, while highly accurate in
137
138 Wilson and Morris
many tertiary referral centers, comes only after extensive examination over a
period of months, during which time specific treatments need to be applied. The
ability to provide a rapid and accurate diagnosis could therefore lead to
significant improvements in dementia treatment.
But how, particularly when the underlying basis for many aspects of
dementia is unknown, is it possible to identify new diagnostic targets, or new
targets for therapy? Can a technique also be applied early enough to provide
presymptomatic diagnosis? Various systems which are currently in use or are in
development allow potential novel diagnostic targets to be identified. These
methods, collectively termed functional genomics, permit the rapid and often
wholesale analysis of cells or tissues in a disease state or the effects of
pharmacological treatment, making the identification of changes possible in a
short space of time.
FUNCTIONAL GENOMICS
The various genome projects that have set out to define the molecular basis for
many major organisms, including man, and have provided the foundation for the
global analysis of cell and tissue function using functional genomics (2). By
cataloguing all the possible genes in an organism, the genome projects have
allowed the production of reagents capable of detecting all the expressed genes,
or a reference with which to identify gene products by. The former is exemplified
by the production of whole genome microarrays, now the method of choice for
determining gene expression, and the latter by the bioinformatics tools which,
when coupled to various mass spectrometry (MS) methods, can be used for high
throughput protein and peptide identification. For the identification of diagnostic
targets in dementia, the application of these methods allows the production of a
catalogue from which to choose suitable candidates for further testing.
spot, often only a few microns across, representing a single mRNA species.
Microarrays are probed using a reverse hybridization procedure where the RNA
sample is labeled, normally with a fluorescent reporter molecule, and then
hybridized to the DNA sequences on the array. Since hybridization is roughly
proportional to the amount of an mRNA species present in the sample, the
presence of this highly ordered array allows each individual gene sequence to be
analyzed for the amount of mRNA present in the sample.
For any given tissue or cell, there will be a specific set of genes that are
expressed and which defines the nature of that tissue or cell. Of the 35,000–40,000
genes in the human genome, normally in any cell only a limited fraction of these
genes will be expressed, perhaps only 20–30% of the genome for any given cell
type (2). The complexities of the brain, with multiple neuronal types, vasculature,
and supporting glia, mean that the gene expression profile of any one brain area can
be extensive, and it has been suggested that perhaps half of the genome is
represented by genes specific to the brain (2). Microarrays have been designed
which are sufficient size that they represent most if not all of the possible genes in
the human genome. For the definition of brain function in health and in disease,
such arrays have the capacity to rapidly assess and identify those genes that are
expressed within a specific tissue. By extracting RNA from brain tissue and
applying it to a microarray, it is therefore possible to determine which genes are
being expressed by that tissue and also the relative amount of the particular
transcript present. Comparing healthy tissue with diseased therefore permits the
identification of which genes are present or absent and also to what extent any
particular gene is being expressed.
One potential problem of brain tissue is its highly complex nature stemming
from the numerous cell types and connections. In the various forms of dementia,
pathology is often restricted to specific anatomical locations, but also key lesions are
restricted to certain cell types (see Chapter 11). For example, neurofibrillary tangle
formation in AD may be restricted to large pyramidal neurons, sparing interneurons
(8–10). Even in areas associated with pathology, only a limited subset of the
vulnerable neurons may be affected, leaving some free of pathology. Because of this
cellular heterogeneity, it can be difficult to detect gene expression changes in a single
cell type due to dilution effects or the effects of pathology. Using laser capture
microdissection, it is, however, possible using a focused laser beam to cut through a
thin tissue section and isolate single cells of a given type, or small areas of tissue (11).
Using this methodology, specific cell groups such as neurons can be profiled
independently of any other surrounding cells, allowing a diseased neuron to be
compared with its unaffected neighbor (12). While there may be only a few
picograms of RNA in a single neuron, this is insufficient to use on a microarray as
normally an array requires microgram amounts of RNA. It is, however, possible to
use linear amplification methods to amplify endogenous RNA in a reliable and
unbiased manner from nanogram amounts of RNA for microarray analysis (13,14).
By combining methods such as laser capture microdissection with linear
amplification of RNA and microarray-based gene expression profiling, it is possible
140 Wilson and Morris
to determine gene expression profiles in isolated neurons or glia in health and disease
(15). This has been achieved to some extent in dementia with expression profiling
having been reported in AD in particular. Perhaps of significance is the report by
Blalock and colleagues who studied AD cases and also individuals with mild
cognitive impairment (MCI) in order to identify genes expressed at the earliest
stages of dementia (16). This study analysing hippocampal gene expression
identified mRNA up-regulation in several pathways involved with oligodendrocyte
differentiation and lipid metabolism, though a down-regulation of energy
metabolism pathways was evident (16). Similarly, analysis of hippocampal CA1
gene expression in AD has been used to show that there is up-regulation of some of
the inflammatory signalling pathways in AD, possibly associated with reactive
gliosis (17). This latter study also showed decreased neurotrophin and apoptotic
signalling, possibly being associated with the loss of pyramidal neurones and
neurofibrillary tangle formation (17).
The primary results of a microarray experiment represent only the
beginning of a particular gene expression study. Standard levels of analysis
identify significantly altered levels of gene expression using relatively
conservative changes, often G2 standard deviations from mean of control
expression levels, making subtle changes in gene expression difficult to detect.
Using bioinformatics approaches, however, allows the detection of specific
pathways that are affected, frequently using clustering algorithms that place
differentially expressed genes into families and pathways based on gene ontology
terms. Since the results of a microarray analysis are at best semi-quantitative, all
microarray experiments require further validation, normally by semi-quantitative
real time PCR (Q-RT-PCR). It is therefore possible to accurately determine the
levels of gene expression for a specific gene identified using the microarray, by
comparison with an appropriate housekeeping gene. Q-RT-PCR systems now
exist which make it possible to determine the levels of expression for either a few
specific genes in several samples, or to determine expression levels of a hundred
or more genes in a single sample. By this route it is a relatively straightforward
task to validate the primary targets from a microarray analysis, but also to analyze
additional components of the pathway identified as being potentially changed on
the basis of bioinformatics analysis. Microarray analysis and Q-RT-PCR
therefore can determine differential gene expression in highly selected
tissue samples.
But what of the tissue to be used in the identification of specific markers for
dementia diagnosis? One approach will be the use of suitable animal models,
most notably transgenic models showing the pathological changes associated
with dementia, which allows considerable control over agonal and pre- and post-
mortem effects. The use of transgenic models offers the opportunity to look not
just at end stage disease as is often the case with human postmortem material, but
to look at early stage pathology and even animals before the development
of pathology. One caveat of such an approach would be the complexity of the
pathology in dementia involving amyloid deposition, tangle formation, gliosis,
Novel Diagnostic and Therapeutic Targets 141
Cases with MCI may be one particularly fruitful source of information since, if
the individuals die early in the disease course without developing dementia, these
cases may provide information relating to the gene expression changes associated
with the primary cause of disease allowing such therapies and markers to be
developed. By using proteomic methods to identify the protein products of these
gene changes in cerebrospinal fluid (CSF), plasma, etc., or by gene expression
profiling of lymphocytes, it may be possible to develop novel tests for rapid
diagnosis. If gene expression changes are confined to the CNS, then the
development of neuroimaging ligands utilizing these targets would be one route
to improving diagnosis. Altered gene expression also provides the foundation for
potential new therapeutic strategies based on specific biochemical pathways
affected in disease, and may afford opportunities for more rational disease-
based treatment.
PROTEOMICS
Proteomics, the analysis of the protein complement of a cell or tissue, represents
the natural extension to functional genomic studies where mRNA expression is
mapped. While the sequencing of the human genome has revealed 35,000–40,000
genes, the proteome is much larger and also more dynamic in nature presenting a
unique challenge. Alternative splicing of genes results in different isoforms of a
protein while post-translational modifications also lead to multiple gene products
from a single gene. Furthermore, proteins also differ physically due to amino acid
composition, for example, strongly basic, acidic, or hydrophobic, as well as
differences in protein folding that have an effect along with protein-protein
interactions. Given the complexity of the brain, and also where comparisons
between healthy and diseased tissue are involved, the data sets generated by
proteomic techniques can be extremely large. While microarray-based gene
expression profiling can provide a rapid means to identify potential candidates for
further analysis, it can be argued that changes in gene expression are not
necessarily reflected by changes in the respective protein (23). High-throughput
proteomic techniques are therefore ideal for the direct identification of
biomarkers of disease, not only in postmortem or biopsy tissue, but also in
serum, plasma, and CSF, making the discovery of novel diagnostics and
treatment markers more effective.
Currently, no single method in a single pass is capable of accurately
identifying all proteins expressed by a cell or tissue due to the enormous variation
in the physical and chemical properties of proteins. A standard approach in
proteomic methodologies, particularly for tissue samples, is therefore to use
fractionation methods to reduce complex samples and simplify analysis. Standard
methods include centrifugation in order to separate a cell or tissue sample into,
for example, cytoplasmic and mitochondrial fractions. Alternatively, the use of
various detergents, high pH, and chaotropes can be applied to samples to extract
insoluble proteins such as membrane proteins followed by chromatographic
Novel Diagnostic and Therapeutic Targets 143
2D Gel Electrophoresis
Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) is the most
commonly used technique in proteomics since it is a highly effective method for
simultaneously separating complex protein mixtures (24,25). It is the only
technique currently available which allows thousands of proteins to be resolved
with one format in a single experiment and remains unchallenged as such.
Proteins are firstly separated by charge [isoelectric point (pI)] using isoelectric
focusing, followed by separation by mass in the second-dimension gel, recent
advances allowing up to 10,000 individual spots to be distinguished on a single
large format gel (26).
First-dimension isoelectric focusing separates proteins according to their pI
(the pH at which a protein has no net charge) using immobilized pH gradients
composed of thin gel strips into which ampholytes (charged carrier molecules)
are incorporated. In an electric field, proteins migrate through the gel due to the
pH gradient effect created by the ampholytes and stop migrating once their
charge is net neutral. These strips are available in a variety of formats, from wide
pH ranges covering many pH units to narrow ranges which cover just one or
two pH units. While broad range strips are ideal for initial experiments, for
example, large scale sample screening, narrow range strips have the advantage of
allowing much greater resolution. A series of gels can be overlapped to provide
greater visualization of proteins within the same range as a broad range strip yet
with much greater resolution, improving image analysis and protein identification
(24,26). Despite the advances in technology, 2D-PAGE remains unsuitable for
highly acidic or basic proteins since a lack of good ampholytes means that
focusing below pH3 and above pH11 is poor. This tends to result in reduced
144 Wilson and Morris
respectively) is detected by MS, allowing direct comparison between test and control
sample. The first generation ICAT reagents have recently been improved by
incorporating an acid-cleavable linker, allowing removal of the biotin affinity tag
prior to MS but leaving the peptide isotopically labeled. This simplifies the analysis
so that greater numbers of peptides can be identified and quantified (51). ICAT
labeling has been employed to study protein changes induced in cultures of cortical
neurons by the chemotherapeutic agent campothecin and analyzed by LC/MS,
demonstrating changes in proteins involved in transcriptional regulation, protein
synthesis, and signal transduction (52). This method is suited to the study of
relatively insoluble proteins such as membrane proteins since these can be extracted
using strong ionic detergent prior to the labeling and digestion steps, creating
peptides which are also more soluble than whole proteins (48). Given that many
ligands for positron emission tomography (PET) and single photon emission
computed tomography (SPECT) are membrane receptors, the use of methods such as
ICAT have the potential to isolate novel targets for the development of new
neuroimaging ligands in dementia.
with signal transduction, the synaptic scaffold, adaptor proteins, and cell-cell
adhesion molecules (57). While some of the proteins were associated with other
cell types such as glial fibrillary acidic protein (57), the use of affinity purification
may enhance the ability to accurately define only those proteins specifically
associated with certain cell or organelle fractions (58).
Perhaps of direct relevance to dementia is the identification of markers
which indicate changes within the CNS and which are amenable to analysis by,
for instance, neuroimaging. Combining methods such as comparative 2D-PAGE,
ICAT, and MuDPIT has the potential to identify either novel pathways indicating
disease mechanisms, or biomarkers that may be present in CSF. As with methods
analysing RNA, analysis of proteins is not, however, without its difficulties. For
human postmortem material, while there is some suggestion that it is possible to
identify phosphoproteins (59), analysis of animal material postmortem suggests
that primary signalling pathways, many which rely on specific phosphorylation
events, may rapidly alter postmortem (60). This may be particularly relevant to
human studies, as similar changes appear to happen, with rapid down-regulation
of signaling pathways postmortem (Wilson, unpublished). Many of these
methods though have the potential to be used in comparative approaches and
several are now being used to provide a preliminary analysis of dementia.
FLUID ANALYSIS
One major goal in the diagnosis of any particular form of dementia would be the
availability of a simple screening tool based on key feature of the biology of the
disease. CSF and plasma testing provides an obvious route to disease marker
identification that could be applied in a routine setting (see Chapter 3). For AD,
markers such as Ab determination in plasma have shown negative results, and
in CSF have yielded conflicting results (61). Identification of elevations in
hyperphosphorylated tau in CSF in AD has improved specificity (62,63), though
the paucity of any studies with neuropathological assessment and the wide
variation of Ab and tau levels with any individual patient perhaps makes the
interpretation of these findings premature (61,64–66). Newer markers (47) have
also not been validated and therefore new markers are required. While peripheral
markers have been suggested to occur in many neurodegenerative diseases, none
have so far proved robust (61). This has been due to the commonly held view that
while there is CNS disease, there is little impact upon the periphery due to presence
of the blood-brain barrier and the very specific cell groups affected in diseases such
as AD (67). This view has recently been challenged, with the finding that there are
marked changes in lymphocyte gene expression following CNS damage (22).
Furthermore, in many psychiatric diseases, there is expression of CNS proteins on
lymphocytes, and their expression mirrors that found in the CNS (68,69). It is
therefore possible that in diseases such as AD there are changes associated with
148 Wilson and Morris
Mass Spectrometry
MS is the preferred method for the identification of proteins by providing
structural information such as peptide mass and amino acid sequence, as well as
information on protein modifications. The data obtained can then be used to
identify a protein by searching various databases. MS measures the mass-to-
charge ratio (m/z) of gaseous ions produced by accelerating an ionized particle, in
this case the protein or peptide, through a rarefied atmosphere to a detector. Often
peptide mass fingerprint analysis is used where the isolated protein to be analyzed
is digested enzymatically, to cleave the protein at specific bonds giving a
reproducible pattern of digestion. MS is then performed on the peptide mixture,
giving masses with high accuracy the mass fingerprint of the protein. This
fingerprint is then compared to databases containing theoretical protein cleavage
data producing a list of the closest matching proteins (70). Much of this has been
achieved by matrix-assisted laser desorption/ionization (MALDI) MS which can
be used not only to identify isolated proteins, but also to directly identify peptides
from tissue samples (71). Ultra high resolution methods such as FT-NMR-MS are
now being used which may allow the detection and characterization of low-
abundance peptides in body fluids (72).
by comparing the peak profiles of the tissue, to identify peptides which differ
between, for instance, case and control.
One major drawback, however, is the use of a relatively mild ionization
procedure which limits the eventual mass resolution of the system. Furthermore,
this also provides only a mass/charge ratio for any peak which could potentially
correspond to several, if not hundreds, of possible peptide sequences. Peptide
identification is therefore difficult unless the peptide is identified in several
different runs, and frequently the specific peptide peak has to be isolated and
identified by more traditional methods such as chromatography. Direct separation
using LC (2D LC) and fractionation of samples (77) along with high-throughput
liquid-handling robotics are now though being coupled directly to MALDI and
ESI-MS to produce more powerful systems. Here it is possible to directly analyze
the individual protein peaks in the peptide profile using TOF/TOF MS and MSn,
and the identity of the peptide can be determined rapidly. There is therefore the
prospect that proteins and peptides identified in a complex mass spectrum as
being differentially expressed between cases and controls can be directly
identified. For example, Ab has been identified in the lens of Alzheimer’s patients
suggesting that the pathological features of the disease overlap between brain and
lens (78). The identification of ovarian and prostate cancer-associated biomarkers
have also demonstrated the usefulness of direct MS-based analysis of biological
fluids for rapid discovery of markers which have extremely high specificity and
sensitivity in a clinical setting (79,80).
Protein Microarrays
Protein microarrays are gaining in popularity as miniature ligand-binding assays
which can be used for complex protein samples since they allow detection and
also quantitation of proteins. With protein arrays, a frequent approach is where
antibodies are immobilized at a high density on a solid support such as a treated
glass microscope slide. When exposed, each individual antibody captures its
target protein from the sample. By arraying hundreds or thousands of antibodies
on a single slide, this method allows large-scale and high-throughput analysis
using small sample volumes and relatively low protein concentrations (81,82).
Like gene arrays, protein arrays are probed by direct labeling of the protein
sample with fluorescent dyes (e.g., Cy3 or Cy5) and the abundance of a protein
being related to the fluorescent intensity of the particular spot on the array (82).
One problem with this approach is low sensitivity, though new approaches
include multiplexed sandwich immunoassays with ultra-high femtomolar
sensitivity (83). A major problem with this approach is the relatively limited
availability of antibodies, and in particular monoclonal antibodies, which allow
arrays to be reproduced particularly on the large scale required in proteomics.
While antibody arrays are an obvious choice for protein detection microarrays, an
alternative is the use of recombinant antibody fragments or immunoglobulin
fragments expressed on the surface of bacteriophage which provides a rapid
150 Wilson and Morris
CONCLUSIONS
The diagnosis of dementia can only be achieved by the skilled clinician using
various clinical tools and personal judgement to assess the individual patient.
Currently the definitive diagnosis of dementia, particularly in the early stages, is
not possible, and reagents and tools are required which will assist the clinician in
achieving this. Using a combination of methods should, however, allow
determination of the gene and protein expression patterns of key brain regions
in health and disease which can define not only the presence of dementia, but of a
specific form of dementia such as AD. Techniques such as those described here
are already becoming routine in the search for biomarkers that can be used to
predict and diagnose disease, and to produce highly specific diagnostic tests. This
will be particularly useful in dementia diagnosis where, for example, symptoms
tend to overlap in the various different common forms, and yet accurate diagnosis
is required if the most effective treatments are to be used. However, if biomarkers
are to be used as diagnostic tools, techniques are required which are not only
sensitive and reliable but also reproducible allowing for multi-center use.
Ultimately, the technologies described here will help to unravel both the genetic
and environmental factors that predispose and precipitate the development of
dementia. These global technologies should therefore be seen, not simply as a
rapid means of identifying biological changes associated with dementia, but as a
route to more traditional methods of cell biological analysis for establishing how
dementia develops, and how new diagnostic tools can be produced.
Novel Diagnostic and Therapeutic Targets 151
ACKNOWLEDGMENTS
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6
Quantitative and Functional Magnetic
Resonance Imaging Techniques
157
158 Frisoni and Filippini
radio waves that are picked up by an antenna. The emitted radio wave signal is
then analyzed and processed.
The signal characteristics denote some of the properties of the emitting
atoms. Clinical MRI makes use of the signal emitted by hydrogen atoms, which
are largely represented by water molecules (99.9%) and also have a strong
resonant frequency (42.577 MHz). The MRI signal gives information about
proton density, i.e., water density, and two more peculiar features are used:
T1 and T2 relaxation times. T1, also known as “spin-lattice,” denotes the time
hydrogen atoms required to revert to their initial magnetic equilibrium state
before magnetization. T2, or “spin-spin,” denotes the time hydrogen atoms
required to revert to their original orientation (dephase). T1 and T2 amplitudes
denote the difference between the MDMs of different tissue in the z-axis and in
the x-y plane, respectively, and the energy exchange of hydrogen atoms with the
surrounding molecules. In the water molecule, energy exchange is low and T1
and T2 are long, while in fatty tissue and in protein rich tissue the opposite is true.
Relaxation times of some tissues are reported in Table 1.
Repetition and echo times (TR and TE) can be modified by the scanner
operator in order to obtain greater tissue contrast based on T1 and T2: a short TR
enhances T1 differences, while a long TE enhances T2 differences.
Brain imaging acquisition time is relatively slow, ranging between 10 and
30 minutes. The bone signal is weak—black—due to low proton mobility, while
water—having long T1 and T2—will show black in T1-weighted and white in
T2-weighted images. A number of CNS (central nervous system) diseases feature
increased brain tissue water content and show as T1-hypointense and
T2-hyperintense images. Gray and white matter can be sharply differentiated
on T1-weighted sequences. Due to its higher water and lower lipid content, the
gray matter is hypointense in T1 and hyperintense in T2 images relative to
the white matter.
MRI sequences that are usually applied in clinical practice are:
T1 (anatomic sequence), where the cerebrospinal fluid (CSF) is black, the gray
matter is dark gray, and the white matter is light gray; T2 (so-called
“myelographic” or “pathological” sequence), where the CSF is white, the gray
matter is light gray, and the white matter is dark gray; fluid-attenuated inversion
Heart 870 57
Liver 250 44
Kidney 560 58
Fat 260 84
Gray matter 920 101
White matter 790 92
Source: From Ref. 1.
Quantitative and Functional MRI Techniques 159
recovery (FLAIR), similar to T2 images, but where the CSF is suppressed and set
to black, to allow better lesion contrast in proximity to CSF spaces; and proton
density (PD), where intensity is proportional to water content. Figure 1 shows the
above sequences of a 40- and a 70-year-old healthy person.
Figure 1 Axial magnetic resonance images of a 40- (left) and 70-year-old healthy person
(right). The images do not show any abnormal findings. The only minimal enlargement of
subarachnoid and frontal ventricular spaces of the older person should be appreciated.
Abbreviations: FLAIR, fluid-attenuated inversion recovery; DP, proton density.
160 Frisoni and Filippini
In active areas, blood flow can rise by 30–50%, while oxygen extraction
increases by only 5%. This leads to both a local increase of oxyhemoglobin
concentration, which has diamagnetic properties, and a reduction of deoxyhemo-
globin, which has paramagnetic properties. Changes of the oxy/deoxyhemoglobin
ratio in brain tissue is considered the physical-chemical basis of fMRI. Indeed
Pauling and Coryell showed that the presence of paramagnetic substances in the
blood could act as vascular markers acting as a natural endogenous contrast agent
(4). As such, the BOLD signal is an indirect marker of brain activity as it does not
evaluate brain activity but hemodynamic changes. Some believe that BOLD signal
changes are generated more by synaptic than neuronal body activity (5); this implies
that the region(s) apparently active during fMRI experiments can be remote from the
true site of neuronal activation. Moreover, as synaptic activity can be either
excitatory or inhibitory, the interpretation of fMRI results may be much less
straightforward than is currently believed (6). Recently Logothetis and colleagues
have analyzed the relationship between BOLD signal and local neural activity by
simultaneously acquiring electrophysiological and fMRI data from monkeys (7–9),
finding that the BOLD signal does reflect a local increase of neural activity (10) due
to both excitatory and inhibitory interneurons (11).
Presently fMRI, based on oxygen consumption, and PET, based on glucose
consumption, are the main functional techniques used to evaluate brain activity.
Although positron emission tomography (PET) and fMRI have a high spatial
resolution, the temporal resolution is limited due to the slower hemodynamic
changes related to neuronal depolarization. This is the reason for the combined
approach of PET/fMRI (high spatial resolution) with EEG/MEG (high temporal
resolution). Comparative studies between fMRI and PET have shown good
agreement, although important differences have also been highlighted (12–14).
Combined measurements of BOLD signal, CBF, cerebral metabolic rate of
oxygen (CMRO2), and oxygen extraction ratio (OER) have been performed. Feng
and colleagues have shown that CBF, CMRO2, and OER changes reached their
maximum approximately one second earlier than the BOLD signal change (15).
