Journal Pre-proof

TFOS DEWS III Digest Report

Fiona Stapleton , Pablo Argüeso , Penny Asbell , Dimitri Azar ,

Charles Bosworth , Wei Chen , Joseph Ciolino , Jennifer P. Craig ,
Juana Gallar , Anat Galor , José A.P. Gomes , Isabelle Jalbert ,
Ying Jie , Lyndon Jones , Kenji Konomi , Yang Liu ,
Jesus Merayo-Lloves , Fabiola R. Oliveira ,
Victor A. Perez Quinones , Eduardo M. Rocha ,
Benjamin D. Sullivan , David A. Sullivan , Jelle Vehof ,
Susan Vitale , Mark Willcox , James Wolffsohn , Murat Dogru

PII: S0002-9394(25)00276-4
DOI: https://doi.org/10.1016/j.ajo.2025.05.040
Reference: AJOPHT 13398

To appear in: American Journal of Ophthalmology

Received date: May 18, 2025

Accepted date: May 23, 2025

Please cite this article as: Fiona Stapleton , Pablo Argüeso , Penny Asbell , Dimitri Azar ,
Charles Bosworth , Wei Chen , Joseph Ciolino , Jennifer P. Craig , Juana Gallar , Anat Galor ,
José A.P. Gomes , Isabelle Jalbert , Ying Jie , Lyndon Jones , Kenji Konomi , Yang Liu ,
Jesus Merayo-Lloves , Fabiola R. Oliveira , Victor A. Perez Quinones , Eduardo M. Rocha ,
Benjamin D. Sullivan , David A. Sullivan , Jelle Vehof , Susan Vitale , Mark Willcox ,
James Wolffsohn , Murat Dogru , TFOS DEWS III Digest Report, American Journal of Ophthal-
mology (2025), doi: https://doi.org/10.1016/j.ajo.2025.05.040

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 TFOS DEWS III Digest Report

Fiona Stapleton1, Pablo Argüeso2, Penny Asbell1,3, Dimitri Azar4, Charles

Bosworth5, Wei Chen6, Joseph Ciolino7, Jennifer P. Craig8, Juana Gallar9, Anat
Galor10, José A.P. Gomes11, Isabelle Jalbert1, Ying Jie12,13, Lyndon Jones14,
Kenji Konomi15, Yang Liu16, Jesus Merayo-Lloves17, Fabiola R. Oliveira18, Victor
A. Perez Quinones19, Eduardo M. Rocha20, Benjamin D. Sullivan21,22, David A.
Sullivan23, Jelle Vehof24,25, Susan Vitale26, Mark Willcox1, James Wolffsohn27,
Murat Dogru28

1. School of Optometry and Vision Science, UNSW Sydney, NSW, Australia

2. Department of Ophthalmology, Tufts Medical Center, Tufts University School
of Medicine, Boston, MA, USA
3. Biomedical Engineering, University of Memphis, Visiting Professorial Fellow,
Memphis, USA
4. Chicago College of Medicine, University of Illinois, Chicago, IL, USA
5. Azura Ophthalmics, Tel Aviv, Israel
6. Eye Hospital of Wenzhou Medical University, National Eye Clinic Research
Center, Peoples Republic of China
7. Cornea Service, Massachusetts Eye and Ear, Department of Ophthalmology,
Harvard Medical School, Boston, MA, USA
8. Department of Ophthalmology, New Zealand National Eye Centre, The
University of Auckland, Auckland, New Zealand
9. Ocular Neurobiology Group, Instituto de Neurociencias, Universidad Miguel
Hernández-Consejo Superior de Investigaciones Científicas, San Juan de
Alicante, Spain
10. Surgical Services, Miami Veterans Affairs Hospital, Bascom Palmer Eye
Institute, University of Miami, Miami, FL, USA
11. Department of Ophthalmology and Visual Sciences, Paulista School of
Medicine / Federal University of São Paulo, São Paulo, Brazil
12. Beijing Institute of Ophthalmology, Beijing Tongren Eye Center, Beijing
Tongren Hospital, Capital Medical University, Beijing, China
13. Beijing Ophthalmology & Visual Sciences Key Laboratory, Beijing, China
14. Centre for Ocular Research & Education (CORE), School of Optometry and
Vision Science, University of Waterloo, Waterloo, ON, Canada
15. Clinical and Translational Research Center, Keio University Hospital, Tokyo,
Japan
16. Department of Ophthalmology, Zhongnan Hospital of Wuhan University,
Wuhan, China

1
17. Instituto Universitario Fernandez-Vega, Universidad de Oviedo, Principality
of Asturias, Spain
18. Department of Clinical Medicine, Division of Rheumatology and Clinical
Immunology, Ribeirao Preto Medical School, University of Sao Paulo,
Ribeirao Preto, SP, Brazil
19. Department of Ophthalmology, University of Miami Miller School of Medicine,
Miami, FL, USA
20. Department of Ophthalmology, Otorhinolaryngology and Head & Neck
Surgery, Ribeirao Preto Medical School, University of Sao Paulo, Ribeirao
Preto, SP, , Brazil
21. Lµbris BioPharma, LLC, Naples, FL, USA
22. Bausch & Lomb, Bridgewater, NJ, USA
23. Schepens Eye Research Institute, Department of Ophthalmology, Harvard
Medical School, and Massachusetts Eye and Ear, Boston, MA, USA
24. Departments of Ophthalmology and Epidemiology, University of Groningen,
University Medical Center Groningen, Groningen, the Netherlands
25. Department of Ophthalmology, Vestfold Hospital Trust, Tønsberg, Norway
26. Division of Epidemiology and Clinical Applications, National Eye Institute,
National Institutes of Health, Bethesda, MD, USA
27. College of Health & Life Sciences, School of Optometry, Aston University,
Birmingham, UK
28. Ichikawa General Hospital Tokyo Dental College Dept of Ophthalmology
Ichikawa, Chiba, Japan

2
Table of contents

1. Abstract ........................................................................................................... 9

2. Keywords ........................................................................................................ 9

3. Abbreviations .................................................................................................. 9

4. Introduction ................................................................................................... 10

5. Sex, gender, and hormones ......................................................................... 11

5.1 Introduction .............................................................................................. 11

5.2 Sex-related differences in the ocular surface and adnexa ................... 11

5.2.1 Lacrimal gland ......................................................................................... 11
5.2.2 Meibomian gland ..................................................................................... 12
5.2.3 Cornea .................................................................................................... 13
5.2.4 Eyelid blinking ......................................................................................... 13

5.3 Sex-related differences and immunity ................................................... 13

5.4 Sex and gender differences in pain assessment .................................. 14

5.5 Hormonal regulation of the ocular surface and adnexa ....................... 15

5.5.1 Androgens............................................................................................... 15
5.5.2 Estrogens ................................................................................................ 16
5.5.3 Progestins ............................................................................................... 17
5.5.4 Sex steroids in the tear film ..................................................................... 17

5.6 Insulin-like growth factor and insulin .................................................... 17

5.7 Thyroid hormone regulation of the ocular surface and adnexa .......... 18

5.8 Gender and DED ...................................................................................... 20

5.9 Future directions...................................................................................... 22

6. Epidemiology ................................................................................................ 22

6.1 Scope of the update ................................................................................. 22

6.2 Operational definitions ............................................................................ 22

3
6.3 Prevalence of DED ................................................................................... 26
6.3.1 Prevalence of DED based on the Women’s Health Study criteria ........... 26
6.3.2 Prevalence of symptomatic DED ............................................................ 26
6.3.3 Prevalence of DED based on signs and symptoms ................................ 27
6.3.4 Prevalence of DED based on TFOS DEWS II criteria ............................. 27
6.3.5 Prevalence based on claims data ........................................................... 27
6.3.6 Prevalence of DED based on clinical diagnosis ...................................... 27
6.3.7 Prevalence of any MGD .......................................................................... 27
6.3.8 Prevalence of clinically significant MGD ................................................. 27

6.4 Annual incidence of DED ........................................................................ 28

6.5 Natural history of DED ............................................................................. 29

6.6 Risk factors for DED ................................................................................ 30

6.6.1 Systemic disorders .................................................................................. 34
6.6.2 Ophthalmic disorders .............................................................................. 36
6.6.3 Surgery and other procedures ................................................................ 37
6.6.4 Medication use (See Section 10) ............................................................ 38
6.6.5 Environmental factors ............................................................................. 39
6.6.6 Demographic factors ............................................................................... 40
6.6.7 Other factors ........................................................................................... 40
6.6.8 Risk factors for MGD ............................................................................... 41

6.7 Morbidity and impact ............................................................................... 42

6.8 Summary and outstanding questions .................................................... 42

7. Pathophysiology ........................................................................................... 43

7.1 Introduction .............................................................................................. 43

7.2 Initiating triggers of disease ................................................................... 45

7.3 Hyperosmolarity....................................................................................... 46

7.4 Proteases .................................................................................................. 47

7.5 Sub-functional or absent glycocalyx ..................................................... 49

7.6 Meibomian gland dysfunction ................................................................ 51

7.7 Inflammatory cell recruitment ................................................................. 52

4
7.8 Immune cell dysfunction ......................................................................... 53

7.9 Future directions...................................................................................... 53

8. Tear Film........................................................................................................ 54

8.1 Introduction .............................................................................................. 54

8.2 Clinical measurements of the tear film .................................................. 55

8.3 The tear lipids – composition and function ........................................... 56

8.4 The tear proteome .................................................................................... 59

8.5 Lipid-protein-mucin interactions ............................................................ 60

8.6 Mucins ...................................................................................................... 60

8.7 MicroRNAs (miRNAs) .............................................................................. 61

8.8 Translational dry eye models of tear film .............................................. 61

8.8.1 In vitro models......................................................................................... 61
8.8.2 In vivo models ......................................................................................... 62

8.9 Summary and future directions .............................................................. 62

9. Pain and Sensation....................................................................................... 63

9.1 Introduction .............................................................................................. 63

9.2 Corneal nerve remodeling in adults ....................................................... 63

9.3 Corneal nerve regeneration after surgery.............................................. 64

9.4 Nerve regeneration in pathological conditions ..................................... 66

9.5 Nerve abnormalities and DED ................................................................. 67

9.5.1 Impact of DED on nerve structure and function in animal models........... 69
9.5.2 Structural and functional nerve alterations in DED in human .................. 71
9.5.2.1 Corneal nerve anatomy in DED ........................................................... 72
9.5.2.2 Corneal sensitivity in DED.................................................................... 78

9.6 Anesthetic challenge in DED .................................................................. 82

9.7 Quantitative sensory testing in DED ...................................................... 83

5
9.8 Brain imaging in DED .............................................................................. 85

9.9 Future directions and conclusions ........................................................ 86

10. Iatrogenic .................................................................................................. 87

10.1 Introduction .............................................................................................. 87

10.2 Topical drug-induced DED ...................................................................... 88

10.2.1 Prevalence ........................................................................................... 88
10.2.2 Topical drugs contributing to DED ....................................................... 89
10.2.3 Mechanism........................................................................................... 90
10.2.4 Role of preservatives and excipients ................................................... 91
10.2.5 Recommendations for management .................................................... 92

10.3 Systemic drug-induced DED ................................................................... 93

10.3.1 Prevalence ........................................................................................... 93
10.3.2 Medications and mechanisms .............................................................. 94
10.3.2.1 Tamsulosin........................................................................................ 94
10.3.2.2 Antihistamines/anticholinergic drugs ................................................. 94
10.3.2.3 Isotretinoin ........................................................................................ 95
10.3.2.4 Chloroquine / Hydroxychloroquine .................................................... 96
10.3.2.5 Corticosteroids and non-steroidal anti-inflammatories ...................... 96
10.3.2.6 Antibiotics.......................................................................................... 97
10.3.2.7 Antidepressants / anxiolytics / mood stabilizers ................................ 97
10.3.2.8 Hormone replacement therapy .......................................................... 98

10.4 Contact lenses and DED.......................................................................... 98

10.4.1 Prevalence of DED in contact lens wear .............................................. 99
10.4.2 Mechanism........................................................................................... 99
10.4.3 Recommendations for management of DED in contact lens wearers 101

10.5 Procedures ............................................................................................. 102

10.5.1 Botulinum toxin .................................................................................. 102
10.5.1.1 Mechanism ...................................................................................... 102
10.5.1.2 Direct inhibition of tear secretion: .................................................... 102
10.5.1.3 Impact on meibomian gland function: ............................................. 102
10.5.1.4 Regulation of inflammatory responses: ........................................... 103
10.5.1.5 Chemodenervation of orbicularis oculi: ........................................... 103
10.5.1.6 Recommendations for management ............................................... 103
10.5.2 Corneal collagen crosslinking ............................................................ 104
10.5.2.1 Mechanism ...................................................................................... 104

6
10.5.2.2 Recommendations for management ............................................... 104
10.5.3 Other procedures ............................................................................... 105
10.5.3.1 Eye cosmetics and beauty treatments ............................................ 105

10.6 Non-ophthalmic conditions................................................................... 105

10.6.1 Radiotherapy...................................................................................... 105
10.6.1.1 Mechanism ...................................................................................... 105
10.6.1.2 Recommendations for management ............................................... 106
10.6.2 Bariatric surgery ................................................................................. 106
10.6.3 Stem cell or bone marrow transplant ................................................. 106
10.6.3.1 Mechanism ...................................................................................... 106
10.6.3.2 Recommendations for management ............................................... 107

10.7 Future directions and conclusions ...................................................... 107

11. Clinical trials design .............................................................................. 108

11.1 Introduction ............................................................................................ 108

11.2 Approvals for DED ................................................................................. 110

11.3 Defining a pathway toward approval .................................................... 115

11.3.1 Solving industry-wide failure rates ..................................................... 115
11.3.2 Overcoming disease heterogeneity with targeted sub-populations .... 115
11.3.3 US FDA clear guidance for industry and flexibility for dry eye approvals
122
11.3.3.1 Trial design ..................................................................................... 122
11.3.3.2 Comparator(s) ................................................................................. 123
11.3.3.3 Trial population ............................................................................... 124
11.3.3.4 Demonstration of efficacy................................................................ 124
11.3.3.5 Safety database .............................................................................. 125
11.3.4 EMA guidance for industry and flexibility for DED approvals ............. 126
11.3.5 PMDA guidance for industry and flexibility for approvals ................... 127

11.4 Devices ................................................................................................... 129

11.4.1 Regulatory pathway (FDA) ................................................................. 129
11.4.2 Regulatory pathway (Japan, PMDA) .................................................. 131
11.4.3 Artificial tears (US, FDA) .................................................................... 131
11.4.4 Artificial tears (Japan, PMDA) ............................................................ 132
11.4.5 Devices approved for DED/MGD ....................................................... 132

11.5 Design features to enhance clinical trial data quality and decisions 133

7
11.5.1 Biomarkers / proof of mechanism / early signal of efficacy ................ 133
11.5.2 Missing data considerations ............................................................... 134
11.5.3 Global trial designs ............................................................................ 134
11.5.4 Compliance monitoring ...................................................................... 135

11.6 Conclusions ........................................................................................... 135

12. Summary................................................................................................. 136

13. Acknowledgements ............................................................................... 138

14. Figures and tables ................................................................................. 138

15. References.............................................................................................. 139

8
1. Abstract

This digest summarises the interdisciplinary research in dry eye disease (DED)
published since the 2017 TFOS DEWS II reports. It comprises seven topics
including Sex, Gender, and Hormones, Epidemiology, Pathophysiology, Tear
Film, Pain and Sensation, Iatrogenic and Clinical Trial Design and explores how
each of these inform diagnostic methodology, disease subtype and management
of DED.

Sex- and gender-related differences significantly influence the ocular surface due
to hormones, sex chromosomes, sex-specific autosomal factors, epigenetics,
care-seeking behaviors, and service utilization. Epidemiological data reveal that
DED prevalence varies by age and sex, influenced by diagnostic criteria and the
multifactorial nature of the disease. New risk factors for DED include
environmental, iatrogenic, systemic diseases and lifestyle domains.

Pathophysiological distinctions between Aqueous Deficient Dry Eye (ADDE) and

Evaporative Dry Eye (EDE) have been clarified. EDE is characterized by a muted
inflammatory response at the ocular surface, meibomian gland dysfunction and
conceivably phenotypic changes in corneal epithelial cells. There is an expanding
role for metabolic, hormonal, physical, neural and cellular stresses, including
hyperosmolarity, mitochondrial stress, and neurogenic inflammation.

Advancements in tear film research recommend new approaches to

understanding DED pathogenesis and identifying biomarkers, such as
microRNAs. Ocular pain perception is linked to structural integrity of corneal
nerves, functional capacities of neurons, and activity of the central and peripheral
nervous systems. Iatrogenic DED can result from medications, contact lenses,
and surgical procedures. Clinical trials now emphasize aligning design and
endpoints with DED subtypes and therapeutic mechanisms, with new
therapeutics and trial designs under consideration.

2. Keywords

Epidemiology, prevalence, risk factors, hormones, sex, gender, tear film,

pathophysiology, neurogenic, clinical trials, iatrogenic

3. Abbreviations
ADDE Aqueous-deficient dry eye disease
BALB/c BALB/C wildtype (mice)
CXL Corneal cross-linking
DED Dry eye disease
DEQ-5 5-Item Dry Eye Questionnaire

9
DEWS Dry eye workshops
DNA Deoxyribonucleic acid
EDE Evaporative dry eye disease
FDA Food and drug administration
(f)MRI (Functional) magnetic resonance imaging
GVHD Graft versus host disease
HLA Human leukocyte antigen
HPMC Hydroxypropyl methylcellulose
ICAM-1 Intercellular Adhesion Molecule 1
IL Interleukin
IGF Insulin-like growth factor
IVCM In vivo confocal microscopy
LASIK Laser-assisted in situ keratomileusis
LIPCOF Lid-parallel conjunctival folds
LLT Lipid layer thickness
MGD Meibomian gland dysfunction
MGYLS Meibomian glands yielding liquid secretions
MMP Matrix metalloproteinase
MUC4 Mucin 4
NGF Nerve growth factor
NIBUT Non-invasive tear film breakup time
NF-kB Nuclear factor kappa-light-chain-enhancer of activated B cells
NRS Numerical rating scale
OCT Optical coherence tomography
OR Odds ratio
OSDI Ocular Surface Disease Index
PRK Photorefractive keratectomy
QoL Quality of Life
SANDE Symptom Assessment iN Dry Eye questionnaire
SPEED Standard Patient Evaluation of Eye Dryness questionnaire
TBUT Tear film breakup time
TED Thyroid eye disease
TFOS Tear Film & Ocular Surface Society
TRPM8 Transient Receptor Potential cation channel subfamily M member 8
VAS Visual analogue scale

4. Introduction
This review updates the evidence for interdisciplinary aspects of dry eye disease
(DED). It considered novel, human and animal evidence-based research
published between 2017 and 2024. The goal of this report was to identify key

10
research published since the 2017 TFOS DEWS II Workshop reports to underpin
the evidence described in the TFOS DEWS III Diagnostic Methodology 1 and
Management and Therapy 2 reports. The topics include sex, gender, and
hormones, epidemiology including prevalence and risk factors, novel concepts
and findings associated with pathophysiology, relevant changes to the tear film
and interactions, the mechanisms related to ocular pain and sensation, iatrogenic
causes of DED and their unique management and a synthesis of clinical trial
designs to inform exploration of new treatment modalities.