Non-invasive methods have been developed to improve the specificity of fMRI
data; one of the most interesting approaches is the so-called arterial spin labeling
(ASL) method, which allows simultaneous measurements of BOLD and CBF
responses providing a direct measurement of perfusion. Disadvantages of ASL
are lower signal-to-noise ratio than BOLD contrast and longer TR (16). With the
ASL method, Obata and colleagues have shown discrepancies between BOLD
and flow dynamics in primary and supplementary motor areas (17), while Uludag
and colleagues have suggested that ASL measurements of CBF change may be a
more reliable marker of neural activity than BOLD (18).
The BOLD signal is divided into three stages (Fig. 2). The “initial negative
dip” (20) appears about one second after neuronal activation and consists of a
mild signal intensity decrease under baseline. It is believed to be due to the
sudden deoxyhaemoglobin increase, subsequent to a stimulus-driven increase in
oxygen consumption, that immediately follows neuronal activation, temporarily
Quantitative and Functional MRI Techniques 161
Figure 2 Schematic representation of the common features of the fMRI BOLD response
to neuronal stimulation. Source: From Ref. 19.
uncoupled with the circulatory response. The “positive BOLD response” that
follows is a marked increase of signal intensity due to increased blood flow and
decreased oxy-/deoxyhemoglobin ratio. In this phase, the signal peak is reached.
At the very beginning of the positive bold response a sudden increase of the
BOLD signal can often be appreciated (“overshoot”), presumably due to a slow
cerebral blood volume adjustment in the face of a swift increase in cerebral
blood flow (21). The same principle applies to the start of the third stage
(“undershoot”), where the end of the task is followed by an abrupt below
threshold decrease of cerebral blood flow and a persistently high cerebral blood
volume which takes longer to return back to baseline values.
In this section two experimental designs typically used in fMRI research will
be briefly considered: block design and event related design. The former lasts
between 20 and 30 seconds, where between six and nine seconds are needed to
reach the activity peak, and between eight and 20 seconds to return to the baseline
level; the latter is variable. A BOLD response based on a typical “block design” is
divided into a “stimulation period” alternated with a “control period” (or “resting
period”).
The fMRI signal believed to be proportional to neuronal activation is the
difference between signal in the active and that in the control condition.
Activation maps are obtained through subtraction of the rest from the active
image. Well-defined functions such as motor and sensory tasks give relatively
sharp fMRI signals (22), while emotions, for example, due to slow and variable
beginning and impossibility to die off quickly, are more problematic. Despite
such methodological problems, some researchers have been able to successfully
study complex functions such as bereavement and empathy (23,24) using a
different technique that is the “event-related design.” In fact, it is difficult to
measure some cognitive states, like emotions, or some tasks not well temporally
defined, such as those analyzed in a typical “oddball paradigm,” with a block
design technique (25). The event-related design is a technique to detect
hemodynamic responses to brief stimuli or events (26). Individual, single trial
162
Table 2 Levels of Increasing Technological Sophistication of Tools to Rate Structural and Functional Imaging Findings in Patients with
Cognitive Impairment
Visual Routine acquisition Visual rating scales: Scheltens’ MTL 1–3 days Visual rating scales: 1–2 weeks
rating atrophy score (14) ARWMC scale (15)
Linear measures: width of the 1–3 days
temporal horn (16)
Volumetry 3D T1 acquisition manual Volumetric measures: hippocampal and 2–4 weeks Volumetric measures: 1 day
or semiautomatic post- entorhinal cortex volumes (17,18) thresholding of
processing WMHs (19)
Compu- 3D T1 acquisition Prospective whole brain assessment: brain – None –
tational software for boundary shift integral (20)
neuro- computerized post-pro-
anatomy cessing serial scans
Expertise denotes the training time needed to obtain accurate measurements.
Abbreviations: ARWMC scale, Age-Related White Matter Changes scale; MTL, medial temporal lobe; WMHs, white matter hyperintensities.
Source: From Ref. 28.
Frisoni and Filippini
Quantitative and Functional MRI Techniques 163
events are measured rather than a temporally integrated signal (27), as happens
for the block design. The main advantage of the event-related design is that
accidents such as habituation, anticipation, and strategy effects are greatly
limited, thus enhancing the possibility to draw powerful inferences.
Visual Rating
Regional Atrophy: Visual Rating Scales
MRI can directly visualise the hippocampus and other critical medial temporal
lobe (MTL) structures in substantial cytoarchitectonic detail. Scheltens and
colleagues (30) have developed a subjective visual rating scale to assess MTL
atrophy on plain MRI films (the subjective MTL atrophy score). T1 weighted
sequences are used and six coronal slices (slice thickness of 5 mm) parallel to the
brainstem axis are acquired from a midsagittal scout image, the first image being
acquired directly adjacent to the brainstem. The score is assigned based on visual
rating of the width of the choroid fissure, width of the temporal horn, and height
of the hippocampal formation (Fig. 3). In 41 patients with AD and 66 non-
demented controls, the subjective MTL atrophy score showed a correct
classification of 96%, comparing favourably with volumetry (93%) (31). In a
prospective study of 31 patients with minor cognitive impairment, the subjective
MTL atrophy score improved the predictive accuracy of age and delayed recall
score for AD at follow-up (32).
Subcortical Cerebrovascular Disease: Visual Rating Scales
The European Task Force on Age-Related White Matter Changes (33) has
developed the Age-Related White Matter Changes (ARWMC) scale (34). This is
a 4-point scale which rates white matter changes separately in five areas: frontal,
parieto-occipital, temporal, infratentorial/cerebellum, and “basal ganglia”
(striatum, globus pallidus, thalamus, internal/external capsule, and insula). The
first three areas are scored as (0) no lesions (including symmetrical, well-defined
caps or bands), (1) focal lesions, (2) beginning confluence of lesions, or (3)
diffuse involvement of the entire region, with or without involvement of U fibers.
Figure 4 Visual rating scales for subcortical cerebrovascular disease: the Age-Related
White Matter Changes (ARWMC) scale. Grade 1, focal lesions; grade 2, early beginning
confluent lesions; and grade 3, confluent lesions with diffuse involvement of a lobe.
Source: From Ref. 34.
Quantitative and Functional MRI Techniques 165
The infratentorial/cerebellum and basal ganglia are scored as (0) no lesions, (1)
only one focal lesion (O5 mm), (2) more than one focal lesion, or (3) confluent
lesions. The final result of the rating are five separate scores to grade subcortical
cerebrovascular disease in the different brain regions in each hemisphere (Fig. 4).
The interrater reliability was found to be moderate (kZ0.48). Figure 4 provides
instances of three patients of increasing severity.
One should not forget to detect lacunes, which can be recognized on MRI
scans as round areas with regular contours, usually larger than 5 mm, located
more often in the basal ganglia, and featuring hyperintensity in T2 and protonic
density and hypointensity in T1 sequences.
Volumetry
Regional Atrophy: Volumetric Measures
A T1-weighted 3D-technique is employed for MRI image acquisition
magnetization prepared rapid acquisition gradient recalled echo or spoiled
gradient (MP-RAGE or SPGR). After acquisition, the digital images need to be
reconstructed on coronal, 1–2 mm-thick slices. The hippocampus is then
manually traced on all the contiguous slices where it can be appreciated
(Fig. 5). In expert hands, the reliability is high, intraclass correlation coefficients
for hippocampal measurements being 0.95 for intrarater and 0.90 for interrater
variability (35).
Subcortical Cerebrovascular Disease: Thresholding of White
Matter Hyperintensities
Quantification of the volume of white matter hyperintensities (WMHs) based on
MRI can provide an objective measure of the severity of subcortical
Computational Neuroanatomy
Recently, advances in neuroscience and neuroimaging have led to an increasing
recognition that certain neuroanatomical structures may be affected preferentially
by particular diseases. Neurodegenerative brain diseases mark the brain with a
morphological “signature”; detecting this may be useful to enhance diagnosis,
particularly in diseases lacking in other diagnostic tools. Moreover, structural
changes provide markers to track the biological progression of disease. In 1993,
Quantitative and Functional MRI Techniques 167
despite a lack of support from clinical variables, interferon beta was approved by
the Food and Drug Administration based on data from MRI (38).
Recent developments in computer science may help detect early sensitive
and specific disease signatures. The new approaches are automated, avoiding
error-prone and labour-intensive manual measurements. Second, such algorithms
can offer unprecedented precision as some can detect brain volume differences of
0.5% between images from the same subject (39).
The effort to develop such algorithms has been referred to as computational
neuroanatomy (40).
The individual algorithms can be categorized into two broad classes:
algorithms devised to detect group differences at one point in time and algorithms
devised to detect prospective changes over time. The first category may be useful
to define disease-specific signatures. The second can be applied to one or more
individuals to track natural disease progression or as modified by treatment.
While most tools have been developed to compare groups, some are being
adapted to analyze individual cases, an issue of the greatest interest for the
practicing physician.
Computational anatomy algorithms generally involve some or all of the
following steps: (1) brain extraction (brain is separated from non-brain voxels),
(2) tissue segmentation (voxels representing gray and white matter and cerebro-
spinal fluid are separated based on intensity values), (3) spatial normalization
(also called registration; the voxels of interest are matched to a template or an
earlier scan from the same individual), and (4) statistical comparison of different
subject groups or points in time. The pivotal step of all methods is registration.
Here, cross-sectional methods match images of interest to a reference stereotactic
template (a typical brain or a typical hippocampus, etc.) or vice versa, while
prospective methods match sequential images of the same patients taken at
different times.
Registration strategies differ in scope (i.e., analysis of the whole brain or
preselected regions-of-interest) and mathematical approach (accounting for
global or local variability of the brain’s size and shape). Some cross-sectional
methods that account for global variability are completely automated (such as
voxel-based morphometry based on statistical parametric mapping by Ashburner
and Friston; see Chapter 7) (41), while those that account for local variability
often require manually positioned landmarks to precisely match the image to the
template (such as cortical pattern matching) (42). Longitudinal methods use the
complexity of each individual’s brain structure to align accurately an individual’s
serial images [such as the brain-boundary shift integral (BBSI) algorithm] (43).
To perform well, all methods need high spatial resolution and clear
differentiation between tissue types. Usually, 3D high-resolution T1-weighted
MR images [spoiled gradient (SPGR) or magnetization prepared rapid
acquisition gradient recalled echo (MP-RAGE)] acquired with conventional
1.5T MR scanners and 1 mm3 voxels (ideally isotropic) across the cranium
provide sufficient detail and contrast.
168 Frisoni and Filippini
Figure 7 Gray matter changes in Alzheimer’s disease assessed with Ashburner and
Friston’s voxel-based morphometry. Black arrows indicate voxels of decreased gray
matter density (p!.0001 uncorrected for multiple comparisons) in the amygdalar/hippo-
campal complex and temporal cortex in 29 mild to moderate (upper row, mean MMSE
21 G4) and three very mild (lower row, MMSE 26 and 27) AD subjects compared with 26
nondemented controls. Source: From Ref. 45.
Quantitative and Functional MRI Techniques 169
Figure 8 Rate of global brain atrophy computed with the BBSI method in 18 Alzheimer’s
disease patients and 31 controls. Tissue loss was 2.78% in patients and 0.24% per year in
controls. Note the lack of overlap between the groups. Source: From Ref. 39.
atrophy compared with controls can be calculated as 85% and 88%. The practical
implications of the results are given in Fig. 9, showing the incremental diagnostic
gain of a positive and negative visual assessment of medial temporal lobe atrophy
at any given pre-test chance (before taking MRI). For instance, in the case of a
pre-test probability of Alzheimer’s disease in a clinical setting of 0.50 (the same
as tossing a coin), a positive result adds 0.27 to give a post-test probability of 0.87
and a negative result lowers the post-test probability to 0.15.
Obviously, the higher the pre-test probability of Alzheimer’s disease, the
smaller the incremental gain obtained through neuroimaging exams. It should be
noted that to date naturalistic studies assessing the incremental value of imaging
exams—as well as studies assessing the incremental diagnostic value of any other
test used for the differential diagnosis of the dementias—are lacking. Thus,
strictly speaking, the inclusionary approach of imaging is poorly supported by
evidence. However, this fate is unfortunately shared by most radiologic and
non-radiologic diagnostic exams currently used in clinical medicine (59).
The Dementias
Alzheimer’s Disease
The preceding sections have already clearly outlined what structural MRI can
show in patients with Alzheimer’s disease, i.e., atrophy of medial temporal
structures (hippocampus, entorhinal cortex, and amygdala), of the parietotem-
poral region, and posterior cingulate. With ordinary analysis tools, i.e., the MRI
film and the physician’s own eyeballs and connected visual cortex, atrophy can
more easily be appreciated in medial temporal structures, while that in the
parietotemporal and posterior cingulate cortex is more difficult to discriminate
from age-associated changes.
Figure 10 shows how regional atrophy can be studied in a patient with AD
using assessment tools of increasing technological complexity. The anatomy of
the medial temporal lobe is such that hippocampal atrophy can be appreciated
even with CT. A CT-based marker of hippocampal atrophy (the radial width of
the temporal horn) has been shown to be sensitive and specific to AD in both
clinical and pathological series (60,61). Values greater than 5.3 mm denote
clinically significant medial temporal atrophy (45). The patient shown in
Figure 10 had a CT-based radial width of the temporal horn of 6.9 mm (above
the 99th percentile of the age-specific distribution), scored 2/4 to the right and
3/4 to the left on the MRI-based visual rating scale for medial temporal atrophy
(normal values 0 and 1), had the smallest (between right and left) hippocampal
volume of 91.5 (below the 2nd percentile of the age-specific distribution), and a
single-case analysis with voxel-based morphometry (age-adjusted at p!.05
uncorrected vs. 173 nondemented controls between 40 and 80 years of age)
showed regional gray matter loss bilaterally in the medial temporal and posterior
cingulate-precuneus regions.
Quantitative and Functional MRI Techniques 173
bodies and AD (72). Barber and colleagues (72) have shown that clinically
significant atrophy affected 100% of AD, 62% of Lewy body dementia patients,
and 4% of controls. Thus, the absence of medial temporal atrophy had 100%
specificity for dementia with Lewy bodies versus AD, while the presence of
atrophy had a sensitivity of only 38% versus controls. These data indicate that in
patients where the differential diagnosis lies between dementia with Lewy bodies
and AD, the absence of atrophy is strongly suggestive of the former. Unfortunately,
this happens in a minority (less than half) of Lewy body dementia patients.
Structural and functional imaging has allowed the elucidation of the
pathophysiology of visual hallucinations in dementia with Lewy bodies. Burton
and colleagues (73) have shown that hallucinations in dementia with Lewy
bodies are not due to occipital atrophy: unexpectedly, the occipital lobe volume
of Lewy body dementia patients with hallucinations is greater than that of
patients without hallucinations, and this holds true also in AD. Notably, despite
structural integrity, functional imaging techniques (see Chapters 7 and 8) have
shown marked occipital hypofunction in Lewy body dementia patients (74).
Figure 12 The “pulvinar sign” in a patient with the new variant of Creutzfeldt–Jakob
disease. (A) T2-weighted, (B) proton density-weighted, and (C) FLAIR magnetic
resonance scans. Source: From Refs. 80, 82.
176 Frisoni and Filippini
Creutzfeldt–Jakob Disease
About 7 in 10 patients with sporadic Creutzfeldt–Jakob disease show hyperintense
lesions in the caudate and putamen and mild thalamic hyperintensities on
T2-weighted images (78,79). The low sensitivity and specificity of these findings
make them of little use in differential diagnosis. However, the new variant of
Creutzfeldt–Jakob disease (due to bovine transmission) shows mild caudate and
putaminal findings and marked thalamic hyperintensities. These are located at the
level of the pulvinar nucleus (“pulvinar sign”) and represent a moderately sensitive
(80%) but highly specific (100%) marker (Fig. 12) (80,81). The sign can be
appreciated particularly well with FLAIR sequences (82). The pulvinar sign has
been included in a suggestion for clinical criteria for the diagnosis of the disease
(81) and appears to be the diagnostically most useful instrumental sign in that the
EEG is often nonspecific and 14-3-3 protein CSF levels have low sensitivity (81).
Vascular Dementia
The clinical diagnosis of vascular dementia is a thorny issue for the lack of a
pathological standard. Consequently, although structural imaging is key to directly
appreciate the vascular lesions, its role and usefulness are still hotly debated.
NINDS-AIREN criteria (83) allow four forms of vascular dementia: (1)
multiple infarcts due to embolism or large vessel disease, (2) strategic infarcts
(due to embolism, large or small vessel disease), (3) multiple lacunes in the basal
Figure 13 Vascular dementia due to (A) multiple infarcts due to large vessel disease but
no small vessel disease; (B) multiple infarcts (1: temporal, 2: striatal) associated with small
vessel disease (3: watershed, 4: white matter lesions of the periventricular and deep
subcortical white matter); (C) multiple lacunes in the basal ganglia; and (D) extensive
lesions of the periventricular white matter associated with one deep frontal lacune in the
white matter.
Quantitative and Functional MRI Techniques 177
ganglia and white matter (generally due to small vessel disease), and (4)
extensive lesions of the periventricular white matter (due to small vessel disease).
Figure 13 shows cases of vascular dementia with multiple infarcts due to large
vessel disease but no small vessel disease, multiple infarcts associated with
small vessel disease, and extensive lesions of the periventricular white matter.
While the number, volume, and site necessary for multiple infarcts to give
vascular dementia is undetermined, strategic infarcts associated with dementia
(or—better—cognitive impairment in multiple domains) have been described in
the left angular gyrus, mono- or bilaterally in the thalamus, and bilaterally in the
globus pallidus (Fig. 14).
The diagnosis of vascular dementia due to extensive lesions of the
periventricular white matter and its differentiation from AD is particularly
difficult, although it is clinically relevant due to the poorer prognosis of the
former (85). Specific diagnostic criteria for subcortical vascular dementia have
been developed where imaging plays a major role, but these await validation (86).
Fazekas and colleagues (87,88) have shown that small punctate lesions of the
white matter (!5 mm, round, and with regular margins), when not associated
with confluent lesions, may not be due to vascular changes (87) and do not
progress over time (88). On the contrary, larger (O5 mm) lesions with irregular
margins (so called “confluent” as they appear to originate from the confluence of
smaller lesions) are due to vascular changes and progress over time (Fig. 15) (88).
Figure 14 Vascular dementia following strategic lesion. (A) Subcortical lesion in the left
angular gyrus (proton density-weighted image). (B) Unilateral (upper, FLAIR image) and
bilateral (lower, T2-weighted image) lesions following occlusion of the thalamic
paramedian artery. (C) Bilateral infarcts in the globus pallidus (T2 weighted image).
Source: From Ref. 84.
178 Frisoni and Filippini
MRI, (63,92) high levels of tau protein in the CSF, and perfusion and metabolic
defects [PET and single photon emission computed tomography (SPECT)] (93–96).
Jack et al. have tested the hypothesis that MRI-based measurements of
hippocampal volume are related to the risk of future conversion to Alzheimer’s
disease in older patients with MCI (63). Eighty consecutive patients who met
criteria for the diagnosis of MCI were recruited from the Mayo Clinic
Alzheimer’s Disease Center/Alzheimer’s Disease Patient Registry. At entry
enrolled subjects underwent an MRI examination, in order to obtain volumes of
both hippocampi, and for a period of time were also followed longitudinally, on
average 32.6 months, with approximately annual clinical/cognitive assessments.
During the period of observation 27 of the 80 MCI patients became demented,
and hippocampal atrophy at baseline was associated with crossover from MCI to
AD. The conclusions of Jack and colleagues were that MCI patients who will
progress to dementia feature lower hippocampal volume as measured through
high-resolution structural MRI.
Whether imaging or non-imaging markers are more promising for future use
in a routine clinical setting is still unknown. The accuracy of single markers varies
between 40% and 100%, (63,92–94,97,98) but studies are poorly comparable for
the doubtful reliability of current criteria when used in different memory clinics,
the heterogeneity of MCI patients enrolled, and the small study groups.
Voxel-based morphometry analysis in MCI has shown a significant agreement
in showing that patients had highly significant gray matter loss predominantly
affecting the medial temporal lobe, the hippocampal regions, the thalamus, the
cingulate gyrus, and extending also into the temporal neocortex (99–102).
A few studies have tried to include use of more than one Alzheimer’s disease
biomarker to discriminate MCI progressors from non-progressors. Okamura and
colleagues (103) studied 30 MCI patients, 22 of whom did and eight who did not
progress to cognitive impairment in the following three years, and found that a
high ratio between tau in the CSF and posterior cingulate perfusion on SPECT
could identify progressors with sensitivity of 89% and specificity of 90%. El Fakhri
and colleagues (104) studied 17 healthy controls, 56 non-demented patients with
memory problems who did not develop AD during three to five years of follow-up,
and 27 nondemented patients with memory problems who developed AD during
follow-up. Combining information coming from SPECT and structural MRI at
baseline allowed the correct classification of 94% of patients. If these early
findings can be replicated in larger and methodologically rigorous studies, the
possibility to diagnose Alzheimer’s disease before dementia has developed might
become a reality using functional and structural tools.
Physical
phenomenon Measure Indicative of Affected by
normal axonal structure, also gives rise to decreased FA, but the ADC is normal
or decreased due to the boundaries to proton motion represented by glial cell
membranes (Table 3). Neurofilament and tau pathology is associated with normal
ADC and FA, cell membranes being intact.
Magnetization transfer imaging is based on the exchange of magnetization
between free and macromolecule bound protons and allows one to indirectly observe
semisolids, such as protein matrices and cell membranes (Table 3). Changes in
tissue structural integrity such as gliosis lead to decreased bound and increased
free protons which show as decreased magnetization transfer ratio (Table 3).
There are grounds to believe that microstructural pathology exceeding the
macrostructural changes can be appreciated through T1-weighted images present
in AD, and this is strongly correlated with cognitive performance. Of the two
pathological hallmarks of the disease, senile plaques and neurofibrillary tangles,
the latter involves the cytoskeleton and neurobiological studies have shown that it
affects axonal transport (105). Moreover, pathological studies have shown that
neurofibrillary tangle density is more closely related to cognitive performance
than senile plaques (106). Structural T1-weighted MRI studies have shown that
the correlation between regional atrophy and global cognitive impairment is
relatively weak (107).
When structural changes are assessed with magnetization transfer and
diffusion imaging, a stronger correlation emerges (107). Bozzali and colleagues
have studied 18 AD patients with MMSE between 5 and 25 and 16 elderly
controls and found that the Pearson’s r correlations with the MMSE were 0.21 for
global brain volume, 0.31 for global diffusivity of the gray matter, and 0.58 for a
magnetization transfer index of the gray matter. A composite score compounding
information on atrophy and magnetization transfer reached rZ0.65, indicating
that 42% of the MMSE variance was accounted for by these two markers (108).
Hanyu and colleagues have studied 35 AD patients with MMSE between 9 and 25
and found that the MMSE was highly correlated with the magnetization transfer
ratio in the gray matter of the hippocampus (rZ0.70, corresponding to 49% of the
explained variance). The same group had previously shown in a group of 23 AD
patients with similar range of severity (four had MMSE !10, 13 had MMSE 11–20,
and six had MMSEO21) that the MMSE had a strong correlation with both
anisotropy (rZ0.65) and magnetization transfer ratio (rZ0.64) (109). However,
the correlation with the total callosal area, a proxy of white matter atrophy, was
significantly higher, reaching rZ0.74 (56% of explained variance). Yoshiura and
colleagues have studied 34 AD patients with MMSE between 3 and 28 and found
that the mean diffusivity of the white matter in the posterior cingulate gyrus was
significantly correlated with the MMSE (rZ0.53, 28% explained variance) (110).
Twenty-five patients with MCI have so far been studied with MT in two studies.
Not unexpectedly, both have found that MT measures of MCI patients are
intermediate between those of normal controls and AD patients (111,112).
Other studies have reported high correlations between global cognitive
performance and measures of diffusion of the white matter (113) and of T of the
182 Frisoni and Filippini
gray and white matter (111), but have included controls in the correlation. This
design can be criticized in that it inflates group variance through the unlikely
assumption that the amount of variance of cognitive performance in healthy
subjects that can be accounted for by MR measures of brain structural integrity is
similar to that in Alzheimer’s patients, and that the biological underpinnings are
also similar.
Taken together, these studies suggest that microstructural pathology of the
gray and white matter might be a strong correlate of cognitive impairment in AD.
However, although one study corrected diffusion and MT measures for brain
volume, thus at least partly accounting for the effect of atrophy (111), all the
others have so far failed to partial out the effect of global or regional atrophy from
that of diffusion and MT measures. Thus, it still remains to be determined
whether microstructural change has an effect on cognitive impairment
independently of the effect of macrostructural pathology.
Individual imaging modalities have variable relationships with cognitive
impairment in AD patients. Future studies will need to combine imaging
modalities in order to obtain a multispectral image of a brain where the variance
inter-modality and among different areas of the same modality might outline a
specific disease pattern. A pioneering study (114) has obtained information on
diffusion, MT, and proton spectroscopy in a single MRI exam in order to define
profiles of microstructural and biochemical changes in individual patients with a
range of neurological diseases (MS, subcortical vascular encephalopathy, major
stroke, AD, Alexander’s disease, and haemangioma), but the specificity of
the profiles still needs to be assessed. Prospective studies will need to assess the
progression of microstructural involvement with cognitive deterioration and
compare this correlation with that of other markers used to monitor the biological
progression of the disease and as surrogate outcome measure in clinical trials (40).
Table 4 Functional MRI Studies with Relevance to Dementia and Alzheimer’s Disease
Reference Aim Results Test performance
Dickerson Study functional activation pattern of MCI Enlarged hippo and parahippo activation with greater
et al., 2004 during a visual encoding task clinical impairment and subsequent memory
(121) decline
Enlarged hippo and parahippo activation was
correlated with better memory performance at
baseline
Grossman Neural basis for verb processing in AD Reduced activation in AD of normally activated Poorer in AD
et al., 2003 cortical regions (post-lat temp and inf fr regions)
(115)
Lipton et al., Study working memory in 2 monozygotic Enlarged parietal and reduced prefrontal cortex Poorer in the AD
2003 (122) twins discordant for AD activation twin
Lustig et al., Study activation-deactivation pattern in a No deactivation in AD of the normally deactivated Poorer in normal
2003 (123) semantic classification task medial post par cortex aging and AD
Enlargement in normal aging and AD of normally
activated regions (dorsal front cortex)
Machulda Study activation patterns of AD and MCI Reduced activation in AD and MCI of normally Poorer in AD and
et al., 2003 patients and normals during a visual activated regions (medial temp lobe) MCI
(116) encoding task Association between activation and performance on
recognition and recall
Rombouts Study working memory in frontotemporal Reduced activation in frontotemporal dementia of Similar in
et al., 2003 dementia regions activated in AD (fr and par cortex) frontotemporal
(117) Greater cerebellar activation in frontotemporal dementia and AD
dementia
Sperling et al., Study the effect of aging and AD on Reduced activation with aging of the dorsolat prefr
2003 (118) activation patterns during a learning task cortex and enlarged activation of par cortex
Frisoni and Filippini
Reduced activation in AD of medial temp and
increased of medial par and post cing regions
Li et al., 2002 Search for a functional MRI marker of Functional synchrony of the hippocampus separates
(124) early AD AD from controls with sens and spec of 80% and
90%
Prvulovic et al., Study visuospatial processing in AD Enlargement in AD of normally activated cortical Poorer in AD
2002 (125) regions (sup temp lobule and occ-temp cortex)
Atrophy partly accounts for differences between AD
and controls
Kato et al., Study nonverbal learning in AD Reduced activation in AD of normally activated Poorer in AD
2001 (119) regions (medial temp lobe and prefr temp and par
association cortices) and enlarged activation of
visual associative cortex
Bookheimer Study activation pattern of apoE4C normal Greater magnitude and enlargement in e4C of Similar in e4C
et al., 2000 persons during encoding task and the normally activated cortical regions (hippoc, par, and e4K
Quantitative and Functional MRI Techniques
(126) predictive power of activation patterns and prefr regions). Greater activation predicted
on subsequent memory decline memory decline after 2 years
Johnson et al., Study the relationship between functional Marked effect in AD of regional cortical atrophy on Poorer in AD
2000 (127) MRI activation and cerebral atrophy activation of the left inf fr but not of the left sup
with a semantic classification task in AD temp gyri
No effect of atrophy in controls
Pihlajamaki Study verbal fluency in normal persons Verbal fluency activates the medial temp lobe
et al., 2000
(128)
Rombouts Study nonverbal learning in AD Reduced activation in AD of normally activated Poorer in AD
et al., 2000 regions (hippoc and parahippoc gyri)
(120)
(Continued)
185
186
Table 4 Functional MRI Studies with Relevance to Dementia and Alzheimer’s Disease (Continued)
Thulborn et al., Study visuospatial attention in AD Activation in AD of silent areas (bilat dorsolat prefr Poorer in AD
2000 (129) cortex)
Reversal of the normal rightOleft activation in the
intraparietal sulcus
Saykin et al., Anatomic substrate of semantic memory Enlargement in AD of normally activated cortical Poorer in AD
1999 (130) impairment in AD regions (inf and middle fr gyrus)
Abbreviations: MCI, mild cognitive impairment; AD, Alzheimer’s disease; MRI, magnetic resonance imaging.
Frisoni and Filippini
Quantitative and Functional MRI Techniques 187
Figure 16 Effects of age and dementia on activation of the posterior cingulate area of the
default network during a semantic classification task: the region is initially activated in all
three groups, but the response in young adults quickly reverses, whereas DAT individuals
maintain activation throughout the task. (Left) Medial parietal posterior cingulate cortex.
(Right) Mean time courses of blood oxygenation level–dependent signal across active task
and passive fixation baseline conditions. Source: From Ref. 123.
activation of the default monitoring system might interfere with the activation of
task-specific regions, leading to poor performance in cognitive tasks. Better
understanding of the impairment of the default network in AD and other
dementias might help to understand the cognitive processes of patients while in
the resting state rather than performance in a challenged state—which has largely
been investigated in the last 30 years—as well as alertness and attention to outer
and inner stimuli. The neural basis of insight and awareness of own cognitive
deficits and of the surrounding environment, the development of agitation
following minor environmental stimulation in some AD patients, and the context-
dependent performance in activities of daily living might also be elucidated.
Some issues should always be kept in mind when interpreting fMRI results
in neurodegenerative conditions. First, fMRI does not allow differentiating
systems that can exert either activation or inhibition. This is a major hindrance for
tracking the cortical remodelling that likely takes place in AD and other
neurodegenerative disorders, whether excited or inhibited. Second, in normal
persons the same task gives rise to greater activation based on subjective
difficulty (cognitive tasks give more activation in persons with lower IQ, the
so-called “neural efficiency” hypothesis) (135,136). As disease specific tasks are
more difficult for patients with neurodegenerative disorders than normal persons,
this might tend to give greater activation. Unfortunately, the specific contribution
of subjective task difficulty cannot be assessed with current experimental designs.
Thus, the fMRI signal of patients with neurodegenerative conditions is the
summation of three trends: (1) lower activation due to neuronal or synaptic
damage, (2) greater activation due to compensatory recruitment, and (3) greater
activation due to subjective task difficulty. Their contribution to brain activation
remains to be investigated.
188 Frisoni and Filippini
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7
Perfusion Imaging with Single Photon
Emission Computed Tomography
197
198 Dougall and Ebmeier
images using both HMPAO and ECD showed that HMPAO is more stable in the
brain with no washout over time (7).
Both HMPAO and ECD underestimate perfusion in high flow areas, a
phenomenon that can be demonstrated with acetazolamide, which causes greater
increases in uptake with IMP than with the other two tracers (8,9).
Uptake of ECD depends on esterase activity, both cytosolic and membrane
bound (10), whereas HMPAO retention depends on the presence of glutathione (11).
This may, most of the time, ensure that variations in perfusion determine the
variability of tracer uptake, but it has to be understood that such variability may also
be due to factors specifically interfering with the tracer’s retention mechanism. This
will be of particular importance in pathological tissue. It appears that HMPAO
uptake in ischemic brain behaves like a rCBF tracer, i.e., is increased in areas of
luxury perfusion, while ECD and IMP do not show increased uptake, just like
regional oxygen consumption, or may even be reduced, as a marker of discrete tissue
damage (12–14). Similarly, ictal, inflammatory, and some neoplastic hyperper-
fusion is more obvious with HMPAO than with ECD SPECT (15–20). However,
relative lack of stability after reconstitution of HMPAO make ECD a logistically
more useful and effective tracer for ictal and peri-ictal studies (21,22).
Vascular lesions and hypo-perfusion in dementia appear to be detected more
easily with IMP or ECD than with HMPAO (23,24), although this may have little
impact on actual diagnostic accuracy (25). The situation may be reversed in AIDS
encephalopathy, with greater diagnostic accuracy for HMPAO than IMP (26).
ALZHEIMER’S DEMENTIA
The characteristic pattern of AD perfusion deficits in parietal and temporal lobes
is generally accepted and has been acknowledged in clinical guidelines (Figs. 1
and 2) (27). SPECT can provide rich information for the differential diagnosis of
dementia, but routine use is still controversial. Research studies have increasingly
documented modulating effects of age and gender on the typical perfusion pattern
of AD. Although confirmation of these effects is still required, the discussion here
will hopefully facilitate image interpretation and improve the usefulness of the
diagnostic changes observed with SPECT.
Effect of Age
Although presenile onset AD is recognized for its faster clinical decline and
greater symptom severity, the wider effect of age on SPECT AD diagnosis across
the whole aging spectrum has yet to be fully explored. One of the first studies by
Burns et al. (28) found age of onset correlated positively with parietal deficits and
negatively with medial temporal lobe perfusion. Jagust et al. (29) then reported
relative left frontal hypoperfusion in presenile AD compared with senile-onset
AD, and Caffarra et al. (30) tried to replicate this finding with patients matched by
illness severity, concluding that presenile AD was not associated with greater
SPECT changes than senile AD. O’Brien et al. (31) confirmed that the right
200 Dougall and Ebmeier
Figure 1 (See color insert.) Tc99m HMPAO SPECT scan of a healthy volunteer
(MMSEZ30); note that the parietal and temporal lobes are well perfused.
Figure 2 (See color insert.) Tc99m HMPAO SPECT scan of Alzheimer’s disease
(MMSEZ22), a characteristic pattern of perfusion deficits in parietal and temporal lobes.
85% of the early onset AD group had reductions in posterior association cortices
compared with medial temporal lobe. In contrast, Nitrini et al. (35) observed
bilateral parieto-temporal perfusion deficits in 23% of an early onset group and
71% of the late onset group—by combining parietal and temporal regions,
perhaps the subtle differences detected by Kemp and Hanyu were lost.
Dougall et al. (36), in a multi-center study comparing young and old AD
patients with age-matched mixed groups of healthy volunteers and depressed
subjects, found that an increase in diagnostic accuracy was achieved for younger
AD but not for the older AD group by using statistical parametric mapping (SPM)
maps in conjunction with SPECT images. Kaneko et al. (37), using 3D-SSP and
posterior cingulate reductions as a marker against other dementias, found that the
frequency of posterior cingulate hypoperfusion was greater in presenile AD than
in senile onset AD patients, thus providing further evidence of the heterogeneity
of AD across age groups.
Summary
Clinicians should be alert to possible age-related changes in the interpretation of
SPECT. Later onset AD appears to involve more medial temporal deficits and
increased image heterogeneity, and is therefore more challenging to diagnose. On
the other hand, younger onset AD has been associated with parietal and posterior
202 Dougall and Ebmeier
cingulate reductions. SPM and 3D-SSP appear to be more helpful in early than
late onset AD.
Effect of Gender
In a retrospective study of 104 SPECT scans of probable AD patients (NINCDS-
ADRDA), Nitrini et al. (35) found that disease severity, age of onset, and being
male was associated with bilateral parieto-temporal hypoperfusion. Thirty-nine
percent of females and 59% of males had this perfusion pattern. In a separate
study by Swartz et al. (38) of 63 probable and possible AD patients, both lower
MMSE scores and male gender significantly predicted reduced temporal and
parietal perfusion.
In a study by Ott et al. (39), left-right symmetric perfusion occurred more
frequently in male than female demented patients with probable AD (NINCDS-
ADRDA). Unilateral perfusion deficits in female probable AD patients were
found to be almost always on the left side; female gender and shorter disease
duration were independent predictors of unilateral left hemisphere
CBF reduction.
increase the spatial extent of reductions observed on SPMs compared with global
normalization (43). In the same multi-center study, Dougall et al. (36) found that
SPM used together with visual assessment increased specificity but not sensitivity.
3D Fractal Analysis
3D fractal analysis (3D-FA) objectively measures spatial perfusion differences by
calculating an index of heterogeneity called a fractal dimension. Yoshikawa
et al. (45) used both 3D-SSP and 3D-FA to compare VaD patients with healthy
volunteers. With 3D-SSP, reduced perfusion in VaD could be divided into two
abnormal patterns: global reduction and a decrease in frontal regions only. 3D-FA
demonstrated a difference between VaD and controls and a complete discrimi-
nation from patients with moderate and severe VaD. A correlation was found in
VaD patients between the fractal dimension of image heterogeneity and cognitive
impairment measured with the MMSE (46).
Using 3D-FA, Nagao et al. (47) found differences between AD subjects and
controls, and since then Nagao et al. (48) have found differences in heterogeneity
between FTD and AD subjects.
Neural Networks
Neural networks (NN) are used to model complex nonlinear datasets where
relationships exist between SPECT perfusion patterns and diagnosis. They are
trained to recognize patterns for disease simply by presentation of a scan training
set, so that new cases can then be classified by likely diagnosis. Using this method,
Chan et al. (49) found high diagnostic accuracy measured by AUC. Dawson
et al. (50) reported a sensitivity for AD of 75% against a specificity against controls
of 69%. Page et al. (51) found NN classified diagnosis more accurately than
alternative statistical techniques and visual assessment ratings. The largest and
most recent study by Warkentin et al. (52) using NN with 133Xe SPECT obtained a
high diagnostic accuracy measured by AUC, with 86% sensitivity for AD and 90%
specificity against controls.
204 Dougall and Ebmeier
COHORT STUDIES
Cohort studies of consecutive patients are particularly relevant as they mimic
clinical imaging practice. To date, published cohort studies are very
heterogeneous in their diagnostic composition. For instance, cohort studies
have recruited consecutive outpatients from a memory clinic (56–59), while other
studies have recruited consecutive patients with memory problems from a nuclear
medicine department (60), a neuropsychiatry unit including neurological
disorders (61), and from neuropsychology (62) (summarized in Table 1).
VASCULAR DEMENTIA
Several studies have attempted to discriminate AD from vascular dementia
(VaD) with varying success—typically using AD criteria of temporo-parietal or
posterior deficits and diffuse multi-focal abnormalities for VaD.
Table 1 SPECT Cohort Studies Are Heterogeneous in Diagnostic Mix and Not Generalizable to the General Population (Continued)
Clinical
Diagnosis definition
confirmed No. of of AD Age MMSE No. non MMSE Sensi- Speci-
First author by follow- AD in patient average mean AD in Clinical definition Age mean tivity ficity
(year) up cohort group (years) score cohort of control group average score (%) (%)
from MC—FTD
(47), VaD (21), DLB
(12), anxiety/depres-
sion (14), others (21)
Velakoulis No 9 AD (clini- 63 NR 47 Non-AD consecutive NR NR 88.9 78.7
(1998) cal cohort of neuropsy-
criteria chiatry unit patients
NR) with cognitive
impairment—FTD
(9), VaD (4), other
dementia (9), neuro-
logical diagnosis (7),
head injury (4), PD
(2), others (12)
(Continued)
207
208
Table 1 SPECT Cohort Studies Are Heterogeneous in Diagnostic Mix and Not Generalizable to the General Population (Continued)
Clinical
Diagnosis definition
confirmed No. of of AD Age MMSE No. non MMSE Sensi- Speci-
First author by follow- AD in patient average mean AD in Clinical definition Age mean tivity ficity
(year) up cohort group (years) score cohort of control group average score (%) (%)
FRONTOTEMPORAL DEMENTIA
SPECT perfusion deficits in anterior areas of the brain (frontal and anterior
temporal cortex) are associated with a frontal lobe dementing process.
Battistin 1990 No 19 Probable 67.7 9.0 15 VaD 71.5 16.9 89.5 80.0
Perfusion Imaging with SPECT
AD
Bergman 1997 Yes by 12 58 Mixed 75.5 21.6 17 VaD 72.9 22.4 55.0 70.6
months; then prob &
every 6–12 poss
months AD
Butler 1995 No 11 Probable 80.2 14.0 11 VaD 79.0 10.6 81.8 81.8
AD
deFigueir- 1995 No 20 Probable 72.3 14.5 22 VaD 77.0 20.8 80.0 86.4
edo AD
Launes 1991 No 36 Assumed 64.9 NR 33 VaD 68.0 NR 63.9 84.8
prob
AD
Lee 2002 Yes average 58 Probable NR NR 30 VaD NR NR 87.9 76.7
11.1 months AD
(Continued)
211
212
Table 2 HMPAO-SPECT Studies Comparing Alzheimer’s Disease and Vascular Dementia (1985–2002 Inclusive) (Continued)
Masterman 1997 Yes at 1 month 51 Probable 76.0 17.0 19 VaD NR NR 74.5 57.9
AD
Pasquier 1997 No 127 Probable 72.8 17.2 21 VaD 71.9 21.6 61.4 47.6
AD
Pavics 1999 No 33 Probable 67.0 19.0 18 VaD 68.0 23.0 70.0 66.7
AD
Sloan 1995 No 43 AD (DSM- 70.1 18.8 25 VaD 76.0 20.2 74.4 92.0
III-R)
Smith 1988 No 26 AD(DSM- 64.6 NR 25 VaD 67.8 NR 73.0 88.0
III-R)
Varma 2002 Yes, 6 monthly 22 Probable 62.8 18.0 18 VaD 66.4 21.0 77.3 66.6
for 1–3years AD
Villa 1995 No 23 Probable 67.9 17.9 12 VaD 68.9 NR 91.0 75.0
AD
Abbreviations: AD, Alzheimer’s disease; MMSE, mini mental state examination; NR, not reported; SPECT, single photon emission computed tomography; VaD,
vascular dementia.
Dougall and Ebmeier
Perfusion Imaging with SPECT 213
Summary
In the differential diagnosis of FTD from AD, the presence of frontal
hypoperfusion is highly specific for FTD even when observed in combination
with bilateral temporoparietal deficits. Anterior-posterior ratios appear more
useful in younger age groups. Whereas posterior cingulate reductions have been
reported as diagnostic for AD, conversely some evidence exists that anterior
cingulate reductions are diagnostic of FTD.
214
Table 3 HMPAO-SPECT Studies Classifying Alzheimer’s Disease from Frontotemporal Dementia (1985–2002 Inclusive)
Diagnosis
confirmed by Clinical defi- Age MMSE MMSE Sensi-
First clinical nition of AD average mean FTD Age mean tivity
author Year follow-up Number group (years) score Number group average score (%) Specificity (%)
Summary
Occipital and posterior cingulate reductions and perhaps also the absence of
medial temporal reductions are probable SPECT markers of DLB. Voxel-based
techniques such as SPM may be more sensitive in earlier detection compared
with ROI analysis.
Table 4 HMPAO-SPECT Studies Classifying Alzheimer’s Disease from Dementia with Lewy Bodies (1985–2002 Inclusive)
Diagnosis Age Clinical
confirmed Clinical defi- aver- MMSE definition MMSE Sensi-
First by clinical nition of AD age mean of control Age mean tivity
author Year follow-up Number patient group (years) score Number group average score (%) Specificity (%)
from
MCI
Huang Average of 82 Progressive 63.4 25.7 20 Stable MCI 59.6 26.9 Area under ROC
(2003) 26.4 months MCI curveZ0.75
Okamura Mean follow-up 22 Progression 71.9 25.6 8 Stable MCI 72.1 26.6 88.5 90.0
(2002) 3.1 years from
MCI to
AD on
follow-
up
Knapp NR 30 Probable 73.6 18.1 50 MCI 65.0 27.3 70.0 70.0
(1996) AD
Johnsona Yes, all subjects 56 Probable 77.4 18.0 27 Questionable AD 72.4 NR 80.4 77.8
(1998) AD (CDRZ0.5)
Johnsona Yes, all subjects 56 Probable 77.4 18.0 18 Converters to AD on 72.6 NR 80.4 94.4
(1998) AD mean follow-up
16.7 m (nZ18)
(Continued)
217
218
Table 5 Summary of Selected SPECT Studies Investigating Alzheimer’s Disease, Mild Cognitive Impairment, and Memory Problems
(Continued)
Clinical
Diagnosis con- definition
First firmed by of AD Age MMSE MMSE Sensi- Speci-
author clinical follow- Num- patient average mean Num- Definition of control Age aver- mean tivity ficity
(year) up ber group (years) score ber group age score (%) (%)
Greene Every 31 Probable 69.9 23.5 24 Controls with 65.4 NR 80.6 66.7
(1996) 6 months for AD memory problems
2 years
Hashi- NR 12 Probable 57.7 NR 18 Controls with 58.7 NR 100.0 100.0
kawa AD headaches or
(1995) dizziness
Mattman NR 128 73 Probable 70.0 NR 48 Controls with 65.0 NR 78.9 64.6
(1997) and 55 memory problems
possible
AD
Dougall and Ebmeier
Mielke NR 20 Probable 68.8 20.9 13 Controls with 59.5 28.8 80.0 65.0
(1994) AD memory problems
Muller NR 116 AD (DSM- 66.0 19.9 20 Controls with 56.0 28.8 48.0 75.0
(1999) III-R) memory problems
Perfusion Imaging with SPECT
and a probable AD group with mild to moderate severity. ARCD subjects had
pronounced asymmetrically decreased temporo-parietal perfusion at baseline
compared with controls that declined further on follow up.
A large HMPAO-SPECT study by Johnson et al. (93) investigated differences
between groups of normal cognition, questionable AD, converters to AD from
questionable AD by two years follow-up, and probable AD (NINCDS-ADRDA)
for preclinical predictors of developing AD. Singular value decomposition was
used to reduce the SPECT data set to 20 vector scores, which were analyzed in
turn by DFA. Probable AD could be differentiated from questionable AD with a
sensitivity of 80% and specificity of 78%, and from converters to AD (mean
16.7 months) with a sensitivity of 80% against a specificity of 94%.
Using patients as their own controls, Kogure et al. (94) observed the
changes of ECD-SPECT scans in 32 patients with MCI as they developed AD
(mean follow-up 15 months). At baseline, MCI patients had bilateral reductions
in posterior cingulate gyri and precunei compared with healthy volunteers—
using SPM for analysis. Further decreases on follow-up and after conversion to
AD were observed in left hippocampus and left parahippocampal gyrus.
Selective perfusion decreases in posterior cingulate gyrus and precuneus in MCI
as measured with sophisticated techniques of analysis may be a marker
predicting future AD.
More recently, a follow-up study by Tanaka et al. (95) retrospectively
examined non-demented subjects with memory loss of whom half converted to
AD within two years. SPECT patterns at initial presentation suggested that both
converters and non-converters had medial temporal and posterior cingulate
reductions in perfusion (using SPM), with additional reductions in parietal and
anterior cingulate in the individuals who later developed AD. SPECT could be
useful in predicting conversion to AD, but typical AD perfusion deficits were also
observed in subjects who did not convert to AD.
Posterior cingulate perfusion was again abnormal in an HMPAO study by
Huang et al. (96) of 54 consecutively recruited MCI subjects, 17 of whom
progressed to AD within two years (DSM-IV criteria). Activity in left posterior
cingulate cortex was found to be decreased in those who progressed compared to
those who retained a diagnosis of stable MCI. In a further SPM analysis of
baseline HMPAO-SPECT images, Huang et al. (97) found bilateral prefrontal
perfusion increases and parietal perfusion decreases (left more than the right) in
the progressive group compared to the stable group and controls; the authors did
not find any differences between groups in posterior cingulate—in contrast to
their previous study.