5. Sex, gender, and hormones

5.1 Introduction

The TFOS DEWS II report on Sex, Gender, and Hormones report 3 addressed
many sex- and gender-related differences that significantly influence the ocular
surface in health and DED. Many of these differences appeared to be due to the
effects of hormones, sex chromosomes, sex-specific autosomal factors,
epigenetics, care-seeking behaviors and service utilization. 3 The purpose of this
section is to highlight some of the relevant research since the publication of that
report. The focus is primarily on studies published after July 1, 2017.

Additional studies related to sexual health and DED may be found in the recent
detailed review, entitled "TFOS Lifestyle: Impact of lifestyle challenges on the
ocular surface". 4

5.2 Sex-related differences in the ocular surface and adnexa

Significant sex-related differences in the lacrimal gland, meibomian gland, cornea

and eyelid, reported since publication of the TFOS DEWS II report on Sex,
Gender, and Hormones, 3 are briefly highlighted below. These differences may
contribute to the increased prevalence of DED in females.

5.2.1 Lacrimal gland

Significant, sex-related differences exist in the gene expression, morphology, and
pathophysiology of the lacrimal gland. The sex-associated differences in lacrimal
gland gene expression may be very important in promoting lymphocyte
accumulation in this tissue and contributing to the onset, progression, and/or
severity of the inflammatory disease process in Sjögren disease. 5 This condition
is an multisystem autoimmune disease affecting the exocrine glands including
the salivary and lacrimal glands that occurs primarily in women, and is
associated with aqueous-deficient DED. 6 Murine models of autoimmune lacrimal
gland disease, such as MRL/MpJ-Tnfrsf6lpr and non-obese diabetic/LtJ mice,

11
have provided critical insights into sex-related immune differences. The extent of
lacrimal gland inflammation in MRL/MpJ-Tnfrsf6lpr mice is, as in humans, far
greater in females as compared with males, whereas the magnitude of lacrimal
gland inflammation is far worse in non-obese diabetic/LtJ males. 7 Results
showed that sex significantly influences the expression of thousands of genes,
and that the immune nature of the glandular response is very dependent on the
Sjögren disease model. Lacrimal tissues of female, as compared with male,
MRL/MpJ-Tnfrsf6lpr mice featured a significant increase in the expression of
genes related to inflammatory responses, antigen processing, and chemokine
pathways. In contrast, it was the lacrimal glands of non-obese diabetic/LtJ males,
and not females, that presented with a significantly greater expression of
immune-related genes. These data suggest that factors in the lacrimal gland
microenvironment may be critically important in mediating these sex-associated
immune effects and in promoting lacrimal gland inflammation. 5

Analyses of single-cell transcriptomes from lacrimal glands of MRL/MpJ-Tnfrsf6lpr,

non-obese diabetic/LtJ and wild-type (BALB/c) mice have also defined the
location of multiple cell-type-specific mRNA markers and proteins, the latter of
which may be secreted into the tear film in a sex-specific manner. 8

As concerns morphology and pathophysiology, significant, sex-linked differences

occur during aging in the magnitude of acinar atrophy, periacinar fibrosis,
periductal fibrosis, ductal dilation, ductal proliferation, fatty infiltration, and
lymphocyte infiltration in human lacrimal glands. 9 Female tissues had a higher
frequency of all observed degenerative changes, except for ductal dilation, which
was significantly more prevalent in male glands. 9 The authors concluded that
female lacrimal glands are more susceptible to degeneration, conceivably due to
hormonal influences, estrogen withdrawal or genetic susceptibility, and that this
susceptibility may play a significant role in the higher prevalence of DED in older
women. 9

5.2.2 Meibomian gland

Significant sex-related differences exist in meibomian gland gene expression, but
the nature of these differences may be primarily species-dependent. 10 Analysis
of the 500 most highly expressed genes from human and BALB/c mouse
meibomian glands demonstrated that only 24.4% were the same. Further,
analysis of 100 genes with the greatest sex-associated differences in the human
and mouse meibomian glands showed that none were the same, indicating that
mice are not optimal models for understanding sex-associated differences in
gene expression of the human meibomian glands. 10

12
The prevalence of meibomian gland dysfunction (MGD) may vary by sex but
results are inconsistent. Population- and hospital-based studies report that MGD
may occur more frequently in males, or in females, or in neither sex. A recent
prospective cross-sectional study indicated that older males had more severe lid
margin abnormities and decreased gland number, height, and area compared
with females. 11 Some studies suggest that the influence of sex on MGD may
depend on the type (e.g. obstructive versus hypersecretory forms, 12 the patient's
age, 13 and/or the individual's medical condition. 14 The inconsistency in the sex-
related prevalence of MGD is notable, given that DED is more common in
females 15-24 and that MGD is a major cause of DED. 6,15,25,26 The prevalence of
any MGD and clinically significant MGD are separately reported in a meta-
analysis by age and sex (see Section 5, Figure 3 F and G). In brief, any MGD
including asymptomatic gland changes is more prevalent in men than women in
older age groups but sex-differences in clinically significant MGD are equivocal.
There remains a need to explore the impact of more detailed diagnostic criteria to
understand sex-related effects on MGD.

5.2.3 Cornea
Significant, sex-related differences exist in corneal thickness, sensitivity, re-
epithelization and DED-induced damage. As has been found previously, males
have greater corneal epithelial thickness in all but the peripheral nasal zone 27
and females have higher corneal sensitivity, 28 and corneal nerve regeneration 29
and slower corneal epithelial wound healing. 30 In addition, aqueous-deficient
DED elicits more ocular pain, anxiety and severe corneal damage in female mice.
31

5.2.4 Eyelid blinking

A sexual dimorphism has also been identified in eyelid blinking. DED appears to
increase sex-related differences in blinking, including heightened exaggeration of
excitability in males and enhanced modifiability of the female trigeminal complex.
This latter modifiability is proposed to explain the female predominance in the
development of focal dystonia and benign essential blepharospasm. 32

5.3 Sex-related differences and immunity

As stated in the TFOS Sex, Gender, and Hormones report, 3 sex-related

differences are well known to occur in both innate and adaptive immunity and
lead to differences in the severity and frequency of infections (male > female)
and the risk of developing autoimmune diseases (female > male). These sex-
based variations appear to be due several factors, including sex steroid
hormones, genetics, the microbiome and non-biological factors.

13
Since that report was published, almost 6,000 articles have been cited in
PubMed addressing the phrase "sex differences and immune." These have
continued to show that sex as a biological factor significantly influences the
distribution of lymphocyte subsets, quality of T cell responses, development of
regulatory T cells, formation of the germinal centers, and the epigenetic
accessibility of B cell loci. 33,34 Sex also impacts transcriptional differences that
are often highly immune cell-specific. 35 For example, more than 50 monocyte
transcripts linked to the interferon pathway, inflammatory cytokines and
chemokines display sex-associated expression, that are prominent in some
female subjects. 35 In addition, changes in sex steroid hormone concentrations
over the course of one's life contribute to sex-related differences in immune
profiles and disease susceptibility patterns. 36

There are also significant sex-related differences in regulatory processes

between transcription factors and their target genes in multiple tissues. 37
Different transcription factors may regulate genes in males and females,
irrespective of whether those target genes are differentially expressed. This sex-
associated pattern of gene regulation may help to explain why males and
females do not often manifest disease in the same way, or respond in the same
way to treatment. 37

All of these sex-related differences in immunity should have relevance to the

ocular surface and adnexa in health and disease.

5.4 Sex and gender differences in pain assessment

The intersection of sex and pain has been widely studied at clinical,
psychological, and social levels. However, pain research often conflates sex and
gender, and little is known about how gender identity diversity, such as gender-
affirming medical procedures (hormonal or surgical therapies), environmental
exposures, and minority status, impact pain. 38

Despite chronic pain and DED being more common in women, most pain
mechanism studies are based on male rodents. 39 With the inclusion of sex as a
biological variable in preclinical research, studies on the influence of sex on pain
and analgesia have increased. Female sex and older age are still the main
factors associated with chronic pain. Experimental pain response differences
suggest a biological mechanism rather than sociocultural gender-related issues.
40

Studies have investigated whether pain sensitivity influences DED symptoms

differently between sexes, accounting for ocular parameters. In a cross-sectional
14
study of a young and healthy cohort (194 women and 93 men), intersex
differences in ocular surface and pain sensitivity were linked to higher DED
symptoms in women. 41 This finding helps explain the disparity between DED
symptom intensity and signs previously described in females.

5.5 Hormonal regulation of the ocular surface and adnexa

5.5.1 Androgens
As detailed in the TFOS DEWS II report, 3 androgens are extremely important in
the regulation of the ocular surface and adnexa and appear to mediate many of
the sex-related differences in these tissues. Androgen deficiency, in turn, is
associated with, and a risk factor for, both aqueous-deficient and evaporative
DED. Recent studies are discussed below.