An attempt to combine 123I-IMP-SPECT and CSF tau protein levels to give
higher predictive accuracy was reported by Okamura et al. (98). Perfusion in
posterior cingulate was found to be lower in both progressive MCI and probable
AD compared with cognitively normal subjects and the stable MCI group. In
addition, CSF tau levels were higher in the progressive MCI and AD groups
compared with normal subjects and stable MCI. Combining CSF and CBF data in
Perfusion Imaging with SPECT 221
a ratio [CSF (tau level)/CBF (posterior cingulate)], and using an optimum cut-off
value, the authors achieved a sensitivity of 89% for progressive MCI and a
specificity of 90% against stable MCI.
Encinas et al. (99) found that medial and anterior temporal perfusion
deficits were of no value in predicting conversion to dementia since both stable
and progressive MCI groups showed these deficits; this is in agreement with
Tanaka et al. (95), who also found medial temporal deficits to be of no predictive
value. In contrast, right and left pre-frontal deficits discriminated stable MCI from
AD converters with accuracy measured by area under ROC curve of 75–78%.
El Fakhri et al. (100) extended the study of Johnson, this time comparing,
both independently and in combination, MRI volume with HMPAO-SPECT
perfusion estimates. Using DFA, both baseline SPECT and MRI were highly
significant predictors of conversion to AD; they discriminated between
questionable AD and converters to AD. Assessed by area under the ROC curve
(AUC), MRI had relatively higher sensitivity and lower specificity than SPECT
and vice versa. Taking MRI and SPECT in combination increased accuracy
further—providing evidence that the two methods contribute independent
information. For SPECT, Johnson reported that the best areas to discriminate
between questionable AD and converters were: amygdala followed by superior
temporal sulcus, basal forebrain, caudal anterior cingulate, hippocampus, and
rostral anterior cingulate. In this study, the classification rules were derived from
the study subjects themselves—testing these classification rules in a new group of
patients would provide further insight as to the usefulness of the methodology.
Not all published SPECT studies have positive findings. A 3-year
prospective study by McKelvey et al. (101) followed up MCI subjects at 9–12
monthly intervals and found that half progressed to probable AD. Of the MCI
subjects, 36% were judged to have normal SPECT scans by visual assessment
and 64% abnormal scans; 52% of those with abnormal scans progressed to
dementia compared with 55% of those with normal scans. Since SPECT
abnormalities did not predict cognitive decline (MMSE score) or conversion to
dementia, McKelvey concluded that SPECT was not useful in predicting
conversion—at least with visual inspection of images.
Summary
SPECT evidence demonstrates that the longitudinal spectrum of decline from
normal cognitive function to MCI, and possible further decline to early AD is
most likely associated with reductions in cingulate gyrus, in particular posterior
cingulate, possible precuneus, hippocampal, and amygdala involvement as well
as temporo-parietal cortex. It would appear that visual inspection of images does
not discriminate between MCI and healthy volunteers. Regions of interest
analysis is not as powerful or statistically robust as the technique of SPM for
detecting subtle changes in small structures prone to partial volume effects.
222 Dougall and Ebmeier
CONCLUSION
Clinical criteria such as NINCDS-ADRDA for “probable AD” against the
neuropathological gold standard have been estimated to have an average sensitivity
of 81% and specificity of 70% (102). Comparing SPECT against clinical criteria
in a meta-analysis finds it to be a test with relatively higher specificity than
sensitivity. Therefore SPECT can be a useful adjunct to clinical criteria in the
differential diagnosis of dementia. Diagnostic accuracy can be improved with
SPECT for those demented patients not conforming with clinical criteria or who
have possible co-morbid symptoms of vascular disease.
Recent research indicates that sophisticated multi-variate statistical
processing of image data such as SPM enhances the diagnostic power of
SPECT. Using such methods could help improve prognostic accuracy—for
example, in patients presenting with memory problems or MCI. Visual inspection
does not appear to be sufficient to discriminate between dementia sub-types in the
early stages. While SPM is readily available and has been successfully
implemented in clinical practice, subtle abnormalities associated with early
disease are more likely to be identified with SPM or 3D-SSP (94,95).
To date most SPECT studies have been concerned with defining accurate
markers for dementia sub-types that could be readily adopted in clinical use.
Future research should investigate the value of SPECT in combination with other
predictor variables, such as biological and cognitive markers, to optimize
prediction and, by implication, therapeutic power.
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228 Dougall and Ebmeier
Karl Herholz
Wolfson Molecular Imaging Centre, University of Manchester,
Manchester, U.K., and Department of Neurology,
University of Cologne, Cologne, Germany
229
230 Herholz
with age, most prominently seen in frontal cortex (10), but this has not been
confirmed in all studies.
Local brain glucose metabolism depends on brain function. During
functional activation lCMRglc increases, and it is decreased by sedation and
during slow-wave sleep (11). Thus, a valid measurement of lCMRglc must
include adherence to a defined functional state. Usually this is resting state
characterized by lying comfortably in the scanner in supine position in a quiet
examination room with dimmed lights. In many laboratories patients are asked to
close their eyes before beginning the examination and to keep them closed. It is
important to make subjects familiar with the surroundings prior to examination to
avoid any unnecessary level of anxiety and restlessness.
Figure 1 (See color insert.) Typical findings with FDG PET in Alzheimer’s disease.
has been used as an indicator with 93% accuracy (23). The same accuracy was
achieved even without image reconstruction by a special pattern extraction
technique from PET sinograms (36). Several discrimination functions combined
with principal component analysis or partial least-squares have been tested for
discrimination between AD and FTD in a sample of 48 patients with autopsy-
confirmed diagnosis and achieved accuracies between 80% and 90% (37).
Use of FDG PET to diagnose AD rests on the typical distribution of these
functional changes in brain. Impairment of local FDG uptake by itself, however,
is not specific for AD pathology and could potentially be caused by many other
disorders, e.g., ischemic lesions, when they just happen to affect those areas that
are typically affected in AD. Correspondingly, a high sensitivity in the order of
90% to 95% for FDG PET to detect AD has been documented in several studies,
but specificity for discrimination from other neurodegenerative disorders is lower
and in the order of 65–75% (38). Patients with Parkinson’s disease (PD) may
show a very similar metabolic impairment (39,40), even in absence of major
234 Herholz
cognitive deficits (41). These changes may even be reversible with successful
electrical stimulation of the subthalamic nucleus (42). With appropriate clinical
information, the “pseudo-AD” metabolic pattern should not be a major problem
because it is seen only in patients with long-standing PD who also have the
clinical motor symptoms of PD. Thus FDG PET scans always need to be
interpreted in the context of clinical history and symptoms. They also cannot
replace structural imaging by computed tomography (CT) or magnetic resonance
imaging (MRI), which are needed to recognize non-degenerative lesions such as
infarcts, tumors, hematomas, or hydrocephalus that may also lead to focal or
generalized cortical hypometabolism (43).
Patients with late-onset AD may show less difference between typically
affected and non-affected brain regions than usually seen in early-onset AD, which
could potentially lead to reduced diagnostic accuracy with FDG PET (44–46). This
could reflect the fact that at older age multifactor damage to the brain is likely to
accumulate, and, actually, also in neuropathological studies the proportion of
unclassifiable dementia is highest in the oldest old (see Chapter 11). Thus, in very
old multimorbid patients, FDG PET is probably of little diagnostic use, which is in
accord with general clinical wisdom.
Atrophy of hippocampal and parahippocampal structures is a main finding of
structural imaging in AD (see Chapter 6). Therefore one would expect also major
functional changes of lCMRglc in this brain area, but this has not generally been the
case (47). It is difficult to identify hippocampal metabolic impairment on FDG PET
scans because it exhibits lower resting metabolism than neocortex and pathological
changes are not obvious by visual image analysis. However, by coregistration of
MRI and standardized placement regions of interest onto the hippocampus in FDG
PET scans to increase spatial accuracy, a reduction, especially of entorhinal
metabolism, has indeed been observed in mild cognitive impairment (MCI) and AD
(48). In normal controls, lCMRglc in the neocortex is correlated with entorhinal
cortex lCMRglc across both hemispheres, whereas in AD patients these functional
metabolic correlations are largely lost (49). Metabolic impairment in the
parahippocampal gyrus had also been noted in a previous study during activation
using a simple memory task (50). As would be expected, hypometabolism in that
region is associated with memory impairment (51).
Atrophy may cause an apparent reduction in local FDG uptake due to
increased partial volume effects, even if local glucose metabolic rate in remaining
cortex was constant. Algorithms to correct for that effect have been applied in
AD (52,53) and concluded that partial volume effects contribute to apparent
hypometabolism but also that in spite of that local glucose metabolism remains
reduced in temporoparietal cortex even after correction. In contrast, the negative
correlation between frontal and perisylvian apparent glucose metabolism and age
seen in normal subjects could largely be resolved by partial volume correction (54).
Partial volume correction is not usually applied to clinical data because it requires
accurate coregistration with high-quality MRI, is very sensitive to slight mismatch
potentially producing severe artifacts, increases image noise, and is not needed for
FDG PET 235
other centrally acting cholinergic drugs generally did not show a main treatment
effect on cerebral glucose metabolism, but when subjects were divided into
responders and non-responders the former showed a metabolic increase that was
mostly located in frontal cortex and associated areas (69–71). These trials studied
the effect of drugs on CMRglc but did not address whether drugs might slow
disease progression. To perform such a trial, it has been estimated that 24 patients
with Alzheimer’s disease per active treatment and placebo group would be
needed to detect a significant 33% treatment response with p!0.05 and 80%
power in a one-year, double-blind, placebo-controlled treatment study (72).
comparing phosphorylated tau protein in CSF with FDG PET in MCI, Fellgiebl
et al. found similar findings with both tests (94). Seven of 16 patients had
increased phosphorylated tau levels, and six of them also had AD-typical FDG
PET findings.
FRONTOTEMPORAL DEMENTIA
Frontotemporal dementia (FTD) is characterized clinically by leading changes in
personality and behavior, such as apathy or disinhibition, whereas memory
impairment may be absent or less prominent (109). In practice, FTD is readily
identified on FDG PET scans by a distinct frontal or frontotemporal metabolic
FDG PET 239
Figure 3 (See color insert.) Asymmetric frontotemporal atrophy and metabolic impair-
ment in frontotemporal dementia.
VASCULAR DEMENTIA
Diagnosis of vascular dementia (VaD) is easily made in the case of multiple
cortical infarcts multi-infarct dementia, but may be a difficult issue in cases with
severe microvascular changes but without major cortical infarcts. There is
currently no consensus about clinical criteria, and correspondence between
existing criteria (e.g., ICD-10, DSM-IV, NINDS-AIREN, CAMDEX) is poor
(135,136). Several studies have suggested that a diffuse global reduction of
cerebral glucose metabolism is a typical finding in VaD (Fig. 4), and that the degree
of that reduction in association cortex is similar to that seen in AD (137,138). In
cases without gross cortical infarcts, the effects of WML are generalized and
frontal hypometabolism correlates with memory and global impairment, cognitive
as well as executive function. The effects of subcortical cerebrovascular disease
appear to converge on the frontal lobes but are diffuse, complex, and of modest
magnitude (139). Thus, the contrast between metabolic impairment in association
areas and preserved metabolism in primary areas, basal ganglia, and cerebellum,
which is typical for AD but not for VaD, seems to provide some distinction with
FDG PET between these two types of dementia (137). Only large white matter
lesions may cause moderate focal reductions of overlying cortical blood flow and
metabolism (140), and it therefore is likely that presence of the typical metabolic
AD pattern in demented patients with multiple deep white matter lesions does
FDG PET 241
Figure 4 (See color insert.) Global reduction of cerebral glucose metabolism in vascular
dementia.
indicate that this is predominant AD rather than VaD (141). However, a definitive
distinction between VaD and AD or the relative importance of vascular and
Alzheimer-type lesions in patients who have signs of both often remains uncertain,
even after autopsy and histopathological examination (see Chapter 11).
VaD may also be caused by strategic infarcts in structures that are essential
for integrating higher cognitive functions, especially the thalamus. Even small
thalamic lesions may lead to considerable deactivation of cortex, as demonstrated
by FDG PET (142–144).
CREUTZFELDT–JAKOB DISEASE
Clinically, this disease is characterized by rapidly progressive dementia, often
accompanied by insomnia, myoclonus, and other extrapyramidal disorders. In all
cases reported so far, cerebral glucose metabolism was severely reduced in a
multifocal fashion (145–149).
SUMMARY
Imaging cerebral glucose metabolism with PET has a proven potential for early
diagnosis of AD at the clinical stage of MCI. The typical regional pattern of
metabolic impairment differs between major neurodegenerative diseases that
may cause dementia. Thus, FDG PET also has the potential to improve early and
242 Herholz
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9
Imaging of Neurotransmitter Systems
in Dementia
Steen G. Hasselbalch
Neurobiology Research Unit and Memory Disorders Research Group,
Copenhagen University Hospital, Rigshospitalet, Copenhagen, Denmark
Gitte M. Knudsen
Neurobiology Research Unit, Copenhagen University Hospital,
Rigshospitalet, Copenhagen, Denmark
INTRODUCTION
Chemical neurotransmission as a tool for cell communication in the human brain
relies on the interaction of neurotransmitters acting on their target receptors. At
the moment, over 100 neurotransmitters with 300 corresponding receptors have
been identified. Malfunction of some of these neurotransmitter systems in
degenerative disorders may provide part of the pathophysiological basis for the
varying symptomatology found in these disorders.
Animal models, peripheral measures of receptors, neurotransmitters and their
metabolites, and clinical effects of neuropharmacological treatment are crucial to
improving our understanding of the underlying pharmacology involved in aging and
cognitive dysfunction. As opposed to these indirect approaches, molecular imaging
of the neurotransmitter systems with positron emission tomography (PET) or single
photon emission computed tomography (SPECT) is able to characterize the
functional state of human central receptor systems in vivo. PET is a powerful
technique for investigating human brain physiology and pathophysiology. When
used with appropriate radioligands, PET can reveal the distribution of
neuroreceptors in the living human brain, and their interactions with neurotrans-
mitters or administered drugs. PET radioligands suitable for these purposes give
large signals that are quantifiable in terms of robust measures of regional receptor
253
254 Hasselbalch and Knudsen
NORMAL AGING
Although the amount of receptors varies considerably between individuals (1) it
is well established that a number of different receptor systems decline with age—
frequently at a rate that varies between the different brain areas. This has been
demonstrated both in postmortem human and in brain imaging studies.
Postmortem studies have shown a loss of dopamine transporters amounting
to 5–7% per decade in striatum (2) and that dopaminergic D1 and D2 receptors
in striatum also decline with age (3). These findings have been corroborated
in imaging studies where van Dyck et al. found a 6.6% decrease per decade in
striatal dopamine transporters (Fig. 1) (4), and postsynaptically, dopamine D1 and
D2 receptors in the same areas seem to decrease at the same rate (5,6).
For the serotonergic system, postmortem studies showed evidence for a
decrease in 5-HT2A receptor binding with age (7). A gender-specific effect of age
on 5-HT1A receptor density was initially reported with an inverse relationship
between 5-HT1A receptor density and age in men, but not in women (8). PET
data from a large sample of healthy control subjects have, however, not been
able to confirm this finding, and the density of 5-HT1A receptors seemed to be
remarkably constant within the age span from 23 to 54 years (9). Neocortical
5-HT2A receptor binding, by contrast, was found to decline by 6% per decade in a
large group of healthy controls aged between 21 and 79 years (Fig. 2) (10),
corroborating previous findings in a smaller sample (11).
256 Hasselbalch and Knudsen
Figure 1 (See color insert.) Loss of dopamine transporters with age. Dopamine trans-
porters measured using SPECT and [123I]-PE2I in a 21-year-old healthy male (left) and in a
70-year-old healthy male (right). Source: Courtesy of the Neurobiology Research Unit,
Copenhagen, Denmark.
2
BP1
0
0 10 20 30 40 50 60 70 80 90
Age (years)
Figure 2 The correlation between age and the averaged binding potential of 5-HT2A
receptors for all cortical brain regions (measured with PET and [18F]-altanserin in healthy
controls). The regression coefficient corresponds to an average decrease of 5-HT2A
receptors of about 6% per decade. Source: From Ref. 10.
Imaging of Neurotransmitter Systems in Dementia 257
ALZHEIMER’S DISEASE
The earliest and most prominent symptom of AD is problems with episodic
memory due to failure of learning and retention of new material. Therefore
neurotransmitter systems involved in memory processes, such as the cholinergic
system, have generally been studied in more detail than other systems, although
lack of specific ligands for this system have hampered in vivo imaging data.
However, behavioural and neuropsychiatric symptoms are frequently occurring in
the course of AD, and even extrapyramidal symptoms are encountered in the
disease, sometimes rendering a firm diagnosis difficult. One of the purposes of past
and recent neurotransmitter imaging studies has been to aid clinicians in improving
diagnostic accuracy by revealing disease-specific neurotransmitter changes in the
most prevalent neurodegenerative disorders, including AD. Another purpose has
been to better understand the pathophysiology behind certain symptoms with the
ultimate goal of improving treatment. In the following, only receptor systems
considered of importance for AD are included. A summary of these main findings
is given in Table 2. Data based both on postmortem studies and in vivo emission
tomography studies will be reviewed and the potential impact of disturbances
within the receptor systems discussed.
Table 2 Single Photon Emission Computed Tomography and Positron Emission Tomography Neuroreceptor Studies in Alzheimer’s Disease
Number of Number of
Receptor system controls patients Ligand Results Author
Cholinergic
Muscarinic receptors 11 14 [123I]-QNB Y Weinberger 1991 (19)
4 8 [123I]-QNB 4 (Y) Wyper 1993 (20)
5 5 [123I]-IDEX Y Claus 1997 (21)
18 18 [11C]-NMPB Y Yoshida 1998 (22)
18 6 [11C]-NMPB 4 Zubieta 2001 (23)
Cholinergic
Nicotinic receptors 3 8 [11C]-nicotine Y (high affinity) Nordberg 1995 (24)
Cholinergic
Acetylcholin-esterase 8 5 [11C]-MP4A Y Iyo 1997 (25)
4 4 [11C]-MP4A Y Herholz 2000 (26)
14 28 [11C]-MP4A Y Shinotoh 2000 (27)
26 14 [11C]-PMP Y Kuhl DE 1999 (28)
GABAergic
GABAA receptors (central 5 6 [123I]-iomazenil Y Soricelli 1996 (29)
benzodiazepine receptors) 4 8 [123I]-iomazenil Y Fukuchi 1997 (30)
6 13 [11C]-flumazenil 4 Meyer 1995 (31)
5 5 [11C]-flumazenil 4 Ohyama 1999 (32)
Hasselbalch and Knudsen
Serotonergic
5-HT2A receptors 37 9 [11C]-setoperone Y Blin 1993 (33)
10 9 [18F]-altanserin Y Meltzer 1999 (34)
26 9 [123I]-R91150 Y Versijpt 2003 (35)
Glutamatergic
NMDA receptors 5 5 [11C]-MK 801 ([) Brown 1997 (36)
Microglia marker (peripheral
benzodiazepine receptors)
8 8 [11C]-PK11195 4 Groom 1995 (37)
15 8 [11C]-PK11195 [ Cagnin 2001 (38)
9 10 [123I]-PK11195 [ Versijpt 2003 (39)
Dopaminergic
D2 (striatum) 9 15 [123I]-IBZM Y Pizzolato 1996 (40)
10 10 [11C]-raclopride 4 Kemppainen 2000 (41)
5 10 [11C]-raclopride Y Tanaka 2003 (42)
D2(D3) (hippocampus) 11 14 [11C]-FLB 457 Y Kemppainen 2003 (43)
Dopaminergic
D1 (striatum) 8 10 [11C]-NNC 756 Y Kemppainen 2000 (41)
Imaging of Neurotransmitter Systems in Dementia
Dopaminergic
DAT 12 15 [18F]-dopa 4 Tyrrell 1990 (44)
15 12 [11C]-betaCFT Y Rinne 1998 (45)
33 34 [123I]-FP-CIT 4 O’Brien 2004 (46)
259
260 Hasselbalch and Knudsen
subtype as a target for specific amyloid binding (63), but at present there are no
suitable tracers available for in vivo imaging of this subtype. Recently,
radiohalogenated analogs of 3-(2(S)-azetidinylmethoxy)pyridine (A-85380)
were used successfully for the in vivo visualization of alpha4beta2 nicotinic
receptors in the human brain with PET/SPECT (64), but so far, data for 6-18F-
fluoro-A-85380 PET imaging in AD are still missing.
Muscarinic Receptors
Although some biopsy and autopsy studies point to nAChR as the receptor
subtype most affected in AD with relative sparing of the mAChR subtype (55,65),
a selective loss of the subtype M2 compared to M1 has been shown (66,67).
Recent studies suggesting a role for muscarinic agonists in regulating the
production of Ab-amyloid raise the possibility that selective M1 agonists could be
useful in treating not only the symptoms, but also the underlying cause(s) of AD.
Unfortunately, due to several methodological problems, imaging of mAChR has
so far revealed contradictory results, and has not been able to demonstrate the
selective loss of M2 as compared to M1 receptors obtained from postmortem
studies. Using non-selective M1/M2 mACh receptor tracers, PET and SPECT
studies have rather consistently shown a reduction in mAChR in AD (19–22).
Thus, using 11C-NMPB and PET, Yoshida et al. found significant reductions in
mAChR binding in the parietal cortex of AD patients as compared to controls.
However, taking some of several methodological problems into account, Zubieta
et al. could not demonstrate a reduction in mAChR in six mild to moderate AD
patients (23). In healthy APOE 34-positive subjects, a higher M2 receptor binding
as assessed with 18F-TZTP PET has been demonstrated as compared to APOE 34-
negative subjects, presumably because of a lower synaptic concentration of
acetylcholine in these individuals (16). So far, no studies have been conducted
with selective muscarinic tracers in AD, but future studies should reveal mAChR
subtypes are differentially affected in vivo in AD.
Choline Acetyltransferase and Acetylcholinesterase
Marked reductions in the enzymes ChAT and acetylcholinesterase (AChE) and
cholinergic neuronal loss have been reported in postmortem studies as well as in
biopsies from AD brains, and this cholinergic dysfunction correlates with the
cognitive function (65,68,69). Currently, there are no tracers to map the key
cholinergic enzyme ChAT, but AChE tracers are in use. Pappata et al. used a
labeled inhibitor of AChE (11C-physostigmine) in normal subjects to demonstrate
tracer distribution corresponding to that found in postmortem studies (70). The
PET tracer 11C-MP4A serves as an in vivo AChE substrate so that the local rate of
radiotracer hydrolysis can be followed by PET. Based on this methodology,
AChE levels were reported to be decreased by 30–40%, especially in
temporoparietal regions in 1997 by Iyo et al. (25), and this finding was later
confirmed by Kuhl and coworkers using 11C-PMP in patients with mild to
moderate AD (28). These data agreed with earlier postmortem data with regard to
262 Hasselbalch and Knudsen
Figure 3 (See color insert.) Acetylcholinesterase activity in the central nervous system
evaluated with C11-MP4A. Source: From Ref. 26.
Imaging of Neurotransmitter Systems in Dementia 263
in younger individuals with major depressive disorder, a PET study has revealed
a more widespread reduction in 5-HT1A receptor binding (80).
The 5-HT2A receptors are highly expressed in neocortex but less so in limbic
cortex and basal ganglia (84). 5-HT2A receptors are also associated with cholinergic
nerve terminals in the cerebral cortex and hippocampus. In a postmortem study of
hippocampus receptor binding, the largest decrease was found for 5-HT2 binding,
with N-methyl-D-aspartate (including glutamate, phencyclidine, and glycine
binding sites), quisqualate, kainic acid, adenosine A1, benzodiazepine, 5-HT1,
muscarinic cholinergic, beta-adrenergic, neurotensin, and opioid receptors showing
parallel but less pronounced reductions in binding (85). Bowen et al. also showed
that 5-HT1 and 5-HT2 receptors (7,86) and serotonin reuptake sites (87) were
decreased in postmortem AD studies with 5-HT2A receptors preferentially affected
over 5-HT1A receptors (86,88). Using 18F-setoperone and PET in moderately to
severely demented AD patients, Blin et al. demonstrated large reductions in 5-HT2A
receptor binding in frontal, temporal, parietal, and occipital cortices (33). This
finding was corroborated in a more recent study by Meltzer et al. (34). They studied
5-HT2A receptor binding using 18F-altanserin in patients with depression and in
patients with mild to moderate AD, including three with concurrent depression, and
compared these results to age-matched controls after controlling for partial volume
effects. No differences were, however, found between AD patients with and without
depression, but the limited sample size prevented any examination of relations
between neuropsychiatric symptoms and 5-HT2A receptor binding. Likewise, using
SPECT in nine AD patients, Versijpt et al. found a generally decreased 5-HT2A
receptor density in the orbitofrontal, prefrontal, lateral frontal, cingulate,
sensorimotor, parietal inferior, and occipital regions (35). In very early AD (mild
cognitive impairment of the amnestic type), reduction in 5-HT2A density has been
demonstrated, suggesting that the 5-HT2A receptors are affected early in the course
of the disease (Hasselbalch et al., unpublished data).
The 5-HT4 receptor is highly expressed in cortex and hippocampus and
seems to enhance memory formation through its action on cholinergic neurons
(89). One postmortem study found significant reductions of 5-HT4 receptors in
hippocampus (67%), and smaller reductions in the neocortex in postmortem AD
brains (90), whereas another study did not find 5-HT4 receptor density affected in
the frontal and temporal cortex in AD (91).
A more widespread decrease in 5-HT transporter sites was also found in
brain tissue from AD patients with depressive symptoms as compared to AD
patients without depressive symptoms (92).
In conclusion, both postmortem and imaging studies have consistently
found evidence for serotonergic dysfunction in AD with 5-HT2A receptors most
prominently affected. Although there is substantial evidence for a serotonergic
dysfunction in AD, no PET studies have so far addressed the relationship between
on one hand the serotonergic system, and on the other hand neuropsychiatric and
behavioral symptoms in AD. In particular, the question of to what extent deficits
Imaging of Neurotransmitter Systems in Dementia 265
Figure 4 (See color insert.) Dopamine transporter loss visualized with FP-CIT SPECT
in the differential diagnosis of dementia with Lewy bodies (DLB). Reductions in FP-CIT
binding occurred in the caudate and anterior and posterior putamens in subjects with DLB
compared with subjects with Alzheimer’s disease and controls. Transporter loss in DLBs
was of similar magnitude to that seen in Parkinson’s disease (PD), while the greatest loss in
all three areas was seen in those who had PD and dementia. In Parkinson’s disease, there
was a greater selective reduction in putamen uptake compared with DLB and PD and
dementia. Source: From Ref. 46.
268 Hasselbalch and Knudsen
FRONTOTEMPORAL DEMENTIA
With regard to neuroreceptor imaging, FTD is the least studied progressive
neurodegenerative disorder. Furthermore, FTD comprise a group of disorders
with different neuropathological findings, and therefore neurochemical post-
mortem studies might reflect this heterogeneity.
In a postmortem study of 16 FTD patients (10 Pick pathology patients, and
six patients with unspecific changes), cholinergic activity (measured as ChAT
activity and M1 receptors) was not significantly reduced in cortical areas in FTD
as compared to controls (108). This data supports the empirical clinical finding of
no effect of cholinergic drugs in FTD. By contrast, in the same study both 5-HT1A
and 5-HT2A receptors were reduced in temporal and frontal cortices in FTD as
compared to controls. Interestingly, serotonergic receptors were also lost in AD,
but only in temporal and parietal areas. Thus, there might be regional differences
in the serotonergic dysfunction in FTD and AD, but whether imaging of
serotonergic receptors might be helpful in the differential diagnosis remains to be
settled, since no imaging studies have explored this possibility.
Since extrapyramidal symptoms are relatively common in FTD, the
expected dopaminergic dysfunction has indeed been supported by one post-
mortem study in which neuronal loss in the substantia nigra was found (109).
In familial FTD with parkinsonism due to a mutation in chromosome 17, one
SPECT study (110) and one PET study (111) have found evidence for a reduced
dopamine transporter binding. In the latter study, striatal D2 receptors were
preserved much in line with what is found in PD. In a 11C-CFT PET study,
dopamine transporter binding was reduced by 18% and 14% in putamen and in
Imaging of Neurotransmitter Systems in Dementia 269
Figure 5 (See color insert.) Differences in cortical GABAA receptor density between
patients with leukoaraiosis and dementia (A) and patients with leukoaraiosis without
dementia (B), indicating that severe white matter ischemic lesions may cause cortico-
subcortical disruption, which may augment cognitive dysfunction. Source: From Ref. 116.
CONCLUSIONS
Among the primary degenerative disorders, AD is the most extensively
investigated when it comes to neuroreceptor studies. In postmortem studies,
marked reductions in some of the receptor systems, in particular 5-HT2 receptors,
nicotinic a4-b2, and a preferential loss of M2 as compared to M1 muscarinic
receptors have been found. Studies of muscarinic and dopaminergic receptors
have shown conflicting results. Some of these findings have been partially
replicated in in vivo imaging studies: a decrease in 5-HT2, nicotinic, and possibly
muscarinic receptors.
In DLB cholinergic and dopaminergic dysfunctions have been established,
and in FTD reductions in serotonergic receptors and dopamine reuptake are among
the neurochemical changes. Some of these results have been replicated in vivo in
neuroimaging studies, and imaging of neurotransmitters may prove useful in the
differential diagnosis between the primary neurodegenerative disorders.
Generally, the development of more selective radiotracers that are
accurately quantifying receptor binding would greatly enhance our knowledge
on in vivo changes in the transmitter systems in dementia disorders. Neurotrans-
mission imaging has an enormous potential for increasing our understanding of the
complex relationship between neurochemical processes and cognitive and
neuropsychiatric symptoms in dementia.
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10
Development and Application
of b-Amyloid Imaging Agents
in Alzheimer’s Disease
INTRODUCTION
The ability to localize and quantify amyloid deposition in the living brain can
advance the study and management of Alzheimer’s disease (AD) in several
important ways. This chapter describes some recent progress in the development
and application of amyloid imaging agents. Sensitive in vivo detection of amyloid
deposition could aid in early, perhaps even pre-clinical, diagnosis. Longitudinal
279
280 Klunk et al.
studies of amyloid deposition could shed new light onto the controversial “amyloid
cascade hypothesis.” The ability to assess amyloid deposition pre- and post-
treatment with anti-amyloid therapies could significantly facilitate the develop-
ment of these promising experimental treatments. Surrogate marker questions
must await clinical trials in which amyloid imaging is performed before and after
treatment with anti-amyloid therapies. While the results of initial studies are
promising, they must be followed by larger studies, employing a wider range of
disease severity, incorporating longitudinal studies, and examining amyloid
imaging agent retention in dementias other than AD.
finding. This focality underscores the need to have an in vivo method capable of
measuring amyloid burden in the entire brain when assessing the effects
of immunotherapy. The promising findings from the Ab-immunization trial have
prompted more intense interest in further refinements of the immunotherapeutic
anti-amyloid approach such as passive immunization with anti-Ab antibodies
(21,22) that should avoid many untoward effects of active immunization
including meningoencephalitis. These passive immunization trials have now
entered early-phase human studies.
three hours) and the half-life of the radionuclide would render such an approach
impractical for in vivo investigations in human subjects.
[18F]FDDNP
The second agent for the in vivo visualization of amyloid deposits was a
radiofluorinated derivative of the solvent and viscosity sensitive fluorophore
2-{1-[6-(dimethylamino)-2-naphthyl]ethylidene}malononitrile (DDNP), termed
[18F]FDDNP. In vitro characterization of FDDNP demonstrated that it bound to
amyloid-beta fibrils with high affinity and fluorescently stained both plaques and
NFTs at high concentrations (43,56). Human studies in AD patients (nZ9) and
controls (nZ7) showed slightly greater retention of [18F]FDDNP in frontal,
parietal, temporal, and occipital cortices at steady-state (60–120 minutes post
injection), though this increased cortical retention was shown to exceed the
reference region (pons) by only a margin of 10–15%. The area of highest
retention at equilibrium was a region encompassing hippocampus/amygdala/
entorhinal (h-a-e) cortex region, which was shown to exceed retention in the pons
by w30% (57). Interestingly, autopsy studies (4) showed that neuritic plaques are
more densely concentrated in neocortex, while the mesial temporal lobe
structures, including the h-a-e cortex region, contain the fewest neuritic plaques.
NFTs, in contrast, are densely concentrated in the mesial temporal lobe where
[18F]FDDNP retention is greatest (3).
Initial analyses of [18F]FDDNP PET data were performed using a novel
kinetic analysis method, termed the relative residence time (RRT), which relates
specific radiotracer binding to the negative net difference between the reciprocal
of the tissue clearance constants (k2) for the reference and target tissues. The
authors demonstrated significantly longer h-a-e RRT values for AD patients as
compared to controls, and that the value of the RRT parameter was significantly
correlated with Mini-Mental State Examination (MMSE) (58). A concern with
this method of analysis is that it is sensitive to both peak and steady-state levels of
[18F]FDDNP in tissue, which could potentially result in aberrations of the
outcome measure by blood flow and transport phenomenon. For instance, brain
areas with nearly identical equilibrium levels of tracer, such as temporal cortex
and occipital cortex, produced RRT estimates that differed by more than a factor
of 15. Additional analyses of [18F]FDDNP have been reported in a preliminary
form comparing 13 AD subjects (age: 76G8 years; MMSE: 18G6) and 10
controls (age: 63G10 years; MMSE: all scored 30). The method used a standard
uptake value (SUV) computed from 60–120 minutes tissue radioactivity
concentrations that were normalized to a reference region devoid of specific
binding (SUVR), such as cerebellum. These analyses have shown significant
differences (p!0.001) in the retention of [18F]FDDNP between control and AD
subjects in regions such as the medial temporal, parietal, and prefrontal cortices,
although the magnitude of the signal in the AD subjects did not exceed the control
value by more than 15% in any region (59). The applicability of conventional
quantitative analysis methodologies, such as compartmental modeling and
286 Klunk et al.
maximal in the left FRC. It is likely that the improved distinction between AD
and control subjects using PIB is a result of more rapid clearance of non-specific
binding. While the pattern of retention of [11C]SB-13 appears to mirror that of
PIB, the latter provides a greater dynamic range and improved distinction of AD
subjects from controls.
Pittsburgh Compound-B
The first human studies with PIB were presented in preliminary form in 2002 (35)
and were followed by a full report in 2004 (62). The initial study included 16
probable AD patients (65.9G11 years; 12.3G4 years education), six elderly age-
matched controls (69.0G7 years; 12.7G4 years education), and three 21-year-old
controls (young controls). The young controls were included in the study because of
the near certainty that these young subjects would represent true plaque-negative
controls. Dementia severity in the probable AD subject group, as evaluated by the
MMSE, varied from mild (MMSEZ18) to very mild/questionable (MMSEZ29)
with a mean MMSE of 24.9G3.4. Subjects were administered w300 MBq (8 mCi)
of PIB, and dynamic PET data were acquired for 60 minutes. In this study, neither
MRI images nor arterial blood samples were obtained for anatomical co-registration
and input function determination, and analyses of these studies were limited to semi-
quantitative SUV measures of PIB uptake. Nevertheless, striking differences in PIB
retention were observed between control and AD subjects in brain areas known to
contain significant amyloid deposits in AD [e.g., FRC, and parietal cortex (PAR)].
The six elderly control subjects showed a brain entry and clearance pattern that was
indistinguishable from that of the three young control subjects. This permitted young
and elderly control groups to be combined to form a unified (HC) group for
comparison to the AD patients. As a group, the control subjects showed rapid entry
and clearance of PIB from all cortical and subcortical gray matter areas, including
the cerebellar cortex (Fig. 1). Cerebellum, an area generally lacking fibrillar amyloid
plaques in AD, showed nearly identical uptake and clearance of PIB in the
cerebellum of HC and AD groups (Fig. 1A). Subcortical white matter showed
relatively lower entry and slower clearance in both HC subjects and AD patients
compared to cortical and subcortical gray matter areas (Fig. 1B). In contrast, the AD
patients showed a markedly enhanced retention of PIB compared to HC subjects in
areas of the brain known to contain high levels of amyloid deposits in AD (Figs. 1C,
D, and E), such as parietal and frontal cortices (2,4).
The regional distribution of PIB retention was clearly different in AD
patients compared to the HC subjects (Fig. 2). PIB accumulation in AD patients
as a group was most prominent in cortical association areas and lower in white
matter areas, a pattern consistent with that described in postmortem studies of
amyloid deposition in AD brain (2). PIB images from HC subjects showed little
or no PIB retention in cortical areas, leaving the subcortical white matter regions
highest in relative terms. In absolute terms, the accumulation of PIB in white
matter was essentially the same in AD patients and HC subjects (Fig. 1B).
A series of axial and sagittal SUV images provides a three-dimensional sense of
288 Klunk et al.
SUV
SUV
2 2 2
1 1 1
0 0 0
0 20 40 60 0 20 40 60 0 20 40 60
Time (min) Time (min) Time (min)
(D) (E)
5 5
Fr Par D Fr Par E
SWM Cb SWM Cb
4 4
Older Control AD
3 3
SUV
SUV
2 2
1 1
0 0
0 20 40 60 0 20 40 60
Time (min) Time (min)
the regional distribution of PIB retention (Fig. 3). The marked difference between
PIB retention in the AD patient and the HC subject is apparent throughout most of
the forebrain. FRC was widely affected in the AD patient, but intense PIB
retention also was observed in temporal and parietal cortices, part of the occipital
cortex, and in the striatum. Lateral temporal cortex (LTC) appeared to have
greater PIB accumulation than mesial temporal areas. Consistent with previous
reports of extensive amyloid deposition in the striatum of virtually all AD
patients (63–66), the striatum was found to have significantly higher PIB
retention in AD patients than in HC subjects. Cerebellar cortex (Fig. 3) showed
little PIB retention and was similar in AD patients and HC subjects. In general,
the observed pattern of PIB retention in AD subjects was found to be consistent
b-Amyloid Imaging Agents 289
Figure 2 (See color insert.) Pittsburgh compound-B (PIB) standard uptake value (SUV)
images demonstrate a marked difference between PIB retention in Alzheimer’s disease
(AD) patients and HC subjects. PET images of a 67-year-old HC subject (left) and a
79-year-old AD patient [AD6; Mini-Mental State ExaminationZ21; (right)]. (Top) PIB
SUV images summed over 40–60 minutes; (bottom): fluorodeoxyglucose (FDG) rCMRglc
images (mmol/min/100 ml). The left column shows lack of PIB retention in the entire gray
matter of the HC subject (top left) and normal FDG uptake (bottom left). Nonspecific PIB
retention is seen in the white matter (top left). The right column shows high PIB retention
in the frontal and temporoparietal cortices of the AD patient (top right) and a typical
pattern of FDG hypometabolism present in the temporoparietal cortex [arrows; (bottom
right)] along with preserved metabolic rate in the frontal cortex. PIB and FDG scans were
obtained within three days of each other. Source: From Ref. 62.
Figure 3 (See color insert.) Serial planes demonstrate the topography of Pittsburgh
compound-B (PIB) retention. Axial (top two rows) and sagittal (bottom two rows) PIB
standard uptake value (SUV) images of the subjects shown in Figure 2. The HC subject
data is shown in rows (1) and (3). The AD patient data is shown in rows (2) and (4). The
reference region, the cerebellum, can best be appreciated in the images at the far right. The
cerebellar peduncles (white matter) show some nonspecific retention, but the cerebellar
cortex shows negligible retention. Scale bar indicates relative levels of PIB SUV values.
Source: From Ref. 62.
amyloid deposits in whom the clinical diagnosis of AD was incorrect and would
not be confirmed by postmortem evaluation. In the elderly control group, the
oldest subject (76 years old) consistently showed the highest cortical PIB
retention and the lowest cortical rCMRglc (boxed circles in Fig. 4). This subject
had not expressed any subjective memory complaints and performed within the
normal range on the neuropsychological test battery except for difficulty copying
a complex cube. This type of case, which could be described as an asymptomatic
amyloid-positive case, highlights the issue of specificity versus early detection.
One possibility could be that a high PIB signal was obtained in the absence of
amyloid deposits (i.e., a false-positive). If this finding does represent the true
b-Amyloid Imaging Agents 291
2 2
1.5 1.5
SUV
SUV
1 1
0.5 0.5
0 *** *** ** 0
Temporal
Parietal
Frontal
Pons
SWM
Cb
Cb
(C) 80 (D) 80
*
rCMRglc (µmol/min/100mL)
rCMRglc (µmol/min/100mL)
60 60
40 40
20 20
0 0
binding assume reversible in vivo kinetics and provide outcome measures, such as
the binding potential (BP, unitless), distribution volume, and the distribution volume
ratio (DVR), which are related to Bmax, the concentration of free receptors, and the
ligand dissociation constant Kd.
Determination of the PIB tissue-to-plasma ratios for a variety of cortical and
subcortical regions showed that a constant ratio (plateau) was achieved after
approximately 20 minutes in the cerebellar regions of both AD and control subjects.
Tissue-to-plasma ratios in regions known to contain significant amyloid deposits in
AD, such as the posterior cingulate gyrus (PCG), also achieved a constant ratio that
was greater than threefold higher than that observed for controls, though the time to
plateau was considerably delayed (45–50 minutes). Furthermore, the specific
binding of [3H]PIB to human AD brain tissue homogenates was shown to be clearly
reversible with an off-rate (koff) of 0.0027/min, or a clearance half-life of
252 minutes (73). These observations were consistent with reversible radioligand
binding and supported the use of models of reversible radioligand binding.
Compartmental modeling of PIB data showed that PIB kinetics were best
described by a model that allowed for influx and efflux from two tissue
compartments (2T-4k) for all subject groups and regions, including cerebellum.
Nonspecific PIB retention was found to be similar across subjects, as no
significant group differences in the cerebellar 2T-4k DV were observed. In
cortical regions, the 2T-4k DVR value showed greater PIB retention in PCG,
PAR, LTC, and FRC relative to control subjects.
The Logan graphical analysis, a regression method for describing
reversible radioligand binding, was applied to regional and cerebellar PIB data
over the 35–90 minutes post-injection interval. Linear regression of the
graphical variables yields slope values that are equivalent to the 2T-4k
radiotracer DV (77). Good agreement between compartmental and Logan DVR
values (e.g., PCG: rZ0.89, slopeZ0.91) was observed, although the
Logan results were considerably less variable. Statistically significant differences
(p!0.05) in Logan DVR values were observed for AD subject relative to
controls in cortical areas after correction for multiple comparisons. The Logan
DVR measure also demonstrated favorable intrasubject variability, as
assessed in five subjects (2AD, 1 MCI, 2C) who returned for a repeat PIB PET
scan 8–20 days later. Test-retest variability averaged 8.4G5.4% across 11
cortical and subcortical regions-of-interest, and 6.1G1.5% across the five areas
with the highest PIB retention in AD: PCG, PAR, FRC, LTC, and caudate.
The intersubject variability was greatest in regions dominated by non-specific
PIB retention, such as the subcortical white matter and pons.
In general, mean MCI Logan DVR values fell between control and AD
means in brain areas that showed high PIB retention in AD. Figure 5 shows that
individual MCI subjects do not fall as a group into an area that is intermediate
between the control and AD ranges, but, instead, MCI subjects are either
indistinguishable from controls, or indistinguishable from AD subjects. This was
equally true for SUV and compartmental analyses (data not shown).
294 Klunk et al.
of bias, test-retest variability, and Cohen’s effect size (AD vs. control) (79).
Carotid volume-of-interest-based methods showed the lowest bias of any method
relative to the metabolite-corrected arterial methods. Although the test-retest
variability was the poorest of all the simplified methods examined, the carotid-
based methods demonstrated acceptable test-retest variability (G7.1% across 11
regions). Cerebellar input methods showed negative biases relative to the
metabolite-corrected arterial methods that were greater in subjects with higher
levels of PIB retention, although these cerebellar reference-based
methods showed the lowest test-retest variability of any method (G4.5% across
11 regions) and a large effect size [w6.5 for PCG]. The single-scan SUV-based
methods showed the largest effect sizes for AD and control group differences (6.9
for PCG) and performed very well in terms of inter-subject and test-retest
variability (G5.0%), although the SUV measures were positively biased relative
to the arterial methods. While the results of the comparison of simplified methods
of PIB analysis are preliminary, they suggest that several simplified methods of
analysis may be applicable to human PIB PET data, some requiring only a
20 minutes PET scanning session.
amyloid deposition may play in the various cognitive decrements found in the
“normal” old. The availability of amyloid imaging techniques that can measure the
amyloid load at multiple time points in living individuals who are not close to death
is thus a major advantage.
very stable and reproducible measure over short time periods with test-retest
variability of only 5–10% (73). Therefore, reductions in amyloid load of 20% or
more should be easily measurable, once corrected for the increase in amyloid
load over the time period of the study determined from natural history studies.
One must consider whether imaging large fibrillar Ab deposits is the
appropriate method for assessing the efficacy of anti-amyloid therapies. Evidence
is accumulating to suggest that small, soluble oligomers may be the toxic species
of Ab in the human brain (91,92). Some would even suggest that large insoluble
aggregates of Ab are beneficial in that they serve to “detoxify” these soluble
oligomers. This concept appears to have some support from in vitro studies of
other amyloid-related pathologies (e.g., prion protein accumulator in yeast) in
which large intracellular aggregates of the protein in discrete cellular organelles
appear to be related to cell survival (93). However, plaques are extracellular
aggregates and are very different from the carefully managed intracellular
“detoxification” programs seen in cellular models of prion disease that use both
the cell membrane and organelle membrane to control the toxic waste. Plaques
are more akin to an unmanaged dumping of barrels of toxic waste into a land fill.
Even when the soluble/free toxin is removed, the plaque serves as a source from
which more toxin can leach into the surrounding extracellular environment. To
keep the potential danger of the insoluble Ab deposits in perspective, we must
remember that in AD brain, soluble forms of Ab make up less than 1% of total
brain Ab (94). In addition, it is almost certain that soluble and insoluble Ab pools
are in equilibrium. This is supported by the fact that it is extremely difficult to
separate soluble and insoluble Ab in the laboratory, because even after
ultracentrifugation and removal of soluble Ab, resuspension of the “insoluble”
Ab results in the appearance of more soluble Ab (unpublished observation).
Other support for this equilibrium comes from the observation that immunization
of amyloid-depositing transgenic mice in a manner that produces antibodies
specific for oligomeric Ab leads to marked reduction of not only oligomeric
forms of Ab, but also results in clearance of thioflavin-S positive plaque forms of
Ab as well (95,96). This suggests that any meaningful anti-amyloid therapy will
need to have a significant impact on insoluble brain Ab deposits for there to be a
long-lasting lowering of soluble Ab. Such an effect should be detectable with
agents that bind to fibrillar amyloid.
SUMMARY
In vivo amyloid imaging technology has largely overcome the hurdle of
achieving sufficient brain entry while retaining adequate affinity for the Ab target.