A one-month administration of the anti-androgen, finasteride, led to the

development of rat lacrimal gland inflammation and aqueous tear deficiency. 42
Conversely, testosterone treatment of female MRL/MpJ-Tnfrsf6lpr mice led to a
striking down-regulation of the lacrimal gland expression of over 60 immune-
associated biological process ontologies (≥ 20 genes/ontology), including those
related to immune system processes, lymphocyte activation, cytokine production,
and inflammatory response. 43 The nature of this androgen effect was dependent
upon murine strain, and the data indicate a major role for the lacrimal gland
microenvironment in mediating androgen effects on immune gene expression. 43

In human meibomian gland epithelial cells in vitro,dihydrotestosterone

administration suppressed proinflammatory gene expression, 44 a hormone
action that may contribute to the typical absence of inflammation within the
human glands. 6,25 Dihydrotestosterone also inhibited the hyperosmolar-induced
expression of TNF-α, IL-8 and IL-6 mRNAs in human corneal epithelial cells, 45
but had no influence on proinflammatory gene expression in unchallenged
human corneal epithelial cells. 44 In contrast, dihydrotestosterone significantly
increased 33 gene ontologies linked to the immune system in human conjunctival
epithelial cells. 44

Anti-androgen therapy for the treatment of prostate cancer and benign prostate
hyperplasia led to a significant increase in the signs and symptoms of MGD and
DED. 46,47 Transdermal androgen therapy applied to the lower abdomen, in turn,
alleviated DED signs and symptoms in androgen-deficient individuals, 48 as well
as DED signs in rabbits with combined androgen deficiency and MGD. 49

Androgenetic alopecia was associated with decreased tear film breakup times,
increased meiboscores, ocular surface symptoms and MGD. 50 However, this

15
form of hair loss is not necessarily linked to androgen excess, but is associated
with an increased risk of polycystic ovary syndrome. 51

5.5.2 Estrogens
It has recently been reported that estrogen receptor-1 deficiency in mice induces
inflammation and lipid deposition in the meibomian gland and lacrimal gland. 52
Estrogen receptor-1 loss however, does not block estrogen receptor-activity, but
results in an abnormal endocrine environment (e.g. increased luteinizing
hormone, estradiol, testosterone, and progesterone levels) and may lead to
insulin resistance. 53-55

Ovariectomy of monkeys 56 and rats 57 engenders the signs of DED, and that
these can be reversed with estrogen administration. 57 Other studies show that
ovariectomy leads to rat anxiety and depressive-like behavior 58 conditions that
promote DED, 4, and that estrogen treatment may amplify rat ocular hyperalgesia.
59
Some of these disparate findings regarding the role of estrogens might be
explained by differences in experimental design, hormone dosage or animal
model. 60 However, complete estrogen absence does not cause lacrimal gland
inflammation, gross alterations in meibomian gland histology, or aqueous-
deficient DED in mice, and does not play a major role in the sex-related
differences of the mouse meibomian gland. 60,61

However, is it possible that this animal research does not reflect the situation in
humans? This question is prompted by the results from aromatase inhibitor
studies, which report an increased prevalence of DED. 62-65 Aromatase inhibitors
block the synthesis of estrogens, induce estrogen deficiency, and are used as
therapy in women with hormone receptor-positive breast cancer post initial
chemotherapy. 66 However, these studies had no control groups. Treatment with
aromatase inhibitors is known to promote anxiety (> 112 PubMed articles),
depression (> 243 PubMed articles), and sleep disturbance (> 25 PubMed
articles) (e.g., 67,68, all of which are associated with DED. 4 It may be that these
mental health factors and sleep disorders contribute to, and possibly account for,
these DED effects seen in those treated with aromatase inhibitors.

Investigators have also suggested that the increased estrogen levels following in
vitro fertilization are responsible for the in vitro fertilization-associated DED
symptoms 69 and/or signs. 69,70 However, there were no controls with these
studies, and the effects may also have been linked to the known in vitro
fertilization-induced anxiety, depression and sleep disturbance. 71

16
Clinicians continue to test whether topical estradiol might serve as a DED
treatment. One of the most recent clinical trials showed no significant differences
in effects between any of the estrogen dosages and the placebo. 72

Lastly, estradiol may inhibit the conjunctival goblet cell response in vitro to an
inflammatory stimulus. 73 This effect may be dose-dependent, given that the
estrogen concentration used (0.1 µM) was considerably higher than the
physiological range. Very high doses of estrogen often suppress, whereas
physiological doses often enhance, immune responses. 74

5.5.3 Progestins
Progesterone appears to suppress ocular pain and discomfort, which are
common features of DED. Within 30 minutes after application to the rat forehead,
1% progesterone gel (i.e. 10 mg/mL) produced corneal antinociception. 75
Forehead application of the same dose, twice per day, for ten weeks led to a
significant decrease in the frequency and severity of ocular symptoms in ocular
graft-versus-host patients. 76 Researchers speculated that this hormone effect
may be mediated by the V1 branch of the trigeminal nerve that innervates the
forehead skin, and that progesterone modifies the signal relay in the rostral and
caudal trigeminal nucleus to dampen nociception. 76 However, given that the
applied progesterone concentration in these studies was so much higher than
that typically found in blood (e.g. ng/ml levels), it might be possible that hormone
action also involved other receptors. Progestins are known to bind glucocorticoid,
androgen and mineralocorticoid receptors, as well membrane receptors, oxytocin
receptors and γ-aminobutyric acid (GABAA). 77,78

5.5.4 Sex steroids in the tear film

Investigators continue to try to measure 17β-estradiol, progesterone and
testosterone in the human tear film. 79-81 However, as explained previously 82,
such measurements are of questionable, or no, relevance, and do not
necessarily reflect the concentration of sex steroids or their metabolites in any
ocular tissue. The processes by which sex steroids are synthesized and
metabolized in humans are addressed in detail in the TFOS DEWS II Sex,
Gender, and Hormones report. 3

5.6 Insulin-like growth factor and insulin

Insulin-like growth factor (IGF)-1 levels in tears may be an indicator of ocular

surface health. Higher IGF-1 levels in tears were found in young adults compared
to older adults, correlating positively with tear film break-up time (TBUT) and
Schirmer test results. 83 In experimental studies, IGF binding protein-3 plays a

17
role in delivering IGF to target cells and works independently of IGF. IGF binding
protein-3 levels decrease in response to hyperosmolarity, a marker of DED,
potentially causing epithelial damage in DED. 84

Insulin insufficiency, as found in diabetes mellitus, has adverse effects on the

ocular surface. A study in South Africa found significantly worse tear film
parameters and a higher frequency of DED in children with diabetes mellitus
compared to healthy controls. 85 An investigation in Turkey found that obese
children had lower tear meniscus parameters and worse TBUT and Schirmer
scores than healthy children, and these results correlated with insulin resistance.
86
A study in India showed worse ocular surface parameters in those with
diabetes mellitus compared to healthy controls. 87

Additional research has linked the DED in diabetes mellitus to insulin impairment.
For example, elevated serum opioid growth factor levels in diabetes are
associated with DED and corneal damage, which can be mitigated by controlling
glucose levels with insulin or opioid receptor antagonists. 88 Topical insulin
therapy, in turn, has shown promise for treating the signs and symptoms of DED.
2,89-91

For insulin topical therapy to be widely adopted for DED, questions about the
optimal concentration, vehicle, and suitable DED subgroups need to be
addressed. 92 Novel formulations using nanotechnology to improve insulin
permeability and exposure to the ocular surface have shown promise in
experimental models. 93,94

5.7 Thyroid hormone regulation of the ocular surface and adnexa

Thyroid eye disease (TED) or thyroid autoimmune orbitopathy, includes a

spectrum of conditions like Hashimoto thyroiditis, Graves’ disease, and Schmidt’s
syndrome. 95

The association between TED and DED is debated. A study in the USA found no
link between TED and DED. 96 However, studies in India, Saudi Arabia, Spain,
Taiwan, and Russia found TED to be a risk factor for DED. 97-100 The variable
clinical manifestations of TED (Figure 1) and different definitions of DED likely
contribute to these discrepancies.

18
Figure 1. Thyroid eye disease

The mechanisms that cause DED in TED are broad and include anatomical
changes in the orbit leading to proptosis and excessive corneal exposure, as well
as inflammation of the ocular adnexa, including the main lacrimal gland and the
eyelids. 101 Animal models deprived of thyroid hormone have shown clinical
responses consistent with DED. As an example, rats made hypothyroid with
methimazole developed higher tear film osmolarity and hypoesthesis. 102

In TED, changes in the lacrimal gland have been observed via magnetic
resonance imaging, with smaller lacrimal glands correlating with more severe
clinical signs and higher inflammatory indices. 103 A study on the overlap of
Hashimoto’s thyroiditis and Sjögren disease found higher expression of four
genes involved in both diseases, which may have diagnostic value. 104 These
observations highlight the role of anatomical changes, genetic factors, and
molecular inflammation mediators in linking TED, thyroid hormone dysfunction,
and DED.

Recent studies have confirmed higher Ocular Surface Disease Index (OSDI)
scores, lower tear TBUT, and MGD in TED patients, compared to healthy

19
controls. These symptoms and signs were correlated with worse proptosis,
higher Clinical Activity Scores, incomplete blinking, as well as corneal damage.
100,105-108
Changes were also observed in euthyroid or inactive TED, indicating
mixed inflammatory and anatomical factors. 109,110 Superior limbic
keratoconjunctivitis was observed more frequently in TED patients (31%) and
was associated with worse ocular surface conditions, younger age, and smoking.
111
Worse Schirmer test results, TBUT, and goblet cell density were found in
children with Hashimoto’s thyroiditis compared to healthy children, even without
symptoms. 112 In a study of 38 individuals with moderate to severe TED in Iran,
over 70% had DED. 113

Non-invasive imaging techniques have recently improved the diagnosis and

monitoring of TED. Magnetic resonance imaging and computed tomography,
commonly used to assess orbital parameters in TED, are now used to study
lacrimal gland volumetry and activity. 114,115 Evaluation of the lids and meibomian
glands has shown an association between MGD scores and clinical activity
scores in TED. 116

5.8 Gender and DED

As reported in the Sex, Gender, and Hormones report of TFOS DEWS II, 3
"gender" refers to an individual's self-representation as a man or woman, and
how social institutions respond to that person based on the person's gender
presentation. 3 Gender is not the same as "sex." Sex distinguishes males and
females based upon their biological characteristics, whereas gender reflects
socially constructed characteristics such as behaviors related to being a woman,
feminine, or being a man, masculine. 3 Both sex and gender affect health and
disease, and gender also affects people's access to and interactions with the
healthcare system. 3 Many health disparities are associated with gender, and
both gender and biological sex influence DED risk and presentation, care-
seeking behaviors, and service utilization.