Brain clearance and the resulting specific labeling of Ab deposits also appears
sufficient in some of the existing tracers, but future radiopharmaceutical
development may result in tracers with even better signal-to-noise ratio and
sensitivity. Existing tracers all appear to target aggregated b-sheet forms of Ab,
but further work is necessary to precisely define the subtypes of Ab deposits that
b-Amyloid Imaging Agents 303
are labeled by each tracer. The early proof-of-concept phase has produced
convincing evidence that amyloid load can be quantitatively assessed in living
humans. Future challenges lie in the application of amyloid imaging to increase
the understanding of the natural history of amyloid deposition and its relationship
to cognitive change. Perhaps the most important impact of amyloid imaging
technology will be in the facilitation of the development of anti-amyloid
therapies, early (even preclinical) identification of cognitively normal individuals
with amyloid deposition who would be good candidates for anti-amyloid therapy,
and individual patient follow-up to monitor the success of anti-amyloid
treatment regimens.
ACKNOWLEDGMENTS
This work was supported by grants from The National Institutes of Health (R01
AG018402, P50 AG005133, K02 AG001039, R01 AG020226, R01 MH070729,
K01 MH001976), The Alzheimer’s Association (TLL-01-3381), The U.S.
Department of Energy (DE-FD02-03 ER63590), and GE Healthcare, Inc. GE
Healthcare entered into a license agreement with the University of Pittsburgh
based on some of the technology described in this manuscript. Drs. Klunk and
Mathis are co-inventors of PIB and, as such, have a financial interest in this
license agreement.
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11
A View on Early Diagnosis of Dementias
from Neuropathology
Kurt A. Jellinger
Institute of Clinical Neurobiology, Vienna, Austria
INTRODUCTION
With an increasingly elderly population the incidence of dementing disorders is
rapidly increasing in frequency and represents a major scientific, humanitarian,
and socioeconomic problem. Due to recent progress in genetic, molecular
biological, imaging, and neuropsychological techniques, and detection of disease
specific biological markers, the diagnosis and classification of these processes
have increased in accuracy. However, despite the establishment of diagnostic
guidelines and consensus criteria, a definite diagnosis still depends on neuro-
pathological examination of the brain at autopsy and, rarely, by biopsy. This is
particularly true for early stages of such processes, in which the validity of
clinical and neuroimaging criteria is limited. Studies of the brains of elderly
individuals without cognitive changes and in those with Alzheimer’s disease (AD)
and other dementias have provided a major input for progress in our under-
standing of brain aging and the lesion thresholds for mild cognitive impairment
(MCI) and initial stages of dementia (1–16).
The quality and pattern of brain lesions in aging and AD, their spreading
along well-defined anatomical pathways, and the consecutive biochemical
changes have been described and correlations with cognitive function and clinical
course have been performed. Recent insights into the cell biology of protein
misfolding and its consequences for neuronal (dys)function offer hope for a better
understanding of the underlying molecular processes (17–21). However, many
311
312 Jellinger
at a later age as amyloid burden increases, suggesting that Ab levels are not
a marker of memory decline that is related to disease progression (101–103),
whereas in the APPSLPS1KI mice massive CA1/2 neuronal loss correlates with
strong accumulation of intraneuronal Ab42 (104), and APP/tau double-tg mice
show accelerated Ab deposition, neurofibrillary degeneration, and neuronal loss
suggesting a reciprocal interaction between Ab and tau (105). In triple-tg
(3xTgAD) mice harboring PSEN1, APP, and tau (P301L) antigens and
progressively developing SPs and NFTs, synaptic dysfunction manifests in an
age-related manner before plaque and tangle pathology (96), suggesting that
plaque or tangle pathology contribute to cognitive dysfunction at later time
points. APP(SL) PSEN1 tg mice develop age-related synaptophysin-IR
presynaptic boutons within the hippocampus, which supports their role in
neurodegeneration (106). Clearance of the intraneuronal AD pathology by
immunotherapy rescues the early cognitive deficits on a hippocampal-dependent
task, strongly implicating intraneuronal Ab in the onset of cognitive dysfunction
(107). This AD model and APP(SL) PS1 K1 mice show massive CA1/2 neuronal
loss with intraneuronal and N-terminal truncated Ab42 accumulation (104). The
introduction of Ab into tau-tg mice has led to enhanced tau pathology with no
change in Ab deposition, further supporting the amyloid cascade hypothesis
(92,95,108). Tangle formation in tg mice expressing mutant human tau is
accompanied by extensive axonal and neuronal damage (109).
Ab Neurotoxicity
Amyloid proteins appear as a subgroup of misfolded proteins, where misfolding
leads to subsequent aggregation. This aggregation may be a generic property of
polypeptide chains possibly linked to their common peptide backbone that does
not depend on specific amino acid sequences. And, in fact, many proteins can,
in vitro, form amyloid-like aggregates, while in vivo, only 20 amyloid proteins
have been so far identified. Although misfolding and aggregation are quite well
studied in vitro, the last step of amyloid deposition, i.e., anchorage to the
extracellular matrix, cannot be so easily approached. Proteoglycans and serum
amyloid P component have nevertheless been identified as key elements involved
in extracellular deposition of amyloid (110).
Mounting data suggest that soluble Ab oligomers (protofibrils), intraneur-
onal and spherical aggregates of Ab42 (amylospheroids), rather than Ab fibrils,
may be the primary toxic species and the principal cause of synaptotoxicity
(111–118). Ab cytotoxicity is mediated by p38 inducing oxidative stress (119).
Soluble Ab oligomers ultrastructurally localize to cell processes and might be
related to synaptic dysfunction in AD brain (46). These soluble neurotoxins
[known as “amyloid b-derived diffusible ligands” (ADDLs) (120,121) and
protofibrils] seem likely to account for the imperfect correlation between
insoluble fibrillar amyloid deposits and AD progression. ADDLs are known to be
potent inhibitors of hippocampal long-term potentiation in vivo, which is
Early Diagnosis of Dementias from Neuropathology 319
a paradigm for synaptic plasticity, and have been linked to synapse loss and
reversible memory failure in tg mouse AD models (111), and binding occurs
predominantly at synapses (117). If such oligomers were to build up in human
brain, their neurological impact could provide the missing link that accounts for
the poor correlation between AD dementia and amyloid plaques. Oligomers in
AD reach levels up to 70-fold over control brains. Brain-derived and synthetic
oligomers exhibit the same striking patterns of attachment to cultured
hippocampal neurons, binding on dendrite surfaces in small clusters with
ligand-like specificity. Binding assays using solubilized membranes show
oligomers to be high-affinity ligands for a small number of low abundance
proteins. Current results confirm the prediction that soluble oligomeric Ab
ligands are intrinsic to AD pathology (121). Recent experiments have detected
the presence of ADDLs in AD-afflicted brain tissue and in tg models of AD. The
presence of high affinity ADDL binding proteins in hippocampus and frontal
cortex parallels the regional specificity of AD pathology and suggests
involvement of a toxin receptor-mediated mechanism. The properties of
ADDLs and their presence in AD-afflicted brain are consistent with their
putative role even in the earliest stages of AD (116). The strong correlation
between soluble Ab brain concentrations and severity of neural circuitry
dysfunction (122) and early aggregates of nondisease-associated proteins are
potent cytotoxics, whereas their final fibrillar aggregates (indistinguishable from
AD fibrils) are not (123). The presence of misfolded proteins in the ER triggers
a cellular stress response called the unfolded protein response (UPR) that may
protect the cell against the toxic buildup of misfolded proteins. Phosphorylated
(activated) pancreatic ER kinase (p-PERK), which is activated during the UPR,
was observed in neurons in AD patients, but not in nondemented controls and did
not co-localize with AT8-positive tangles. These data show that the UPR is
activated in AD, and the increased occurrence of p-PERK in cytologically
normal-appearing neurons suggests a role for the UPR early in AD neuro-
degeneration. Although the initial participation of the UPR in AD pathogenesis
might be neuroprotective, sustained activation of the UPR in AD might initiate or
mediate neurodegeneration (124).
On the other hand, recent data suggest that synapse-associated Ab is
prominent in regions relatively unaffected by AD lesions, and that amyloid
accumulation in surviving terminals is accompanied by gliosis and alteration in
the postsynaptic structure (125). In accordance, single systemic administration of
an antibody against Ab produces rapid improvement in behavioural performance
of APP-tg mice without affecting total amyloid levels, obviously because of
blockage of a diffusible Ab species (126). The binding of Ab to membrane lipids
facilitates Ab fibrillation which in turn disturbs the structure and function of
neuronal membranes (127). Fibrillary Ab deposition has been shown to be more
detrimental to neuronal circuitry than previously thought (128). Ab toxicity
mediated by the formation of APP complexes (129) but not by secreted Ab (130)
and mitochondrial dysfunction (131,132) induce oxidative stress-mediated
320 Jellinger
Figure 1 (See color insert.) Spreading pattern of (A) cytoskeletal/tau pathology and
(B) of Ab deposition. Source: From Refs. 218, 229.
as the basis of AD or enhances the tau pathology. For the more frequent sporadic
form of AD, the pathogenic trigger has not been unambiguously identified.
Whether Ab is again the main cause remains to be determined, although tau-tg
mice and tissue-culture models provide insight into the biochemical mechanisms
of tau aggregation and nerve cell degeneration (244,245). Chronic activation of
microglia, via the secretion of cytokines and reactive molecules, may exacerbate
plaque pathology and enhance the hyperphosphorylation of tau and the subsequent
development of NFTs. Suppression of microglial activity in AD brain has been
considered as a potential treatment of AD and may slow disease progression (246).
Recent results clearly indicate that Ab25-35, the peptide region to which the
cytotoxic properties of Ab can be assigned, interacts with the peptide region of tau
protein involved in microtubule binding; intracellular binding of Ab oligomeres to
soluble tau may promote tau phosphorylation. This interaction produces the
aggregation of tau peptide and the concomitant disassembly of Ab25-35, offering an
explanation for the lack of co-localization of NFTs and SPs in AD, and suggesting
the possibility that tau protein may have a protective action by preventing Ab from
adopting the cytotoxic, aggregated form (247).
Recent data indicate nonoverlapping but synergistic actions of both
pathologies in sporadic AD (248,249). APP dysfunction/mismetabolism inducing
Ab accumulation in the brain is believed to be the primary influence driving AD
pathology due to a failure of an autoregulation feedback reducing neuronal
activity and could contribute to cognitive decline in early AD and to disease
progression (41,250). Tau colocalizes with Ab42 and is induced by Ab42 in vitro.
Recent evidence suggests that plaques, NFTs, and caspases share a common
pathway (251). Caspases cleavage of APP and tau has been demonstrated in AD
(158,252,253). Ab accumulation triggers caspase activation which, in turn, leads
to caspase-cleavage of tau which is an early event that precedes hyperpho-
sphorylation in the evolution of AD tangle pathology (252,254,255). Caspase-
cleaved tau (Dtau) may initiate or accelerate the development of tangle pathology
(164,253). Its accumulation may represent a common pathway associated with
abnormal intracellular accumulation of tau or a-synuclein (AS) and may be less
dependent on the extracellular accumulation of Ab in non-AD dementias (256).
Dtau occurs early in the development of tangle pathology within AD brains and in a
tg mouse AD model, suggesting that caspase cleavage of tau plays an important
role in the development of NFT pathology. Alterations in tau phosphorylation and
cleavage by caspases have been reported in neuronal apoptosis. The presence of
activated caspase-6 in pre-tangles suggests that it occurs early and supports
previous studies demonstrating caspase activation in MCI but not in AD subjects.
While caspase-6-cleaved tau was found in NPs, NTs, and NFTs, active caspase-6
localized primarily to neurites (161). This is consistent with the hypothesis that
apoptosic-like mechanisms can damage synapses, axons, and dendrites, without
causing overt neuronal death. These results also lend support to the hypothesis that
the activation of apoptosis-like mechanisms may be involved in AD pathogenesis
(160,251). Caspase inhibition prevents tau cleavage without reversing changes
Early Diagnosis of Dementias from Neuropathology 327
!50 a 0 C C C
50–75 a 0 B C C
O75 a 0 A B C
PrP)
immuno-
histo-
chemistry
Key: K, lacking; G, rare; C, occasional; CC, moderate; CCC, severe.
Abbreviations: AD, Alzheimer’s disease; CBD, corticobasal degeneration; DLB, dementia with Lewy bodies; FTD, frontotemporal dementia; GCIs, glial
cytoplasmic inclusions; LBV/AD, Lewy body variant of Alzheimer’s disease; MSA, multiple system atrophy; PSP, progressive supranuclear palsy.
333
334 Jellinger
defined neuropathological criteria for DLB and the presence of LBs in many
cases at autopsy with non-DLB clinical presentations. The distribution of cortical
LBs in DLB does not follow the spread of NFTs (196), while the spreading
pattern of AS pathology with onset in the lower brainstem and progression to the
midbrain, dorsal forebrain, amygdala, limbic cortex, and final extension to the
neocortex is similar to that in sporadic PD (306). The late stages 5 and 6 of LB
pathology (involvement of sensory association and prefrontal, primary sensory,
and motor areas) suggest transition between PD and DLB (275,307,308). DLB
was classified into the limbic type and neocortical type according to the degree
of Lewy pathology, including LBs and LNs, and, on the other hand, into the
pure form, common form, and AD form according to the degree of Alzheimer
Early Diagnosis of Dementias from Neuropathology 335
(A) (B)
Morphological diagnosis n % n %
pathology including NFT and amyloid deposits by Braak staging (218). These
combined subtypes were lined up on a spectrum not only with Lewy pathology
but also with other DLB-related lesions including Alzheimer pathology, neuronal
loss in the SN, spongiform changes in the transentorhinal cortex, and LNs in the
CA2-3 region. There were some similarities in the extent of Lewy pathology
between PD and DLB, although Lewy pathology of PD was below the lowest
stage of Lewy pathology. In contrast, AD did not meet the stages of DLB Lewy
pathology, and there were also no similarities in other DLB-related pathologies
between AD and DLB. In addition, LBs of AD showed characteristics different
from those of DLB in the coexistence of LBs with NFTs. These findings suggest
that DLB has pathological continuity with PD, but can be pathologically
differentiated from AD. While this would suggest that DLB exists as a discrete
pathological entity, as with AD and PD (309), others showed no (310) or only
336 Jellinger
Brodmann
Cortical region area Anatomy Score
little difference between Parkinson’s disease with dementia (PDD) and DLB
(310a), in particular, more frequent involvement of the amygdala-limbic system,
and neocortex in DLB (308).
LB pathology may progress in a systematic fashion through the brain
regardless of the clinical phenotype. Except for some deviations in the severity and
distribution of lesions in substantia nigra pars compacta (SNc) and a more frequent
involvement of the CA2-3 region of hippocampus in DLB, these similarities
suggest morphological and pathogenetic relations between both disorders. DLB
is frequently associated with AD pathology of variable intensity and extent.
A subgroup with diffuse Ab deposition and neuritic AD lesions absent or restricted
to the hippocampus is referred to as “pure” DLB, while those showing significant
neuritic pathology that fits with the diagnosis of definite AD (281) have been
classified as LBV/AD (290,311). They occupy higher Braak stages of AD
pathology than age-matched controls, but lower stages than pure AD (290,312).
Among 96 autopsy cases, 62% were classified as “pure” DLB and 37% had severe
additional AD pathology (Braak stages 5 and 6). LBV/AD was present in 33% each
of the limbic and neocortical subtypes (276,313). “Pure” DLB shows no significant
differences in neocortical synapse density and synaptophysin immunoreactivity
compared to controls, while severe synapse loss comparable to AD is seen in LBV/
AD (314). Although neuronal loss in the cholinergic basal forebrain is consistently
found in DLB, PDD, and AD (1), early and more widespread cholinergic decline
differentiates DLB from AD (300), while LBV/AD shows an increase of cortical
M1 muscarinic cholinergic receptors compared to both AD and controls (315).
Early Diagnosis of Dementias from Neuropathology 337
Table 7b Modified Criteria of Dementia with Lewy Bodies, Cases Without Brainstem
Lewy Bodies Excluded
Maximum number of Lewy bodies per field Rating score for each region
0 0
1–5 1
6C 2
Region sampled
PDD, largely determined by the temporal order of symptoms) and also about
pathological findings (LBD, AD). The latter “categories” will need to be further
illustrated by details of lesion density and distribution (330). Thus, a practical
consensus for the diagnosis of DLB at present is not available.
Pick’s Disease
PiD is a rare variant FTD that accounts for 1–2% of all elderly dementia cases; its
classical type shows frontotemporal lobe and limbic atrophy with neuronal loss,
spongiosis, and gliosis including microglial activation, achromatic (Pick) cells,
and intraneuronal globose inclusions (Pib) in hippocampus, in particular in
dentate granule neurons, in cerebral cortex, and in selected brainstem nuclei.
Ultrastructurally, they are composed of straight and long-period twisted
filaments, made up of 3-repeat tau doublets (60 and 64 kDa) and a minor
Early Diagnosis of Dementias from Neuropathology 341
68 kDa band (332,378). Tau-positive glial inclusions, mainly in the white matter,
NFTs, and a network of dystrophic neurites differentiate PiD from FTD with
tau-negative astrocytes (379). In sporadic PiD cases, in addition to 3- repeat
tau deposits, isolated filaments formed from 3- and 4-repeat tau isoforms, and tau
phosphorylation-dependent and exon 10-specific epitopes are found. Thus,
accumulation of Pibs with 3- and 4-repeat tau pathology distinguish PiD from
other tauopathies (380) along with a lack of any association with the tau H2
haplotype (381). A novel presenilin 1 mutation with familial Pick-type tauopathy
without tau gene mutation and without Ab plaques (382), a familial FTD with
Pick-like pathology associated with Q336R mutation of the tau gene (383),
another phenotype with a missense mutation of S305N (384), and a family with
the R406W mutation and pathology consistent with NFTD (362) have been
reported recently.
Familial Tauopathies
Several genetically distinct groups of inherited FTD have been identified:
(1) FTDP-17; (2) FTD-MND linked to chromosome 3; and (3) FTD linked to
chromosome 3 (FTD-3). Tau mutations have been found in 25% of familial cases
of dominantly inherited FTD, but only in 4% of sporadic cases (389). Tau on
chromosome 17 is the only gene where mutations have been identified, while Ub
and tau inclusions have been found in the frontal cortex of patients from a Danish
family with FTD-MND that shows more diffuse cortical involvement than other
forms of FTD (390,391). Autosomal-dominantly inherited disorders linked to
chromosome 17q21–22 show diverse but overlapping phenotypes (392), with tau
pathology in neurons and glia, but no Ab deposits or other disease-specific brain
lesions (393). Two classes of tau mutations have been found in 10–40% of
familial FTD cases—those directly affecting the microtubule-binding sites of tau
and those that alter tau splicing (378,387). The majority of the currently known
tau mutations are located in exons 9 to 13. P301L mutation was detected in 11%
of familial FTD cases. The H1 haplotype was not overrepresented, but the P301L
342 Jellinger
1. 3-repeat tauopathies—PiD.
2. 4-repeat tauopathies—CBD.
PSP.
Argyrophilic grain disease (AGD).
3. 3- and 4-repeat tauopathies; tangle-predominant dementia.
4. DLDH.
5. Diseases with motor neuron disease-type inclusions (DMNDI).
Another classification distinguishes pathological subtypes of FTD (352):
† with tau-immunopositive inclusions (with and without Pib)—CBD
and AGD
† with Ub-immunopositive inclusions—FTD-MND/FTLD-U
† lacking distinctive histology (DLDH).
FTLD-U or FTLD-MND should be considered in the differential diagnosis
of progressive FTD with an akinetic-rigid syndrome and PSP (399). An
additional, recently described form is neuronal intermediate filament inclusion
disease (NIFID) (372,403), representing a neuropathologically distinct, clinically
heterogenous variant of FTD that may include parkinsonism or MND and is
distinct from other FTDs (405). Genetic alterations can give rise to phentoypes
more or less similar to any of the above. Tau mice recapitulate the key phenotypic
hallmarks of human tauopathies (404).
Early Diagnosis of Dementias from Neuropathology 343
Other Tauopathies
Hippocampal Sclerosis Dementia (with Tauopathy)
In a subset of elderly individuals, hippocampal sclerosis (HS) is the only
remarkable morphological finding (406–412). Its frequency in autopsy series of
elderly demented ranged from 0.4 to 26% (413), while it is almost never seen in
nondemented oldest-olds (414,415). HS shows a wide range of severity and
distribution, with damage of the hippocampus and subiculum ranging from
neuronal loss and gliosis to frank infarction. It is occasionally accompanied by
multiple small infarcts in other brain regions or leukoencephalopathy or both,
while only rarely such brains show additional AD lesions (410,413). The cause of
HS has been suggested to be due to occult hypoxic-ischemic episodes (408,409)
or as a sequela of limbic encephalitis (416). Classical HS dementia (HSD),
clinically being more similar to FTD than to AD (417), is morphologically
characterized by severe loss of neurons and gliosis in the hippocampal CA1
region and subiculum. The EC, temporal pole, inferior frontal neocortex, and
frontal pole may also be affected. Frequent tau-positive neurons and/or glial cells
in neocortex, basal ganglia, thalamus and/or limbic regions, resembling to FDTP-
17 and a mixture of 3- and 4-repeat tau isoforms, suggested the term “HSD with
taupathy” (HSDT) (413). “Pure” HSD (406), where no other cause of dementia
could be identified, shows morphological similarities to DLDH or ubiquitinated
neuronal inclusions, similar to those of MNDID (363,418), but with brain levels
of soluble and insoluble tau being normal, suggesting that “pure HSD” may
represent FTD and, in particular, its MNDID variant (419). In addition, several
cases had argyrophilic grains (413) that are also seen in argyrophilic grain disease
(AGD). Selective degeneration of the CA2 sector of the hippocampus is rare, but
when detected, it is associated with 4-repeat tauopathies, particularly AGD (420).
Argyrophilic Grain Disease
This neurodegenerative disorder of the elderly is seen in about 5% of demented
patients and in subjects progressing from MCI to dementia (421–423). It may or
may not be associated with a cognitive decline in the presence of only moderate
amounts of AD-related pathology (424) and may be associated with anxiety,
restlessness or depression (422) or may present with FTD (425). Morphologically
AGD displays abnormal argyrophilic grains (AG), coiled bodies, and “pre-
tangles” mainly in the limbic system (medial temporal lobe, amygdala) and
insular cortex containing 4-repeat tau deposits with isoforms of 64 and 69 kDa
(380,426–428). It shows genetic differences to other tauopathies (422,429). AGD
is frequently associated with AD-related changes, low Ab-load, and mild to
moderate NFT pathology. A recent comparative morphological study showed
that demented AGD cases showed lower stages of AD-related pathology, but
higher ones than non-demented AGD patients. AGD associated dementia was
associated with Braak stages 2–4 and Ab phases 2–3, while those stages were not
associated with dementia in the absence of AGD (424). Another recent study
344 Jellinger
Vascular-Ischemic Dementia—Vascular
Cognitive Impairment
Vascular dementia (VaD) is a heterogeneous group due to lesions caused by
different pathophysiological mechanisms and with different combinations of
brain pathologies. It is therefore necessary to identify the various types of
vascular brain lesions in order to correlate correlation with clinical symptoms
and for diagnostic purposes to search for risk factors and therapeutic strategies.