Transgender individuals experience difficulties accessing appropriate health

care.117 Since that report, a case series reported ocular findings in a transgender
and gender diverse population receiving gender-affirming hormone therapy. The
treatments involved daily doses of estrogen and spironolactone or testosterone
to 10 male-to-female and 7 female-to-male individuals, respectively. The major
findings included intracranial hypertension in female-to-male and chorioretinal
diseases in male-to-female. Regarding the ocular surface, one female-to-male
patient developed dendritic keratitis, two male-to-female patients had prior DED,

20
and two others were diagnosed with de novo DED due to low tear film TBUT and
punctate keratitis. 118

Providing appropriate healthcare for transgender and gender diverse individuals

is challenging due to a lack of specialized knowledge and social and cultural
barriers. Clinical research and education for healthcare professionals are
essential to address these challenges. 119-121 The interactions of sex hormones
with ocular tissues and their documented effects on systemic health indicate that
the impact of gender-affirming hormone therapy on vision and ocular health
needs further investigation (Table 1). This includes exploring potential
associations with DED and other ocular surface diseases and developing specific
preventive and therapeutic strategies, such as perceived in geriatrics and
gastroenterology conditions. 122,123

Table 1. Systemic effects of gender-affirming hormone therapy with

potential impact on ocular surface health and DED.

Reference Gender-affirming Systemic Relationship with

hormone therapy manifestation DED and ocular
type surface disease
Betsi et al., 2024 Puberty Bone mass Anxiety and
124
suppression with retardation, mood Depression
GnRH fluctuation
Heng et al., 2024 Testosterone Breast atrophy and Potential risk of
125
reduction of breast other exocrine
epithelia glands atrophy
Hashemi et al., Testosterone Higher risk for Potential impact of
2024 126 metabolic syndrome Metabolic syndrome
on DED
de Silva et al., Estrogen Venous Ischemic damage to
2024 127 thromboembolism ocular and adnexal
tissues
Nieves-Rios et Estrogen and Corneal
al.,2023 118 spironolactone for epitheliopathy
MTF or attributed to HSK in
Testosterone for FTM and DED and
FTM Punctate keratitis in
MTF
Tienforti et al., Hormonal Sex hormone Sexual hormone
2024 128 imbalance, imbalance and DED
menopause,
andropause
DED: dry eye disease, FTM: female-to-male transgender; GnRH: gonadotrophin release hormone;
HSK: herpes simplex keratitis; MTF: male-to-female transgender

21
 5.9 Future directions

There have been significant research advances linking sex, hormones and
gender to DED. Aging, cancer and hormone therapy increasingly broaden the
interdisciplinarity in this field over time. Despite the significant impact of gender-
affirming hormone therapy on the entire endocrine system and its effects on
physical and mental health, there is limited information on its impact on ocular
health. Variations in age, health profile, gender-affirming hormone therapy
compliance, and barriers to accessing regular healthcare limit the documentation
of side effects. Clinicians and future research should consider these variations,
as recommended in a recent systematic review on the medical aspects of the
transgender and gender diverse population. 123

6. Epidemiology

6.1 Scope of the update

This epidemiology update aimed to assess and summarize knowledge on the

prevalence and incidence of DED from well-designed population studies and to
perform a meta-analyses of existing study data to determine prevalence of DED
using different diagnostic approaches, stratified by age and sex.

6.2 Operational definitions

As per the TFOS DEWS II report, 129 the subcommittee examined data from a
large range of cohort studies and considered different methods of disease
ascertainment and definition, including studies involving the type, frequency and
severity of symptoms, patient self-report of a diagnosis of DED by an eyecare
practitioner, and studies that involved a clinical examination.

An updated search of published peer-reviewed literature was conducted using

PubMED (https://pubmed.ncbi.nlm.nih.gov/) which includes MEDLINE for articles
that reported the prevalence or incidence of DED. The following terms (dry eye
syndrome, OR dry eye disease, OR meibomian gland dysfunction, OR
keratoconjunctivitis sicca, OR blepharitis) AND (prevalence, OR epidemiology,
OR incidence) were used to identify potential additional articles. Human studies
published since the date applied in the previous TFOS DEWS II update (18
September 2015 onwards) were considered for inclusion. For the meta-analyses,
human studies captured in the TFOS DEWS II meta-analysis report and those
published subsequently were included.

22
Eligible studies included those reporting prevalence of either or both dry eye
symptoms and signs. Observational studies (cross-sectional or cohort) were
included if they were population-based and presented the study outcome as DED
versus non-DED. Studies were excluded if no variance in the measure of
prevalence was available in the manuscript, if it was not possible to calculate it
from the data presented, if the sampling criteria were not explicitly stated or if no
denominator was reported. Detailed exclusion criteria are described in Figure 2.
Article author and date, setting (region; population or hospital), numbers and
characteristics of participants within each study group (age, sex, ethnicity),
prevalence, and incidence data were extracted from each article. When required,
data were extracted from manuscript figures using open-source software
(available at https://plotdigitizer.com/app). 130 Where multiple studies were
published from the same dataset, those with minimal overlap, distinctly different
diagnostic criteria and age/sex disaggregation were preferentially chosen.

A meta-analysis was conducted to determine the prevalence of DED for different

diagnostic criteria stratified by age and sex. Using the search strategy described
above, population-based prevalence studies published since 1980 (per TFOS
DEWS II 2015 search) were included. Prevalence data were extracted firstly by
age group as follows; 6-9 years, 10-19 years, 20-29 years, 30-39 years, 40-49
years, 50-59 years, 60-69 years, 70-79 years and above 80 years and secondly
by sex. For studies that had age categories that overlapped with the decade-
based categorizations (e.g. 35-44, 45-54, 55-64 etc), the weighted averages of
data from each contributing interval were computed for each decade. Confidence
intervals for the measures of prevalence were computed by using standard
methods for computing standard error of a proportion, or, if prevalence was zero,
by computing the Poisson 95% confidence interval, dividing it by 2 to provide an
estimate of the standard error as above. Studies were combined where the
diagnostic criteria were broadly similar and in line with the 2017 approach and
included a new category for studies consistent with the diagnostic methods
described in the TFOS DEWS II Diagnostic Methodology Report 131 as follows:

1. Women’s Health Study criteria

2. Symptomatic DED (OSDI above 13 or significant ocular or visual
symptoms, where signs are not reported)
3. Insurance claims data supporting diagnosis or treatment of DED
4. Symptoms and signs (e.g., fluorescein staining, TBUT, and Schirmer
score)
5. Diagnostic criteria broadly aligned with that described in the TFOS DEWS
II Diagnostic Methodology Report. 131

23
 6. Clinical diagnosis or prior diagnosis of DED
7. Any MGD reported and separately, clinically significant (grade 2 and
above, as reported) MGD

A random effects model was used to combine prevalence data. To compute the
standard error, the formula SE = √p*q/n, where p was the proportion with DED
was used. For studies where prevalence was 0, the exact Poisson confidence
limits for the proportion computed and the width of that interval divided by 3.92 to
approximate the standard error.

For descriptive analyses, box plots were produced using SAS version 9.5 (SAS
Institute, Cary, North Carolina, USA). For most diagnostic categories, the number
of studies available for each age category was small. When there was only a
single study in the category, the prevalence is reported as originally reported by
the study. If there were two or more studies in the category, a pooled rate was
computed using a suite of SAS macros models developed by Weir and Senn.
(Senn et al., 2011) These macros include summary statistics for the DerSimonian
and Laird. (DerSimonian & Laird, 1986) The Weir and Senn macro forest was
used to produce forest plots of the results.

The flowchart describes the studies identified since 2015 and reasons for
exclusion (Figure 2). A total of 2687 articles were identified through PubMed
search (updated 29 June 2024). The extracted data summary is available here
(Supplementary Table 1. Studies included in the meta-analysis). For the overall
prevalence estimation and meta-analysis, the final dataset combined both 2015
and 2024 results. Subsequently meta-analyses were conducted by age and sex
by decade where data were available.

Estimates of the prevalence of DED from 76 large international cohort studies (24
and 52 from the 2015 and 2024 datasets, respectively) are summarized in Figure
2 and below. Supplementary Table 1 describes the characteristics of the
additional included studies from the 2024 dataset.

24
Figure 2. Prisma flowchart of the literature search outcome for prevalence
studies published between 18 September 2015 and 29 June 2024.

Table 2 summarizes the overall prevalence range by diagnostic criteria and

Figure 3 disaggregates the prevalence by age and sex where there were
sufficient data and includes data from both the 2015 and 2024 datasets.

Table 2. Prevalence range for DED for each of the diagnostic criteria

Diagnostic criteria Prevalence range (%)*

Women’s Health Study criteria 2.7 - 30.1

Symptomatic DED 7.3 - 31.6

Claims data 2.8 - 8.5

Symptoms and signs of DED 4.7 - 62.9

TFOS DEWS II criteria 5.4 - 44.2

Clinical or prior diagnosis of DED 1.0 - 15.3

Any MGD 0.0 - 66.3

Clinically significant MGD 1.8 - 23.3

25
 *Noting the lower end of the range relates to the rates in children. See Figure 3 below for
prevalence disaggregated by age

Figure 3. Prevalence of DED based on age and sex for different diagnostic
criteria

6.3 Prevalence of DED

6.3.1 Prevalence of DED based on the Women’s Health Study criteria

Figure 3, panel A represents prevalence by age and sex based on report of severe
symptoms and/or a diagnosis of DED by a practitioner (n=12). 132-143 The prevalence
ranges from 2.7% in those 20-29 to 30.1% in women over 80. The rate increases by
age particularly after the age of 40 in both sexes and women have a higher rate of DED
above the age of 50, with the sex difference in prevalence becoming more marked with
age. One study has shown high prevalence rates in those 10-19 years old without sex
related differences. (See supplementary Table 1 for individual study data).

6.3.2 Prevalence of symptomatic DED

Figure 3, panel B represents prevalence of symptomatic DED by age and sex. The
prevalence ranges from 7.3% in those 6-9 years old to 31.6% in females aged 20-29
years, however confidence intervals are wide (n=33). 133-135,139-142,144-168 There were
higher than expected rates of disease in the age groups under 30, except a low rate in
children aged 6-9 years. Rates were consistent in the adult groups aged 40-80 years,
with rates in men in the order of 20% and women 30%. Higher rates were observed in
the over 80s age group and in all ages above the age of 10, women had a consistently
higher prevalence.