Dementias related to cerebrovascular disease (CVD) and ischemic brain
damage were previously considered to be the second most common type of
dementia (432–436) in the Western world after AD and DLB, with a possible
incidence of 8–15% (415). Clinical diagnostic criteria for VaD–ICD-10 [WHO
93 (437)] exists: DSM-IV (438), the SCADDTC (439), and NINDS-AIREN
criteria (440), supported by the Hachinski Ischemic score (HIS) (441,442),
show variable sensitivity (average 50%) and specificity (range 64–98%)
(443–446) and may exclude a number of subjects with VaD (447). NINDS-
AIREN neuroimaging criteria have been found not to distinguish stroke
patients with and without dementia (448). In an interobserver study, use of the
operational definitions for the NINDS-AIREN criteria improved agreement but
only for already experienced observers (449), whereas in another study none of
the currently used clinical criteria identified the same group of incidentally
demented subjects (450). On the other hand, integration of neuropsychological
and neuroimaging data was suggested to be sufficient for the diagnosis
of subcortical VaD (451). The classification may be based on: 1) primary
vascular etiology, 2) primary type of ischemic brain lesions, 3) primary
location of the brain lesions, and 4) primary clinical syndrome. Subcortical
ischemic VaD is an example of such a subgroup. Vascular cognitive disorder
(VCD) has been recently limited to cases without dementia, i.e., vascular
Early Diagnosis of Dementias from Neuropathology 345
left-side infarcts, strategic infarcts, and white matter lesions appear to be the main
predictive factors of PSD (488). During a four-year follow-up the incidence of
PSD increased gradually, shifting from an initial AD-type picture to a VaD-type
later (489). Many patients with lacunar infarcts have a good functional outcome
at five years. For older patients, for patients with an initial severe stroke, and with
additional vascular risk factors, however, the prognosis is more severe, with an
increased risk for mortality, stroke recurrence, and physical and cognitive decline
(490). Dementia showed a correlation with widespread small ischemic lesions
throughout the CNS, mainly lacunes, microinfarcts, and hippocampal injury, and
much less with larger infarcts, many brains showing more than one type of CVLs,
although in cognitively normal aged controls similar lesions were present
(297,415,460,461,491). The size of WMLs in the elderly may progress with time
and may relate to clinical symptoms (492–494) and thus can be regarded as a risk
factor for cognitive impairment (495), whereas others may show territorial
infarcts and older age as predictors of cognitive impairment in the first year after
stroke (496). They impair frontal functions regardless of their location (497),
are associated with cortical more than entorhinal and hippocampal atrophy (498),
and increase the risk of dementia (499), particularly in patients with lacunar
Early Diagnosis of Dementias from Neuropathology 347
infarcts (500,501), but WMLs and lacunes may be independently associated with
cognitive dysfunction (502). Other studies showed that subcortical vascular
disease on computed tomography (CT) is frequent in older patients with MCI,
but does not appear to be associated with the severity of cognitive deficits (503).
Postmortem detection of WMLs by MRI was less sensitive than pathology (504).
These and recent neuroimaging data indicate that subcortical lacunes, WMLs,
and multiple disseminated microinfarcts resulting from small vessel disease
that damage structures of the prefrontal-subcortical circuits (505) are the most
common pathological features of VCI/VaD, whereas large infarcts are less
frequent (506).
Mixed Dementias
Mixed type dementia (MD) is characterized by combined pathologies of both AD
and VaD or other dementing disorders, but the distinction between these two
pathological diseases is controversial (356,434,460,507–509). Recent emphasis on
co-morbidity of AD and CVD (510–512), the link between AD and atherosclerosis
(513), cognitive impairment associated with amyloid angiopathy (514,515),
significant cerebral microvascular pathology (516,517), and deficient clearance
of Ab across the BBB in AD (518,519) all indicate that vascular disorder is an
important feature of the chronic neurodegeneration in AD. Therefore, neurovas-
cular dysfunction could have a major role in the pathogenesis of AD (140,520).
ApoE E4 and E2 with its potential amyloidogenic role may be responsible for some
of the microvascular changes found in AD (521). Many patients with dementia
have radiological and neuropathological features of AD and VaD, with the
classical NFTs and Ab plaques of AD together with the cerebral infarcts of VaD.
A close relationship between AD and VaD has been suggested since the age-related
changes in cerebral blood vessels that are the basis of CVD and VaD may also be
responsible for the failure of elimination of Ab from the brain in AD (522).
Neuronal overexpression of human APP renders the brain more vulnerable to
ischemic injury and describes the factors that are involved in increased neuronal
Early Diagnosis of Dementias from Neuropathology 349
susceptibility to ischemic stroke (523). Criteria for the clinical diagnosis of mixed
dementia are variable (439,440,524–528), and it has been questioned whether
mixed dementia really exists as a separate entity (529,530). Generally accepted and
validated histopathological criteria for the diagnosis of MD are currently not
available, and its true frequency is unknown. Its prevalence rate in autopsy series
showed a wide range from 2 to 56% in retrospective studies and 2.9 to 54.2% in
prospective studies with means around 15% (13,460,461,477,531). Complicating
the studies, many persons exhibit neuropathologic changes similar to AD, VaD, or
mixed dementia, but do not meet clinical criteria for dementia (2,532).
Patients with AD frequently have other concomitant pathological lesions
(533) and strokes are common and increase with age (534). Elderly people with
silent brain infarcts have an increased risk of dementia and a steeper decline in
cognitive function than those without such lesions (535). The severity of cognitive
impairment was correlated to the total volume of infarcts with impact on lesions in
limbic and medial association areas, frontal cortex, and white matter (536). In the
Nun study, patients with autopsy-confirmed AD and CVLs had a higher prevalence
of dementia than those without infarcts (24). The risk of being demented was
20-fold higher in subjects with AD and lacunar infarcts, but much lower and non-
significant when a large territorial infarct was present (525,527,537). Other
studies, however, showed lower Mini-Mental State Examination (MMSE) scores
in AD patients without than in those with concomitant CVLs, the latter having
a significantly more frequent history of strokes (Table 10). The presence of
cerebral infarcts in AD increases significantly with age, but has little influence on
the clinical features, and cannot be predicted from common vascular risk factors.
In spite of a trend, there are no major differences in neurodegenerative lesion load
(Ab and NFT) between AD and AD with cerebral infarcts groups, except when
cerebral infarcts are located in the temporal lobe (including hippocampus) where
AD pathology was frequently lower, suggesting that this location may be
important in the pathophysiology of mixed vascular and AD dementia (538).
In a population-based study in the UK among 209 autopsies of elderly subjects,
48% being demented, 78% with CVD, and 70% AD pathology, the proportion of
multiple vascular pathology was higher in the demented group (539), in which only
21% showed “pure” AD (471). In a health maintenance organization dementia
registry, only 36% of patients had AD and no other findings, while 45% had
pathologically definite AD plus coexistent CVLs, and 22% had AD plus DLB
features (540). Comparison of 186 AD cases and 13 individuals with no cognitive
impairment (NCI) suggested that neocortical core plaques were more related to
dementia severity than the density of total SPs and NPs, while diffuse SPs were not.
In those patients with infarcts, hemorrhages, or PD, NFT and SP densities were
lower despite the presence of dementia (6). Among 333 autopsied men in the
Honolulu-Asian Aging Study (120 demented, 115 marginal, 96 normal cognition),
24% of all dementias were linked with CVLs, and dementia frequency more than
doubled with coexistent CVLs (45% vs. 20%). Findings suggest CVLs are
associated with a marked excess of dementia in cases with low neuritic plaque
350 Jellinger
mainly related to the severity and extent of AD pathology (Table 10). Hence, the
combination of two or more pathologic processes may influence the severity
of cognitive deficits and represents a major diagnostic challenge.
Guidelines for the Diagnosis of Mixed Dementia
At present, there are no generally accepted and validated clinical or neuro-
pathological guidelines for the diagnosis of MD. Criteria for AD and VaD are of
limited value for the diagnosis of MD, and more distinct criteria for this diagnostic
category are necessary.
Other Dementias
The pathological findings and diagnostic criteria for other types of dementia will
not be discussed here (292,566).
the hippocampal area CA1 in older ones (73–89 years), while MCI cases had
more NFTs in the same areas. Severely demented patients had large numbers of
NFTs in neocortex, while only part of the nondemented group showed a few
primitive plaques, and all the MCI to mildly demented groups had large numbers
of primitive or mature SPs. Unlike NFTs they were not considered a feature of
aging up to 80 years (14).
In another cohort, all nondemented individuals had NFTs in (para)-
hippocampal areas, their mean density showing an exponential increase with age,
even in the absence of SPs. A group of nondemented elderly showed widely
distributed DPs and NPs in the neocortex and limbic areas; those with NPs were
suggested to represent preclinical AD, CDR 0.5 in this particular study being the
threshold of very mild dementia. NFTs in EC, suggested to be the earliest stages
of AD, closely paralled cell loss in very mild demented subjects, but NFT
formation was not associated with age-related cell loss, which was not found in
nondemented subjects (237,238,589).
In another study, no clear differences in the frequence of NFTs between
cognitively normal persons and CERAD “possible” AD cases were seen. Cogniti-
vely normal individuals had Braak stages 1 or 2 while “possible” AD cases had
Braak stages 1–3 (590).
In a further study, diffuse SPs were seen in all cases with moderate dementia
(mean age 60 years), in 65% with mild dementia, and in 58% with questionable or
no dementia. SP density increased as a function of dementia severity and was
correlated with NP and NFT formation (4). Diffuse SPs may not be part of normal
aging but instead represent presymptomatic AD. The high density of DPs in
controls just at the threshold of detectable dementia is consistent with the
hypothesis that Ab deposition is an initial event in development of AD (237).
This was confirmed in a 74-year-old female with advanced AD and
her nondemented 47-year-old daughter, both dying from homicidal strangula-
tion (591). While the mother showed fully-developed AD pathology, the
daughter’s brain revealed only perineuronal deposition of diffuse Ab in cerebral
cortex and abnormalities of the endosomal lysosomal system, without NFTs
or glial changes, suggesting that amyloid deposition and endosomal-lysosomal
changes are early events in late-onset AD that precede the onset of dementia.
No genetic information about this AD family was available.
Elderly cognitively normal individuals had no or only few neocortical SPs
or NFTs in limbic areas (Braak stages 0–2). While there was no significant
relation between age at death and density of limbic or neocortical NFTs, the rate
of cognitive change was correlated with NFT burden in the neocortex and to
a lesser degree in limbic areas. NFT density decreased in order from the EC
through amygdala, subiculum, and CA1, to infratemporal region. In patients who
became demented, more severe NFT and SPs were seen (3).
These and data from the Vienna Prospective Dementia Study (mean age
81.7G8.6 years) confirm the significant negative correlation between neuropsy-
chological status and Braak stages in patients without other pathologies (509).
Early Diagnosis of Dementias from Neuropathology 355
Neocortical Braak stages 5 and 6 were mainly, but not exclusively, seen in
severely demented individuals, while limbic stages (2–4) were associated with
a wide range between the cognitively normal to overt dementia (Fig. 2). Similar
correlations have been observed in both younger (219,223,274,298) and
older subjects (224,592,593), indicating a “gray” transitional zone of mild to
moderate dementia showing a wide range of AD lesions (Fig. 3). These data
support a continuum in which AD is infrequent in healthy, cognitively stable
seniors, and preclinical pathology precedes cognitive impairment. All MCI
patients had a tau pathology, but not necessarily Ab pathology, whereas not all
patients with tau pathology did not have MCI (219).
Among 11 MCI cases (mean age 89 years; CDR 0.5) five patients showed
frequent NFTs and NTs in limbic regions and moderate DPs with sparse NPs and
NFTs in neocortex, but were insufficient for the diagnosis of definite AD, while
the others showed extensive tau pathology in the medial temporal lobe suggesting
very early AD (594).
In the Religious Order Study (ROS) fewer NFTs and NTs in the
parahippocampus were seen in cognitively unimpaired subjects compared to
MCI and AD (10,295). NFT density within peri- and entorhinal cortex correlated
with measures of episodic memory. NTs were not correlated with any cognitive
ability. The NT burden increased from normal to MCI and then decreased in AD.
Major findings were (1) that granulovacuolar lesions and NFTs correlated with
measurements of episodic memory; (2) NTs precede NFTs that in turn precede
Figure 3 Column scatter plot of Braak stages versus Clinical Dementia Rating (CDR)
scores in 100 oldest-old people (mean age at death 92.5G2.5 years).
NPs; and (3) conformational tau changes emerged as a result of in situ reactivity
to different antibodies, reflecting its structural status in NFTs that correlated with
memory deficits (216). The authors presented a model for PHF degeneration
beginning with NT formation, followed by NFT development and ending with
appearance of NPs. Nondemented subjects showed the hierarchical pattern of
NFTs, while the distribution of NPs varied among individuals. MCI cases had
higher NFT densities than the cognitively normal in the medial temporal lobe,
this being related to performance on memory tests, while amyloid SPs
showed no correlation (11). It seems that the progression from Braak stages I
and II to stage III, which may or may not be associated with signs and symptoms
of mild AD or MCI, takes decades to occur and is presumably critical for
developing AD.
The transition to clinical dementia (CDR R1) was associated with PHF-tau
pathology in areas A9/10, A22, A23, and A39. A9/10 represents a prefrontal
cortical network that provides “executive control” over complex goal-directed
behaviors, while the other areas provide afferents to other frontal systems and
have been implicated in early AD (595). Executive MCI in the absence of
dementia has been suggested to be associated with decrease of soluble Ab in
the frontal cortex (596) and with frontal and anterior cingulate tau and Ab load
that may be early features of the frontal variant of AD (597).
Among 162 older Catholic clergy (mean age 75.8 years; 31 MCI, 57
cognitively unimpaired, and 53 clinical AD), nearly all brains had some AD
pathology. Its average level in MCI was significantly higher than in controls and
significantly lower than in AD. The occurrence of infarcts and LBs showed no
difference between MCI and the two other groups. This suggests that patients
with MCI show a level of Alzheimer type pathology that is significantly different
Early Diagnosis of Dementias from Neuropathology 357
from both AD and cognitively unimpaired individuals. LBs were usually not
related to MCI (16), although MCI can evolve into DLB in some cases (598).
Similar findings were reported recently from the ROS: among 180 participants
(60 normal, 37 MCI, and 88 demented), nearly all had some AD pathology,
cerebral infarctions were present in 35.3%, and 15.6% had DLB. Persons with
MCI were intermittent in terms of Braak stage, CERAD, and NIA-RI
neuropathologic criteria for AD compared to the other two groups, and the
relationship between cognition and AD pathology did not significantly differ
between the three groups which also showed intermediate levels of cerebral
infarction, while only 8% had LB pathology (583).
In the Nun study, Braak stages 1 and 2 showed the greatest variation in
cognitive profile, while in other reports stages 3 and 4 seemed to be the most
diverse in terms of cognitive deterioration. Although stages 1 and 2 have been
described as “clinically silent” (218), some subjects in these stages already had an
identifiable cognitive impairment. Only cognitively unimpaired subjects were
classified as either Braak stage 0 or 2–3, while those with MCI showed all Braak
stages (24), but others found no or little pathological differences between persons
with MCI and cognitively normal individuals (25), and between MCI and
AD (216). Of the cognitively unimpaired individuals, 87% showed allocortical
NFTs, only 37% displayed neocortical NPs, 19% had hippocampal NPs, and none
exhibited neocortical NFTs, though the latter were not detected in 10% of AD
brains or in nearly 50% of mild AD. Although NP and NFT density increased
with dementia severity, significant differences emerged for NFTs alone. The
increase in NPs and NFTs, in patients with even mild AD compared to normal
controls, suggests that both lesions are associated with the earliest symptoms of
AD (566a,599). Hence, NFT and NP pathology may constitute pathological
substrates for memory loss in AD but also in normal aging and MCI.
However, demonstration of a distinct sequence of Ab deposition suggests
that nondemented individuals with Ab or NPs may also represent early stages
of AD (229). An asymptomatic prodromal stage appears to be associated with
increasing Ab in the brain. The initial symptoms of MCI may develop when
a threshold level of neuronal and synaptic loss is reached in the EC and
hippocampus concurrent with NFT formation and gliosis. Subsequent disease
progression and conversion to clinical AD are associated with progressive
neuronal and synaptic loss, and NFT formation on a background of elevated
Ab that reaches a “ceiling” early in the disease. NFT formation and synapse loss
continue throughout the disease course (270). Dementia conversion rates increase
as cognitive impairment increases, and the likelihood of subsequent conversion
of MCI to dementia is usually related to the severity of local AD pathology (24),
particularly in the ventromedial temporal lobe (566a).
In general, neuropathology data do not permit extrapolation in terms of
prediction of progressive cognitive decline in individual cases. However,
respondents with a final MMSE score in the “medium MMSE” group, analogous
to but not identical with clinical MCI, show indices of AD pathology which
358 Jellinger
appear intermediate between high and low performing groups: namely a paucity
of neocortical tau pathology but with more advanced temporal lobe pathology
than high performing patients.
cortex in AD, while chromogranin A was less reduced. This altered availability
may be responsible for impaired transmission and reduced function of dense core
vesicles (623,624). Loss of synaptophysin mRNA in NFT-bearing neurons in the
hippocampus (625) and defects in expression of genes related to synaptic vesicle
trafficking in frontal cortex of AD occur as well (626).
Caspase 3, a marker of apoptosis, is activated in the parahippocampal gyrus
in subjects with MCI and low CERAD/intermediate Braak scores; caspase-like
immunoreactiviy has a tangle-like appearance and co-evolved with PHF
pathology in neurons, suggesting activation of apoptosis and of caspase-cleaved
APP occurs very early in the medial temporal lobe in aging and AD (627,628).
Mitochondrial defects in AD include COX-deficient hippocampal neurons that do
not appear to be prone to apoptosis or directly participate in the overproduction of
tau or Ab. They are likely to contribute to the overall CNS dysfunction and
possibly cause neurodegeneration via mechanisms other than apoptosis (629).
Microarray analyses of hippocampal gene expression revealed a major
transcriptional response comprising thousands of genes correlated with AD
markers. There was up-regulation of many transcription factor/signalling genes
regulating proliferation and differentiation, tumor suppressors, glial growth
factors, protein kinase A modulators of adhesion, apoptosis, lipid metabolism,
initial inflammation processes, and down-regulation of protein folding/transport,
some energy metabolism, and signaling pathways. These findings suggest a new
model of AD pathogenesis in which a genomically orchestrated up-regulation of
tumor suppressor-mediated differentiation and involution processes induces the
spread of pathology along myelinated axons (630).
behavioral studies support the possibility that aging reveals the vulnerability of
an abnormally regulated cortical cholinergic input system. Its decline and the
decline in cognitive functions are further accelerated as a result of interactions
with APP processing (634). Extracellular Ab aggregation may affect cholinergic
terminations prior to progression onto other neurotransmitter systems (635), and
there may be relations between calbindin-D-28k, FADD expression, and
phosphorylation of tau within the basal forebrain (636), but the interaction
between cholinergic dysfunction and Ab are not fully clarified (637,638).
Cognitively normal subjects and early stage AD (average Braak stage !2)
in plaque-containing cases showed a significant decrease of ChAT activity
(71–79.5%) compared to plaque-free cases, and in inferior temporal cortex,
ChAT had an inverse correlation with Ab concentration (639), suggesting
a preclinical onset of the cholinergic deficit in AD (640). NFTs and AT8
immunoreactive neurons in the basal forebrain were seen even in cognitively
unimpaired subjects, but the percentage of tau-positive basal forebrain neurons
was greater in MCI cases and showed a significant correlation with memory
scores (27). These results and reduction of cortical AChE activity (640a) indicate
that the cholinergic deficit occurs at an early stage of AD before the onset of
clinical symptoms. Whereas both the subicular cholinergic fibers and basal
forebrain neurons were lost in AD, no cholinergic deficit was found in the temporal
cortex in “pure” VaD (641), and subicular cholinergic fibers are unimpaired in
Binswanger’s disease (642).
Changes in some cholinergic basal forebrain markers, e.g., the high affinity
TrkA receptor, but not others [e.g., cortical ChAT activity, the number of ChAT
and vesicular acetylcholine (ACh) transporter-immunoreactive neurons], suggest
specific phenotypical changes, but not the frontal neuronal degeneration that
occurs early in cognitive decline. APOE E4 and E2 genotype, a predictor of
cholinergic deficits (643), is accompanied by lower metabolic activity in the
basal forebrain neurons in AD patients and also in aged controls as indicated by
the size of the Golgi apparatus and may represent a risk factor for cognitive
impairment (644). Others saw no correlation between APOE E4 and E2 and
cholinergic decline (ChAT activity) in AD, while the presence of two E4 alleles
was an important determinant of both NP and NFT accumulation in the
neocortex. By contrast, a strong relationship between APOE E4 and E2 alleles
and decreased neocortical NP counts suggests a putative protective role for E2 in
AD (645). However, the fact that cholinergic neuron numbers and brain ChAT
activity are not altered in MCI suggests that compensatory mechanisms occur in
the remaining cholinergic perikarya that normalize cortical ChAT levels.
Reductions of cortical TrkA in prodromal AD may be one of the earliest signs
of subtle neurodegeneration in the basal forebrain system leading to the inability
of this system to utilize NGF and may have important consequences for
cholinergic basal forebrain function during the transition from MCI to AD (646).
The accumulation of proNGF in the basal forebrain cholinergic target zones and
the lack of retrograde signals are crucial for nuclear transcription of genes
Early Diagnosis of Dementias from Neuropathology 363
mediating cell survival. Increases in cell cycle proteins may be a response to the
lack of cell survival signals, suggesting that basal forebrain neurons undergo
degenerative events during the prodromal stages of AD (647). Progressive loss of
cortical acetylcholinesterase (AChE) activity, indicating a progressive dysfunc-
tion of the ascending cholinergic system, correlated with cognitive decline (648).
A significant reduction in the number of basal forebrain p75 neurotrophin
receptor-immunoreactive neurons was seen in MCI (38%) and mild AD
(43%). Its correlation with performance in the MMSE, some tests of working
memory and attention, and other data lend support to the hypothesis that MCI is a
preclinical stage of AD (649).
While strong correlations have been found between the density of NFTs in
the hippocampus and neocortex with the degree of cholinergic pathology and
dementia (650,651), many elderly subjects with MCI or early/very mild AD do
not display deficits in ChAT activity (297,652), or show an upregulation in
hippocampus and frontal cortex (9,647). A significant elevation of hippocampal
ChAT in MCI was found selectively in the limbic (3/4) Braak stages which may
reflect a compensatory response to the progressive denervation of the
hippocampus by lost EC input (653). Hippocampal upregulation was no longer
present in mild AD cases compared to cognitively unimpaired subjects, while
severe AD showed markedly depleted ChAT levels. Cognitively normal
individuals, MCI and mild-moderate AD, showed a positive correlation between
hippocampal ChAT activity and progression of NP pathology in this area. In MCI
and mild AD, ChAT activity was normal in inferior frontal, superior temporal,
and anterior cingulate cortex (9). This suggests that earliest cognitive deficits,
e.g., short-memory loss in MCI, do not involve cholinergic deficits, but more
likely relate to disrupted entorhinal-hippocampus connectivity. Increased medial
temporal NFT correlates strongly and better than hippocampal ChAT activity
with impairment of episodic memory (654). Increased cholinergic activity during
the early stages of AD may contribute to synaptic scaling, but the mechanisms of
these changes are unknown (655). No changes in cortical or hippocampal levels
of NGF were found in the brains of MCI individuals, and they did not correlate
with an increase in ChAT activity in these regions, suggesting that brain NGF
levels appear sufficient to support cholinergic plasticity (656). In vivo studies in
mild to moderate AD showed significant reduction of ACh levels in amygdala
and neocortex, whereas ACh activity and glucose metabolism appeared
preserved or even increased in the basal forebrain. This suggests that changes
in the cholinergic system are an early and leading event in AD rather than the
consequence of neurodegeneration of basal nuclei (657). Cortical AChE activity
is more robustly associated with functions of attention and working memory
compared to performance on primary memory tests in AD (658).
While early-onset AD patients show a wide range of expression levels of
the hippocampal cholinergic neurostimulating peptide (HCNP) precursor protein
mRNA, in late-onset AD it is progressively decreased in hippocampus but not in
the dentate gyrus (651). Since it stimulates the production of ChAT, this low
364 Jellinger
Alzheimer’s Disease
According to the guidelines of an international consensus group, a biomarker for
AD should detect a manifestation of the fundamental neuropathology and be
validated in morphologically confirmed cases. Its sensitivity for detecting AD and
its specificity in differentiating AD and other dementias should exceed 80%.