26
6.3.3 Prevalence of DED based on signs and symptoms

Figure 3, panel C represents prevalence of DED based on the presence of signs and
symptoms by age and sex (n=10). 97,132,133,142,146,152,158,162,163,165,169 The prevalence
ranges from 4.7% in those 6-9 years old to 62.9% in females aged 20-29 years,
however confidence intervals are wide. The rates of disease are reasonably consistent
across the adult age groups with some reduction in symptomatic disease above the age
of 70 in both sexes. Sex differences were not pronounced in most age groups except for
in those aged over 70 where women have a higher rate than men.

6.3.4 Prevalence of DED based on TFOS DEWS II criteria

The overall prevalence is broadly similar to diagnoses based on signs and symptoms of
DED (n=3) 132,133,170 with prevalence ranging from 5.4 (in 6-9 year olds) to 44.2%. Above
the age of 30, the prevalence is higher amongst females than males, and males show a
more obvious age-related change.

6.3.5 Prevalence based on claims data

Overall prevalence based on claims, either identifying DED based on the International
Classification of Disease code or using insurance claims data based on diagnosis or
treatment code was generally low (n=1) 171, ranging from 2.8-8.5%. There was
insufficient data to disaggregate prevalence by age and/or sex.

6.3.6 Prevalence of DED based on clinical diagnosis

Figure 3, panel D represents prevalence of DED based clinical diagnosis by age and
sex (n=8). 135,140,148,149,151,155,172,173 The prevalence ranges from 1.0% in men over 80, to
15.3% in women aged 50-59 years. Rates were reasonably consistent over age in
adults with lower rates in those 10-15 and over 80 years of age. Women had a higher
rate of clinically diagnosed DED at all ages.

6.3.7 Prevalence of any MGD

Figure 3, panel E represents prevalence of any MGD by age and sex (n=6). 97,132,174-177
The point estimates of prevalence ranges from 0% in those under 20 years old to 66.3%
men over 80. Rates appear to markedly increase above the age of 40 and in 40 plus
age groups, MGD is significantly more prevalent in older men aged 70 and above than
in older women (p<0.05). Confidence intervals in most age groups are wide.

6.3.8 Prevalence of clinically significant MGD

27
Figure 3, panel F represents prevalence of clinically significant MGD (Grade 2 or above)
by age and sex (n=2). 165,175 Rates appear to increase with age although sex differences
are equivocal. There are no studies reporting rates of clinically significant MGD in
younger individuals.

Thirty-three of 52 studies included in the systematic review from the 2024 dataset did
not contribute to the meta-analysis due to non-availability of by age decade or by sex
data, or due to duplicate data. 20,23,97,140,141,161,162,164,166,167,169,177-198 The overall
prevalence rates reported include all international cohort studies (24 and 52 from the
2015 and 2024 datasets, respectively). Meta-analysis includes only those studies
publishing disaggregated age and sex data. Supplementary Table 1 shows the
complete list of studies and data extraction for those identified in the recent dataset. The
2015 dataset is included as supplementary data previously. 129

6.4 Annual incidence of DED

A limited number of population studies have attempted to assess the incidence of DED
since the last TFOS DEWS II update. In a retrospective analysis of approximately 6.7
million medical claims from the United States Department of Defense Military Health
System, the annual DED incidence in those aged 2 years and above was low but to
gradually increase over time from 0.55% in 2008 to 0.87% in 2012, respectively. 171
DED annual incidence was consistently higher in women than men and increased with
age. 171

A similar retrospective analysis of electronic medical records of 1,458,830 new patients

presenting to Indian hospitals across four states was conducted and revealed an
annualised incidence (average across 8 years) of DED signs and symptoms of 1.46%.
199
An analysis of the Taiwanese National Health Insurance Research Database
between 2001 and 2015 (covering over 23 million inhabitants) yielded an age-adjusted
annual incidence of DED ranging from 0.15% (in 2001) to 0.37% (in 2015). 200

An ancillary cohort study of the Vitamin D and Omega-3 Trial (VITAL), which assessed
the incidence of DED in 12,174 men aged 50 years and older and 11 349 women aged
55 years and older during a median (range) 5.3 (3.8-6.1) years of follow-up in the United
States. All participants were initially disease free and 2% of participants experienced a
clinically incident DED. 201 The annualised incidence (average across 5 years) of
clinically diagnosed DED and of clinically diagnosed DED plus incident reports of severe
DED were 3.8 and 16 per thousand, respectively. 201 The Salnés Eye Study 2 re-
examined a cohort of 264 Spanish individuals from the Salnés Eye Study 1 now aged
51 years and older, 11 years later. 202 The annualised incidence of DED signs and
symptoms in these individuals was 2.3% (95% confidence interval 1.8-2.8). 202

28
Participants (n=1,682) from the Singapore Malay Eye Study (SiMES) were re-examined
6-years later, and symptoms of DED were evaluated using the Salisbury Eye Evaluation
Study dry eye questionnaire. 203 The 6-year incidence of DED symptoms was 5.1% (95%
confidence interval 4.1-6.4). 203

The incidence of DED is typically difficult to estimate as some with the disease will
report resolution or a reduction in symptoms over multiple sampling periods. This may
particularly occur for milder cases of DED where symptom report may vary over time.
Incidence from claims data may be influenced by changes in reimbursement or the
introduction of new therapies during the study.

6.5 Natural history of DED

Longitudinal studies of DED are rare. A cohort of 784 patients from 1000 with DED from
the Women’s Health Study and Physicians’ Health Study were surveyed about change
in their disease one year after enrolment in the study and medical records were
obtained for 261 of the participants. 204 The mean disease duration was 10.5 years, and
most participants reported no change to their disease status over time. Ocular surface
symptoms were unchanged in 32% of participants and improved in 44%. Visual
symptoms were unchanged in 52% and improved in 19%. Social impact was
unchanged in 71% and improved in 19%. Risk factors associated with progression of
ocular surface symptoms included a higher spend on treatment (more than $20 per
month), history of more severe DED symptoms and use of systemic beta-blockers.
Worsening of visual symptoms was additionally associated with a history of ocular
surgery, untreated depression and blepharitis or MGD. Disease presentation and
treatments varied by sex where women were more likely to present with corneal staining
but worsening of symptoms was not predicted by corneal staining, however treatments
beyond level 1 205, higher symptoms and having a tear breakup test performed were
associated with progression of symptoms.

One retrospective study followed clinic-based participants with a diagnosis of either

Sjӧgren (n=101) or non-Sjӧgren (n=101) DED over a 7-year period. 206 In this arguably
more severe disease group, with escalation of treatment, predominantly with use of
topical anti-inflammatory/immunomodulatory therapies or in office treatments, both
groups experienced a significant reduction in ocular surface staining although no patient
reported-outcomes were recorded.

In a small retrospective study of DED in the absence of treatment (n=73), 207 changes in
signs were observed with an increased bulbar redness, reduced tear meniscus height
and lipid layer thickness after 8 years. Change in symptoms over time was not reported.

29
In a registry-based study, signs of MGD and evaporative DED were apparent at an
earlier age than aqueous-deficient DED (Wang et al., 2020; Wang et al., 2019),
although there may be some confounding due to ethnic background, where up to 2/3 of
those with signs of MGD do not experience symptoms (non-obvious disease). 175 There
is indirect evidence to suggest that signs of MGD may precede other disease markers
by up to 10 years (Wang et al., 2020), which may have implications for the timing of
treatment of non-obvious MGD. In a small treatment trial of adults with symptomatic
MGD who were treated for 12 months, 1/3 of participants had improvement in MGD
signs and improvement was greater in younger participants (those under 40) and with
less ocular surface damage. 208

In summary, there is reasonable evidence for development of signs before symptoms in

untreated disease, which may impact on treatment considerations. There is evidence
that treatment improves corneal staining but limited data on the sequence of
improvement of signs and symptoms. Longitudinal natural history studies are much
needed.

6.6 Risk factors for DED

Conclusive and probable risk factors for DED grouped into major categories described
in Table 3 and have been divided into modifiable and non-modifiable factors.

In clinical practice, administering a questionnaire to the patient (such as via paper or a

digital application) for completion before the consultation is recommended to save time
and to ensure that all commonly associated factors have been addressed. The following
section briefly highlights the most important risk / associated factors by category.
Irrespective of a possible causal mechanism, all associated factors described may be
useful in elucidating the etiology of DED and seeking to identify the possible driver(s) in
an individual.

Table 3. Risk factors associated with DED.

Risk factor for DED Evidence level* Modifiable?