Ideally, a biomarker should also be reliable, reproducible, non-invasive, simple
to perform, and inexpensive. Of particular interest is its ability to detect
the disease at the earliest possible stage, i.e., before significant irreversible disease
progression has occurred. Biomarkers are potentially useful to study disease
pathogenesis in vivo and to study presymptomatic AD (725).
The relationship of functional neuroimaging to early morphological changes
are reviewed in the respective chapters (see Chapters 6 and 8) (726). They appear
superior to cognitive testing for early diagnosis of AD (727), but combined
evaluation of neuropsychological, structural, and functional imaging offers greater
predictive value (580). High resolution magnetic resonance imaging (MRI)
showed that the volume of frontal grey matter was strongly associated with age,
less with occipital grey and white matter, while the reduction in hippocampal
volume (727a) was best modelled as a cubic regression model. Common changes
in brain morphology as observed by cerebral MRI are associated with diminished
cognitive functions in elderly individuals (728). Cerebral atrophy seems to closely
parallel the distribution of neuritic pathology (and less the distribution of SPs).
Significant correlations were observed between antemortem hippocampal
volumes and both dementia severity and the density of hippocampal
neurofibrillary tangles at autopsy. Total cerebral volumes, in contrast, were
significantly correlated with the density of hippocampal SPs. This data suggests
that hippocampal volume assessed in living subjects with probable AD is both a
good marker of dementia severity and of the AD disease process (729). Voxel-
based morphometry and automated brain volumetry showed a high discrimination
accuracy between AD and controls, and could open up new possibilities for
early diagnosis of AD (730,731). MRI shows differences in Braak stages and
hippocampal volume between normal individuals and mild AD (601,732) with
increased changes related to the duration of disease and may predict AD pathology
(733). Early predictors of onset of AD in nondemented elderly subjects and of
progression of amnestic MCI to AD include increased annual atrophy rate in
the medial temporal lobe (734), inward deformation of the left hippocampal
surface in a zone corresponding to the CA1 subfield (735), atrophy of the
hippocampus (736), in particular of the EC (737), whole brain MRI spectroscopy
(738), and reduced hippocampal metabolism (739–741). Findings from the
Nun study indicated that both delayed recall measure and neuritic AD pathology
in the CA1 region reflect associated hippocampal atrophy (742,743). Memory
complaints in patients without any cognitive impairment were associated with
smaller left hippocampal volumes and more depressive symptoms (744).
As a result of neuron loss in the right amygdala-hippocampus complex it is
Early Diagnosis of Dementias from Neuropathology 369
Large numbers of NPs were strongly associated with lower CSF Ab42 levels
that may reflect processes implicated in amyloid pathology (783). Studies in 106
patients with dementia and four NCI persons confirmed the association between
elevated CSF p-tau premortem and the pathological hallmarks of AD, indicating
that elevated CSF p-tau levels strongly support a diagnosis and may be helpful in
distinguishing AD from other dementing disorders (784). A reduced ratio of Ab42
to total Ab in CSF of AD patients was consistent with previous clinical reports, but
because of the broad range of Ab values provided no meaningful additional
diagnostic information when tau was elevated. Meta-analysis of CSF levels in a
large number of AD cases (only 23% confirmed at autopsy) and controls showed
significant decrease of Ab42 and an increase of tau (785). A small autopsy cohort
showed a significant decrease of postmortem CSF melatonin in aged individuals
with early AD changes in the temporal cortex, suggesting that this may also be an
early event in the development of AD (786). In view of diverging data between
in vitro and postmortem CSF, this study awaits confirmation. Although CSF tau
and Ab42 may not be specific for AD, their evaluation combined with
neuroimaging may improve diagnostic specifity (787). Quantitative proteomics
of CSF from AD patients compared to age-matched cotrols, as well as from other
neurodegenerative diseases, will allow us to generate a roster of proteins that may
serve as specific biomarker panels for AD and other geriatric dementias (788).
With regard to plasma Ab40 and Ab42 levels, these may increase with
age but are not specific for MCI or sporadic AD (789). Reports show these to
have been elevated before and during the early stages of AD though then decline
thereafter (790). These findings may be associated with APOE E4 (791) and may
be a risk factor for microvascular damage (791a), although there are no reports of
any neuropathological confirmation of this data. The same findings are evident for
APP in platelets and other peripheral marker for sporadic AD (792). Morover,
plasma Ab42 levels are highly influenced by concomitant medications (793).
Salivary AChE activity may prove to be a useful marker of central
cholinergic activity (794). It is important that any future studies on biomarkers
are long-term prospective investigations with the aim of confirming the diagnosis
pathologically (795).
Frontotemporal Dementias
Based on the severity of gross atrophy in FTD, four progressive stages were
identified (822): stage 1: initial mild atrophy in the orbital and superior medial
frontal regions, and hippocampus; stage 2: atrophy of the other anterior frontal
regions, temporal cortices, and basal ganglia; stage 3: involvement of all
remaining tissues on coronal slices; stage 4: marked atrophy in all brain areas.
Severe atrophy of the frontal lobe, amygdala, and hippocampus in stage 2 suggests
that they are among the earliest affected in FTD (823). Comparison of PID, DLDH,
and FTD-MND found no differences in these subtypes, suggesting that disease
372 Jellinger
stage rather than FTD subtype determines the pattern and extent of neuronal
degeneration (824). This scheme provides the framework to determinate
correlates relating to disease progression, e.g., with neuroimaging data (825–828).
FTD revealed elevated CSF tau (though lower than in AD), high APOE E4
and E2 frequency, and decreased Ab42 levels (829). However, these values are in
the intermediate range between FTD, AD, and other dementias, and are not useful
for the diagnosis of FTD (830,831), and others showed significant reduction of t-tau
levels in CSF (832). CSF t-tau was not increased in FTD patients with tau mutations
(833), while p-tau 231 and p-tau 181 yielded excellent distinction between AD and
FTD (834). It could be useful to develop techniques to study tau-isoform
patterns in CSF, because they are disease specific when measured in brain
homogenates (835). Unfortunately, none of these CSF studies was confirmed by
postmortem studies, and a sensitive and a specific biomarker enabling differen-
tiation of FTD from other conditions is still lacking.
Vascular Dementias
Previous neuroimaging studies in vascular cognitive impairment (VCI) have been
partly contradictory (452,836). Independent correlates of post stroke dementia are
the combination of infarct feature (volume of infarcts in middle cerebral artery,
higher frequency of left-sided infarcts), extent of WMLs, medial temporal atrophy,
and host factors (836). The volume of functional tissue loss may be more important
than of total tissue because it also includes the effect of deafferented cortex
(837,838). Absence of CVLs on CT and MRI is evidence against a vascular
etiology of dementia, which is in agreement with neuropathological studies
(460,461,477). Although the hippocampus is less affected by subcortical CVLs,
dementia due to microvascular pathology showed significant hippocampal
neuronal loss (478,839), and hippocampal atrophy may increase the development
of post-stroke dementia (840). To date, however, it has not been possible to
determine the specific amount of WML necessary to cause cognitive impairment,
but it has been found that even relatively mild WMH can have deleterious effects
on cognition (472). A standardized assessment of subcortical CVD on CT films
can be combined into a unique score that is in good agreement with
neuropathology. This supports the validity of the CT-based visual rating scale
as a valid tool to detect subcortical vascular changes in elderly persons (841).
MRI studies confirmed that infarcts located in strategic areas have a role in
cognitive impairment, and their quantification correlates with neuropathological
findings (842). There is a substantial overlap between cases classified as AD by
NINCDS/ADRDA, and VaD by modified SCADDTC criteria. The substantial
contribution of vascular disease would be missed without inclusion of MRI.
Treatment of risk factors for VaD could have an important impact on the
incidence of dementia (843).
Studies of cerebral blood flow (CBF) and metabolism, in general, failed to
identify diagnostic features of VaD and to provide a robust, easily applicable
diagnostic technique because of the heterogenous pathology underlying dementia.
Early Diagnosis of Dementias from Neuropathology 373
lesions (533). Among the most frequent coexisting pathologies are CVLs with an
incidence of 18–80% (460,461,463,539,551,553,554), followed by LB pathology
in 12–20% (275,540). Cortical LBs have been observed in 7–71% (334,335,850),
and AS pathology in the amygdala and other brain areas in 15–60% of sporadic
AD cases (275,332,338,339). The pathogenesis and clinico-pathological impact
remains to be determined. While the APOE genotype usually shows no differences
between AD and vascular disease, cerebral infarcts in regions typically destroyed
by AD (e.g., limbic areas and the thalamus), and cortical LBs which frequently
occur in both AD and DLB, all may exacerbate the clinical manifestations of AD
and thus contribute to the heterogeneity of the disease (299,533).
CONCLUSIONS
In general, there is a good correlation between tau pathology and cognitive state as
well as between the progress of both neuroimaging and the clinical manifestations
of disease. These morphological markers (tau pathology, amyloidosis, and
synaptic proteins as well as biochemical markers, i.e., ChAT and related mRNAs)
usually identify cases with moderate to severe dementia. However, due to variable
overlaps these changes often fail to distinguish between cognitively intact aged
subjects from those with MCI/preclinical or mild AD. In particular these latter
groups show a wide variety in the intensity and pattern of AD-related lesions.
Although they often differ from “normal” aging, only a small proportion of
cognitively intact aged individuals are free of AD pathology, while up to 50% may
show various AD-related alterations or even definitive AD pathology. Additional
difficulties arise from frequent coexistence with other pathologies that have a
synergistic effect and may act by contributing to cognitive impairment.
In view of these difficulties, further studies are needed to increase the
diagnostic validity of current diagnostic criteria of AD and its distinction from
“normal” aging and from other disorders causing dementia.
ABBREVIATIONS
3-R 3-repeat
ACh acetylcholine
AChE acetylcholinesterase
AD Alzheimer’s disease
ADDL amyloid-ß derived diffusable ligand
ADRDA Alzheimer’s Disease and Related Disorders Association
376 Jellinger
ACKNOWLEDGMENTS
The study was supported by the Society for the Support of Research in
Neurological Sciences, Vienna, Austria. Special thanks are due to Mr. E. Mitter-
Ferstl, PhD, for secretarial and computer work.
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428 Jellinger
Karl Herholz
Wolfson Molecular Imaging Centre, University of Manchester, Manchester,
U.K., and Department of Neurology, University of Cologne,
Cologne, Germany
Daniela Perani
Departments of Neuroscience and Nuclear Medicine, Vita-Salute
San Raffaele University and San Raffaele Scientific Institute, Milan, Italy
Chris Morris
Wolfson Unit of Clinical Pharmacology, Chemical Hazards and Poisons
Division–Newcastle, Health Protection Agency, and School of Neurology,
Neurobiology, and Psychiatry, The Medical School, University of Newcastle
upon Tyne, Newcastle upon Tyne, Tyne and Wear, U.K.
INTRODUCTION
Early diagnosis of dementia has many aspects. There is no, and probably will not
be any, diagnostic measure that can be expected to suit all diagnostic situations
equally well and therefore a differentiated approach is necessary. Three gross
scenarios can be distinguished: (1) diagnosis of mild dementia that is clinically
manifest, (2) diagnosis of a dementing disease in a subject who has cognitive
impairment but is not yet demented, and (3) detection of a dementing disease in
an asymptomatic individual.
In addition to these diagnostic aspects, new methods may play an important
role in clinical trials, e.g., by defining efficient inclusion criteria particularly at a
presymptomatic stage, or by providing pathophysiological insights into treatment
effects that have not been available in the past. In this context, imaging and
biomarkers are expected to play an increasing role. A list of criteria for such
markers to be clinically acceptable has been defined by a joint working group of
429
430 Herholz et al.
the Ronald and Nancy Reagan Research Institute of the Alzheimer’s Association
and the National Institute on Aging (1):
1. There should be at least two independent studies that specify the
biomarker’s sensitivity, specificity, and positive and negative
predictive values
2. Sensitivity and specificity should be no less than 80%; positive
predictive value should approach 90%
3. The studies should be well powered, conducted by investigators with
expertise to conduct such studies, and the results published in peer-
reviewed journals
4. The studies should specify type of control subjects, including normal
subjects and those with a dementing illness but not Alzheimer’s
disease (AD)
5. Once a marker is accepted, follow-up data should be collected and
disseminated to monitor its accuracy and diagnostic value.
Some imaging techniques now appear to fulfill or come close to these
requirements, although the request for 90% positive prediction obviously
depends on the selection of subjects. Even excellent biomarkers providing more
than 95% specificity in selected samples would fall short of that goal in the
context of population screening. It can be achieved only in subjects who have
been selected on the grounds that their clinical symptoms already include
dementia or at least a severe memory deficit.
using certain neuropsychological tests for the detection of the earliest stages of
dementia, though these may only be able to detect individuals with certain types
of dementia amnestic mild cognitive impairment (MCI). While requiring
longitudinal validation, the application of such tests should in the future provide
much simplified diagnosis. Thus, in a clinical setting, the degree of impairment
can be assessed neuropsychologically, but fulfillment of different dementia
criteria is ultimately determined through clinical judgement using information
from these tests within a framework that includes other tools. New biomarkers
probably could not reasonably aim at replacing the detection of dementia by
clinical judgement, but should always be seen as part of that process.
Claims for the more ambitious goal of an early distinction between
dementing diseases are difficult to verify because the appropriate gold standard
for this is histopathological postmortem examination and, as pointed out by
Jellinger (see Chapter 11), these standards are lacking at the earliest stages of
dementing diseases. Thus, only longitudinal studies that cover the five to 10 years
that may pass between onset of dementia and death with definitive pathological
diagnosis could have a chance to achieve this. The lack of such studies, though,
demonstrates an urgent scientific need for carefully designed longitudinal studies
that include modern diagnostic procedures that assess specific pathophysiological
processes. Particularly in very elderly individuals with dementia, they should also
address the issue of how various pathophysiological processes interact, such as
cytoskleletal and synaptic changes, deposition of amyloid, tau protein, and
a-synuclein with vascular pathology, and especially during the progression of
dementia. Such studies are costly and difficult to organize, but the payoff is likely
to be enormous.
Somewhat less firm evidence can be gathered when comparing new
diagnostic procedures against each other and against standard clinical criteria.
From the correspondence of diagnostic classifications, we can at least get an
upper estimate of their accuracy and, by considering their pathophysiological
rationale and closeness to purported core pathophysiological events, an idea of
their potential. Most of the studies that are available today fall into this category
and guide our ideas about their relative merits and perspectives.
Qualitative judgment of atrophy on CT and MRI is already in common use
as part of clinical assessment. Progress is to be expected by using age-adjusted
quantitative measures or, at least, standardised performance and reading of scans,
as described by Frisoni and Filippini (see Chapter 6), but the specificity of using
simple measures of atrophy will probably remain low. New techniques, e.g.,
diffusion-weighted imaging and magnetisation transfer techniques, could be
added relatively easily to the standard clinical test batteries in use and show some
promise in improving diagnostic specificity. Molecular laboratory blood tests
could be integrated easily, but none with high predictive power is currently
available. Cerebrospinal fluid (CSF) testing is less likely to be adopted widely due
to its invasiveness. Functional imaging by single photon emission computed
tomography (SPECT) (see Chapter 7) and 18F-2-fluoro-2-deoxy-D-glucose
Guideline and Perspective 433
proposed. The evaluation of memory is the basis for the differential diagnosis of a
dementia in its early stages and cognitive changes related to normal aging. The
availability of proper normative data, however, represents a current challenge
that requires a solution. In the last few years, specific criteria for the diagnosis
of memory deficits in elderly have been proposed. The criteria for “age-
associated memory deficit” and for the nosographic picture of MCI underline
that, in the two conditions, the diagnosis is based on very different psychometric
criteria of memory performance. In MCI an impairment on tests of episodic
memory is generally the most predictive measure (6), but also tests of many
different cognitive capacities have been shown to be predictive of a future
diagnosis of AD among memory-impaired subjects (9–11).
The results from a general practice study of cognitive impairment have
suggested guidelines for the detection of MCI (12). The role of the general
practitioner in the diagnosis of MCI and the potential feasibility of general
practice screening is particularly relevant. It may be possible for the family
practitioner to verify cognitive complaints and to screen for MCI with a high
degree of accuracy using a brief test battery derived from empirical observations
in population studies. Neuropsychological tests with the highest predictive value
for dementia conversion and suitable for use in general practice comprise three
tests (delayed auditory verbal recall, verbal fluency, and visuospatial
construction), giving a specificity of 99% and sensitivity of 73%. In addition,
MCI detection should not be limited to cognitive performance alone. Proxy
observations of behavioral change and information relating to loss of ability to
perform activities of daily living should also be used to improve sensitivity and
also to provide information needed in patient management. However, the
assessment of specific domains of complex instrumental activities of daily living
that might be impaired in MCI need to be determined.
There is a consensus that cognitive and functional abilities need to be
considered in the evaluation of MCI. Individual slopes of decline in both
functional and cognitive performance may be better measures than deficits
assessed according to age-specific norms. However, a true consensus can only be
achieved after longitudinal studies establish the age-specific levels of cognitive
functioning, as well as normal rates of cognitive decline over specific time
periods.
Thus, there is a substantial need for improvement in detection by means of
more reliable diagnostic methods. We are also lacking appropriate diagnostic
terms to label individuals who present clinically as MCI with additional evidence
that this is early AD. Even when the histopathological signs of AD are found at
postmortem examination, standard diagnostic criteria require evidence of
dementia during life to diagnose AD since the presence of some plaques and
tangles is quite common in cognitively intact elderly subjects and the correlation
between these changes and cognitive status is not very close. Quite obviously, the
issue of whether those individuals with histopathological signs of AD but no
dementia would eventually have become demented cannot be clarified based on
Guideline and Perspective 435
ASYMPTOMATIC SUBJECTS
On a population basis, early diagnosis may mean screening of asymptomatic
individuals who may be at increased risk of AD because they are old or have a
relative with AD and carry a common risk factor such as the APOE epsilon 4
allele. In the current situation without proven interventions that could
prevent onset of dementia, there is usually little reason to perform such screening
other than for the purpose of epidemiologic studies, but individuals may still be
worried and seek advice (28).
Genetic counselling is usually recommended for subjects who have
multiple (typically three or more) family members who developed early onset
AD, especially when age of onset was at 55 years or younger. Up to 70% of such
families may have mutations in the amyloid precursor protein (APP), presenilin-1
(PSEN1), or presenilin-2 (PSEN2) genes that can be identified by comprehensive
genetic screening (29). Yet, even in such cases, genetic testing may not provide a
conclusive answer in some families (30). In those rare families where mutations in
APP, PSEN1, or PSEN2 are identified, or in other families where there is a clear
indication of genetically defined autosomal dominant dementia, there are good
reasons to consider predictive testing in at risk family members since, in the vast
majority of cases, the presence of a mutation indicates a high likelihood of
developing dementia (see Chapter 4). This, however, brings with it certain ethical
issues particularly since there are currently no disease slowing therapies. As with
other neurodegenerative disorders with a clearly defined genetic basis, prior to
Guideline and Perspective 437
MONITORING TREATMENT
The traditional approach for assessment of treatment outcome involves clinical
endpoints that are directly related to symptoms of the disease under investigation,
and this is the approach required by drug licensing agencies, such as the United
States Food and Drug Administration, for approval of drugs. For dementia, these
endpoints are usually based on clinical ratings and neuropsychological test scores
to demonstrate an improvement of symptoms (34). It is much more difficult to
prove a reduction in progression of the underlying disease by clinical trials, but
438 Herholz et al.
that could perhaps be achieved by the use of a biomarker that is closely linked to
disease pathophysiology (35,36).
In a recent trial with Ab immunization that was stopped because some
patients developed meningoencephalitis, a decline in CSF tau was observed in
antibody responders compared to placebo controls (37). In a trial of the cholesterol-
lowering drug simvastatin, CSF alpha-sAPP and CSF beta-sAPP were
significantly reduced, but the CSF levels of tau, phosphorylated tau (p-tau),
and Ab (42) and the plasma levels of Ab (42) were unchanged after 12 weeks of
treatment (38). Thus, molecular analysis of CSF appears useful in order to obtain
some insight into possible mechanisms of drug action, but because of the invasive
nature of CSF sampling and a lack of a clear association with clinical benefits, it is
unlikely to be useful as a surrogate marker to determine drug efficacy.
Imaging proposals have been put forward for measurement of atrophy
progression by MRI (39), although it is still difficult to achieve consistency of
measurements across scanners and centres. Nutrition and hydration status may
also have an influence on brain volume (40), but they are unlikely to influence
specifically local mesial temporal atrophy that is typical of AD. Feasibility of
quantitative PET analysis has been demonstrated in multi-center studies (41,42).
Tools for efficient quantitative regional image analysis have been developed for
multi-center trials that are now underway (43).
Measurement of the progression of hippocampal atrophy by MRI appears
to have some face validity as a marker of degenerative disease progression and is
beginning to be used in drug trials (44). Analysis of the recent immunization trial,
however, has suggested that under certain circumstances there may be a
dissociation between clinical scores and progression of atrophy, e.g., if volume
changes were due to amyloid removal and associated cerebral fluid shifts (45).
Measurement of functional changes in the brain in therapeutic trials have
been accomplished with SPECT (46–48) and PET (49) (see Chapter 8 for further
citations), and also more recently with magnetic resonance spectroscopy (MRS)
(44,50) and functional magnetic resonance imaging (fMRI) (51). In these studies,
there was generally a good correlation between clinical response and an increase
in regional blood flow and glucose metabolism. Such data suggests that
functional imaging techniques have the potential to be used as a surrogate
endpoint in clinical trials although a number of limitations such as the technical
variability across centers and a clear definition of reproducible functional
conditions during studies still applies (35).
Molecular tracers also permit measurement of specific drug effects. The
degree of acetylcholinesterase (AChE) inhibition in the brain of AD patients has
been measured by PET (52,53), demonstrating that cognitive effects are related to
local drug activity which varies among patients and may not be very well
predicted by preclinical studies (54). Amyloid imaging tracers are likely to
provide the most direct in vivo evidence of whether new drugs are actually able to
reduce Ab deposition in the brain and whether that is related to clinical benefits
(see Chapter 10). In addition, drug companies are increasingly likely to study
Guideline and Perspective 439
human regional pharmacokinetics and receptor binding using PET and SPECT at
the preclinical and early clinical phase of drug development (55).
CONCLUSION
A large number of new tools have been developed in recent years for the
improved early diagnosis of AD. New neuropsychological screening tools
facilitate diagnosis of MCI as a condition with substantially increased risk for
developing AD. Functional imaging methods and MRI volumetry have
demonstrated usefulness in identifying subgroups of MCI patients with a very
high likelihood of developing AD within a few years. Molecular CSF tests and
imaging techniques even hold the promise for the early distinction of AD from
other disorders that lead to dementia prior to its onset, and new molecular
techniques for identifying new diagnostic targets may provide improved
methods of detection. Genetic counselling is recommended in early-onset
familial AD, where specific monogenetic mutations may play a causative role.
In the future we should see integration of these new possibilities into longi-
tudinal and therapeutic intervention studies that should lead to more effective
development of drugs to slow progression or even prevent development of
dementia.
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Index
443
444 Index
inclusions
('protective sinks' & 'delayed-toxicity')
Figure 9.1 Loss of dopamine transporters with age. (See page 256.)
Figure 9.3 Acetylcholinesterase activity in the central nervous system evaluated with
C11-MP4A. (See page 262.)
Figure 9.4 Dopamine transporter loss visualized with FP-CIT SPECT in the differential diagnosis
of dementia with Lewy bodies (DLB). (See page 267.)
Figure 9.5 Differences in cortical GABAA receptor density between patients with leuko-
araiosis and dementia (A) and patients with leukoaraiosis without dementia (B). (See page 269.)
Figure 10.6 Examples of Pittsburgh compound-B (PIB) Logan distribution volume ratio
(DVR) images. (See page 295.)
Figure 11.1 Spreading pattern of (A) cytoskeletal/tau pathology and (B) of Ab
deposition. (See page 325.)