C= consistent, P= M= possibly
probable modifiable, N =
non modifiable

1. Systemic disorders

a. Autoimmune disorders
209
Sjögren disease C N

30
 209,210
Rheumatoid arthritis C N
209
Systemic sclerosis C N
209,211
Systemic lupus erythematosus C N
209
Sarcoidosis C N
210,212,213
Thyroid disease C N

Inflammatory bowel disease (Crohn’s disease and ulcerative C N

214,215
colitis)
216
Psoriasis C N

b. Hormone disorders or status

210,212,217
Diabetes C N
218-220
Androgen deficiency C M
221
Polycystic ovary syndrome P M

c. Dermatological and atopic disorders

212,222,223
Acne rosacea C M
214,224
Acne vulgaris P M
212
Eczema C M
225
Asthma C N
226
Allergy C M

d. Pain disorders
227
Irritable bowel syndrome C N
227
Fibromyalgia C N
214,228,229
Chronic pelvic pain C N
227,230
Migraine C N
230
Other headache disorders C N
212,214
Osteoarthritis C N
227
Back pain C N
228
Temperomandibular joint disorder P N

e. Psychiatric disorders
210,212,231-233
Depression C M

31
 231,232
Anxiety C M
212
Stress (including post-traumatic stress disorder) C M
214
Burnout P M
214
Autism P N

f. Sleep disorders
234
Obstructive sleep apnea syndrome C M
141,235
Insomnia and poor sleep quality C M

g. Other disorders
212
Osteoporosis C M
236
Parkinson’s disease C M
214
Sinusitis P M

2. Ophthalmic disorders
237,238
Meibomian gland dysfunction C M
239-241
Anterior blepharitis C M
226
Allergic conjunctivitis C M
212
Glaucoma C M
214,242,243
Facial nerve paralysis (Bell’s palsy) C N

Ocular surface disorders interfering with the tear film or C M

inducing inflammation (e.g. pterygium, pingueculum,
160,212,244-250
conjunctivochalasis)
251-254
Thyroid eye disease (including Graves’ disease) C M
222,223
Ocular rosacea C M

3. Surgery or procedures

Eye surgery (e.g. refractive surgery, cataract surgery, C N

intravitreal injections, glaucoma surgery, retinal surgery)
212,214,255-261

261,262
Eyelid and periorbital surgery C N
261,262
Periocular botulinum toxin injections C N
261-263
Cosmetic periocular and ocular procedures P N

Hematopoietic stem cell transplantation (graft-versus-host C N

264,265
disease)

32
 4. Medications

Anticholinergic medications (antihistamine, antiarrhythmic, C M

bronchodilator, antidepressant, anti-Parkinson’s, and
261,262,266-268
antispasmodic medications)

Eye drops (e.g. preservative-containing, anti-glaucoma, C M

262,266,267,269
antihistamine, anaesthetics, some NSAID’s)

Anticancer drugs (e.g. chemotherapeutic agents and C M

262,270-274
hormone therapy)
275-277
Vitamin A derivatives (including isotretinoin) C M
278-280
Dupilumab C M
213,218,262
Hormone replacement therapy P M
218,281,282
Anti-androgen therapy C M
214
Proton pump inhibitors P M
214
Psychostimulant agents used for ADHD P M

5. Environment
283
Air pollution from NO2 and CO P M
283
Low humidity C M
283
High or low temperatures P M
283
Air conditioning and wind P M
212,262,284
Contact lens wear C M
212,285-287
Screen use C M
263
Cosmetics C M

6. Demographic factors
212,288-290
Increasing age C N
212,288-290
Female sex and gender C N
212,291
Asian race / non-white race C N

7. Nutrient / gene related

292
Vitamin A deficiency C M
292
Vitamin B12 deficiency P M
292
Vitamin C deficiency C M

33
 292-294
Vitamin D deficiency P M
292
Omega-3 fatty acids deficiency C M
292,295-297
Altered gut microbiome P M
136,298,299
Genetic predisposition P N

ADHD = Attention deficit hyperactivity disorder.

*Consistent evidence implies the existence of multiple adequately powered and otherwise well-conducted
studies published in a peer-reviewed journal, along with the existence of a plausible biological rationale
and corroborating basic research or clinical data . Probable evidence implies the existence of at least one
adequately-powered and otherwise well-conducted study published in a peer-reviewed journal. Risk
factors with mixed or inconclusive results from multiple studies or from a systematic review or meta-
analysis are described in the Diagnostic Methodology Report. Non-modifiable = a lack of evidence that
change will impact DED.

6.6.1 Systemic disorders

Numerous systemic disorders (Table 3) have been associated with an increased risk of
DED, emphasising the multifactorial origin of the disease.

Autoimmune disorders are well-established risk factors of DED. Up to 90% of patients

with Sjögren disease develop DED 209,300. Rheumatoid arthritis, systemic lupus
erythematosus, and systemic sclerosis are also associated with a vastly increased risk
of DED 209,211,301. Other autoimmune diseases that have been consistently linked with
an increased risk of DED are sarcoidosis, thyroid disease, inflammatory bowel disease
(Crohn’s disease and ulcerative colitis), and psoriasis 209,210,212,214,215,302,303. Less
commonly investigated autoimmune disorders, also associated with DED in some
studies, include lichen planus, lichen sclerosus, granulomatosis with polyangiitis,
polyarteritis nodosa, primary biliary cirrhosis, mixed connective tissue disease,
antiphospholipid syndrome, and dermatomyositis 209,214,240,301,303-305. Infiltration of the
lacrimal gland and accessory lacrimal glands by lymphocytes and other immune cells
that leads to impaired tear secretion and aqueous-deficient dry eye has been suggested
as a core mechanism. However, there is no peer-reviewed evidence for inflammation in
the meibomian gland in obstructive MGD 240,306.

The endocrine system plays an important role in maintaining ocular surface

homeostasis. Hormone disorders such as androgen deficiency, polycystic ovarian
syndrome, and thyroid disease have been linked to an increased risk of DED
212,213,218,221
. Sex hormones exert effects on all major components of the tear film 218. In
thyroid disease, immune-mediated lacrimal gland dysfunction, and exposure
keratopathy due to orbitopathy (such as in TED), are important mechanisms. 213,218,307
Diabetes has also been consistently, although mildly, associated with DED in
systematic reviews and meta-analyses (odds ratio (OR) of around 1.2) 210,212,217.

34
Decreased corneal sensitivity and impaired reflex tear secretion have been suggested
to be linked 308 alongside factors such as hyperglycemia, advanced glycated end
product accumulation, oxidative stress, metabolic disease and vascular disease 309.
Menopause has also been proposed as a risk factor for DED, but definitive evidence is
lacking 129,218,310.

In addition to psoriasis, other dermatological disorders may also be associated with an

increased risk of DED. Rosacea without systemic involvement may present with ocular
symptoms alone and can lead to chronic blepharoconjunctivitis and MGD (see Section
3.5.3.3.3)212,222,223. Acne vulgaris has been linked to DED, possibly by an associated
risk of MGD or as a result of side-effects of isotretinoin therapy 214,224. Atopic disorders
such as eczema, asthma, and virtually all types of allergy have been consistently linked
with an increased risk of DED 212,214,225,226,283. Allergic eye disease may aggravate DED,
and suggested mechanisms include increased inflammation at the ocular surface,
altered epithelial barrier and corneal innervation, and tear film instability. Antihistamine
use, which may result in anticholinergic side-effects, may also partly explain the
association. Periocular eczema has also been linked to blepharitis. Alternatively, DED
may aggravate allergic eye disease, since allergens may be less efficiently removed
from the ocular surface in the presence of reduced tear turnover and DED is associated
with an upregulation of inflammatory mediators, including some of those common to the
allergic pathway, such as complement. Complement also plays a role in DED in the
absence of allergy 311. Complicating the association between DED and allergy is the
overlap in symptoms such as burning, itching, and tearing. 225,226,312 It is therefore
important to consider allergic conjunctivitis in patients with DED, and particularly in
patients with a history of atopic disease. Specific questioning about eyelid margin
itching may suggest the presence Demodex blepharitis 313.

Chronic pain disorders (the body's heightened sensitivity to non-noxious stimuli) can
present with ocular discomfort symptoms similar to those of dry eye.; fibromyalgia,
chronic pelvic pain, irritable bowel syndrome are among the most established risk
factors 227. Development of neuropathic pain within the trigeminal somatosensory
system has been proposed to underlie this link 227. A study of twins in the United
Kingdom found evidence for shared genetic factors underlying these disorders and DED,
suggesting a common chronic pain predisposition that accounts for clustering of pain
disorders 299. Other pain conditions that have been linked to DED are migraine and
other headache types, osteoarthritis, back pain, and temporomandibular joint disorder
214,227,228,314
.

There is also strong evidence for an association between DED and depression, anxiety,
burnout, and stress (including post-traumatic stress disorder) 212,214,227,231-233. A meta-
analysis of studies in patients with DED found an overall prevalence of 40% for

35
depression (OR of 1.8), and of 39% for anxiety (OR of 2.3) 231. These relationships are
likely bidirectional, and studies show an association with psychiatric disorders with DED
symptoms that is greater than with signs. Proposed mechanisms that link psychiatric
disease to DED include altered pain perception, somatization, and increased serum
inflammatory markers 227. Clinicians may find it beneficial to enquire about mental health
history in patients with DED, as appropriate, and consider referrals to mental health
professionals when indicated.

Obstructive sleep apnea has been linked to DED 212,235. It increases the risk of floppy
eyelid syndrome, which may lead to exposure keratopathy at night and to increased
conjunctival inflammation and MGD 315. Leakage of air from a continuous positive
airway pressure (CPAP) mask may also lead to irritation, conjunctivitis and DED 316,317.
Insomnia and poor sleep quality may be a consequence of DED, but could also result in
DED 141,227,235. For example, obstructive sleep apnea is associated with a persistent
low-intensity inflammatory state which may be pertinent to DED etiology 318.

Other disorders that have been convincingly linked with DED are osteoporosis (which
may reflect common underlying mechanisms such as sex hormone or vitamin D
deficiency, or a role of purinergic signalling) 319, autism (which may reflect
hypersensitivity to external stimuli) 214, Parkinson’s disease (which may be linked to
decreased blink rates and lower tear secretion) 236,320 and sinusitis (which may
sometimes involve the tear ducts, or reflect common mechanisms like allergy, or may
represent referred pain) 214.

6.6.2 Ophthalmic disorders

MGD may be the most important factor associated with DED 237. Its prevalence varies
widely between studies, but has an estimated global prevalence of 35.8%, and the
majority of patients with DED have underlying MGD that is considered to be a major
cause of evaporative DED 237,238,321. Signs of MGD occur earlier in the natural history of
DED progression (typically between 24-29 years) than other clinical signs such as tear
film instability, hyperosmolarity (31-38 years) and ocular surface staining (46-52 years)
322
.

Anterior blepharitis is a common chronic inflammatory disorder of the eyelid margin

located anterior to the gray line (see section 3.5.3.3.1). This area includes the eyelid
skin and the eyelashes and its follicles. Anterior blepharitis is traditionally
subcategorized based on its etiology: bacterial, seborrheic, fungal or parasitic. Anterior
blepharitis is a risk factor for other forms of ocular surface disease including DED, MGD,
chalazion and keratitis 239,323.

36
Other disorders of the ocular surface have also been associated with a disruption in the
tear film or inflammation at the ocular surface, potentially leading to DED (see Section
X). Conditions that have been most associated with increased risk of DED are
pterygium 160,244,324, pingueculum 245-247, conjunctivochalasis 248-250, and allergic
conjunctivitis (see section 3.1.2.1.1).

Patients with glaucoma are also at increased risk for DED, most often associated with
use of antihypertensive eye drops. Both active ingredients 269 and preservatives can
affect the ocular surface. In addition, glaucoma surgery can affect the ocular surface.
212,325,326

Facial nerve paralysis, including Bell’s palsy, can lead to exposure keratopathy through
incomplete blinking, and has also been associated with impaired lacrimation and MGD
214,242,243
. Facial nerve dysfunction may lead to lagophthalmos and reduced blink force,
compromising tear film distribution and postulated to compromise meibomian gland
expression 327.

6.6.3 Surgery and other procedures

Eye surgery has consistently been linked to an increased risk of DED. The greatest
amount of evidence is available for refractive surgery, 256-258,328, cataract surgery 255,259
and intravitreal injections, 260 but it is likely that all eye surgery comes with an increased
risk of DED. 212,214,261,262 Mechanisms that lead to DED after eye surgery include
incisional procedures leading to transectional nerve damage, surgical trauma to the
ocular surface during surgery, phototoxicity, toxicity from eyedrops (e.g. from povidone
iodine, anaesthetics, or preservatives), and damage and stress from repeated drying,
irrigation and exposure to surgical illumination. 261,329,330

Eyelid and periorbital surgery, such as blepharoplasty, ptosis surgery and brow surgery,
have also been associated with an increased risk of DED. 261 Mechanisms include
lacrimal gland injury and post-operative incomplete eyelid closure and incomplete
blinking. Similarly, periocular botulinum toxin injections can lead to lagophthalmos. The
TFOS Lifestyle Report discusses the impact of eye, eyelid and periocular procedures on
the ocular surface in further detail. 261,263

Allogeneic hematopoietic stem cell transplantation, mostly used in the treatment of

various haematological diseases, can lead to graft-versus-host disease. In this disease
the graft’s immune cells attack the host’s body cells, and this can become chronic. It
can affect multiple organs including the eyes and principally involves the ocular surface
which can lead to severe DED. More than 50% of recipients develop DED, mostly 6 to
24 months after transplantation 264.

37
6.6.4 Medication use (See Section 10)

A large population-based study in the Netherlands found that 52 of the 99 most

commonly used medications were associated with an increased risk of DED. 266
Although not all these medications may be causally linked to DED, and some of the
associations may reflect an association with underlying disease (severity), it highlights
the importance of assessing medication status in patients with DED. This section
discusses various medications that have been most frequently associated with DED, but
it is recognised that medication-associated DED is not limited to these medications only.

Medications with anticholinergic (side-)effects have been consistently and causally

linked to DED, such as antihistamine, antiarrhythmic, bronchodilator, some
antidepressant, anti-Parkinson’s, and antispasmodic medications. 261,262,266-268 These
drugs affect the muscarinic receptors of the lacrimal glands and conjunctival goblet cells,
causing decreased aqueous and mucous secretion.

Several medication-related pathophysiological mechanisms have been demonstrated:

allergic, toxic, immune-inflammatory effects; chemical interaction with the tear film
leading to a disrupted lipid layer; reduced aqueous secretion; damage to goblet cells
and epithelium of the cornea and conjunctiva; and neurotoxic effects on the corneal
nerves and eyelids including meibomian glands. 261,262,274,325,326

Systemic chemotherapeutic and other anticancer agents may have cytotoxic effects at
the ocular surface and affect tear film quality and reflex tear secretion. 262,270-274,331
Examples include alkylating agents such as cyclophosphamide, antimetabolites such as
5-fluorouracil and methotrexate, monoclonal antibodies such as rituximab and the
aromatase inhibitors.

Vitamin A derivatives or retinoic acids, including isotretinoin, are used for acne vulgaris
and in anti-ageing regimes, and may be administered topically or orally. They are
secreted into the tear film by the lacrimal gland 332, and are associated with tear film
instability, lower Schirmer test scores, and atrophy of meibomian glands, leading to
DED. 263,275-277,306,333

Interleukin (IL)3 and 4 receptor antagonists (such as dupilumab) are increasingly used
in patients with atopic dermatitis with good effect but may cause several adverse effects
at the ocular surface including a mild to sometimes severe conjunctivitis, punctate
keratitis, blepharitis, and loss of meibomian glands. 278-280

Hormone replacement therapy has been associated with increased prevalence of DED.
213,218,262

38
Proton pump inhibitors, antacids, and psychostimulant agents used for attention-
deficit/hyperactivity disorder have also been linked to a highly increased odds of having
DED, but biological pathways are currently unclear. 266,267

6.6.5 Environmental factors

Climatic risk factors may play an important role in the etiology of DED. The TFOS
Lifestyle Environmental Conditions report has recently and comprehensively
summarized evidence for environmental risk factors for DED. 283 Both low and high
temperatures have been associated with DED, as has low humidity, in several
experimental and population-based studies. Air-conditioning and wind are well-known
risk factors that patients often report as triggering symptoms, 334 although limited
evidence is available from research studies. Air pollution such as from NO 2 and CO was
found to be probably associated with DED, and soil pollution from chromium is likely
associated with DED and Sjögren disease. Evidence for a risk of other air, soil, and
water pollutants was not conclusive. 283

Contact lens wear has been reported as a factor associated with DED in many cross-
sectional population-based studies, but prospective studies are lacking. It has been
postulated that mechanisms leading to DED may include thinning of the tear film after
insertion of the contact lenses, increased friction between the lens and the ocular
surface, leading to meibomian gland dropout, decreased tear film stability, ocular
surface staining and lid wiper epitheliopathy. 238 It is important to recognise that contact
lens discomfort symptoms overlap with DED, and symptoms do not necessarily reflect
underlying DED. 212,262 (See Section 10 Contact lenses and DED)

Screen or visual display terminal use has been consistently linked to DED. 285-287,335 As
few as 1 to 2 hours of screen use per day may even be associated with adverse ocular
surface effects. 285 Important mechanisms include decreased blink rate and incomplete
blinking that leads to increased evaporation and decreased lipid release from
meibomian glands. 336 It is also important to consider digital eye strain, independently of
DED, in the differential diagnosis of persons who are heavily exposed to a digital
environment. 286 This requires a full refractive correction to be determined for the
distances required and a binocular vision assessment to ensure suboptimal visual input
is not the cause.

Cosmetic products may be associated with adverse effects at the ocular surface and
may aggravate or initiate DED symptoms. The TFOS Lifestyle cosmetics report
identified 10 ingredients that are commonly present in cosmetics that particularly have
significant adverse effects: benzalkonium chloride, chlorphenesin, formaldehyde-
releasing compounds, parabens, phenoxyethanol, phthalates, prostaglandin analogues,
retinoids, salicylic acid, and tea tree oil. 263 Finally, recent reviews have found impacts

39
on the ocular surface from: environmental endocrine disruptors in foods, packaging and
pesticides. 337 No clear or consistent link between smoking/vaping and DED has been
identified, although effects on the tear film are evident. 227,338-341 While a direct causal
relationship between smoking and DED may not be certain, it is reasonable to suggest
that smoking cessation should be encouraged as part of a comprehensive approach to
promoting ocular surface and holistic health in patients with DED.

6.6.6 Demographic factors

Female sex and gender are strong risk factors for DED (see Section 6). As discussed in
Section 5, sex-related differences in prevalence are underpinned by differences in the
ocular surface and adnexa including anatomy and immunity, a higher prevalence of
autoimmune disorders and psychiatric disease (and their related medications), higher
general pain sensitivity in women and reduced androgen levels (already lower than in
men) after menopause. 218,289,342

Increasing age is also a well-established risk factor for DED, but DED is prevalent in
both the young and the old. Systematic reviews have found that signs show a stronger
relationship with age than symptoms. MGD, also, increases in prevalence with age.
288,321,343-346

Prevalence values for DED and MGD are higher in Asian countries than in Western
countries 289 and a pooled analysis of interethnic differences suggested differences in
disease type and age at which signs manifest in Asian and Caucasian populations. 291
There is also evidence that white races are less affected than non-white races in
Western studies. 212 It is difficult to unravel precise causes for these differences, which
likely reflect socio-economic, cultural, genetic, anatomical, lifestyle and environmental
differences between groups.

6.6.7 Other factors

Nutrition may play a role in the development of DED, but there is no strong evidence to
support recommendation of an optimal diet. Poor nutrition that drives systemic disorders
may be associated with risk factors for DED. Limited but increasing evidence links
alterations in the gut microbiome to ocular surface health, possibly by altering the
immune system. 292,296,297,347-350 The TFOS Lifestyle Nutrition report concluded that
there is evidence that deficiencies of vitamins A and C and omega 3 fatty acids are risk
factors for DED. Moderate evidence was also found for vitamin B12 and D deficiencies
as risk factors.292 There is no clear evidence that alcohol use, 227,292, a Mediterranean
diet 351 and water intake 292 represent risk or protective factors for DED.

40
Genetic factors likely contribute moderately to DED. A large twin study in the United
Kingdom established heritability accounted for 29% for DED symptoms, and 41% for
DED diagnosis. The remaining 60-70% was attributed to unique environmental factors.
136
A recent genome-wide association study in Taiwan with over 14,000 DED cases and
almost 26,000 controls found eleven independent risk loci, including MUC16, which
encodes for a mucin protein that is expressed at the ocular surface. A polygenic risk
score including 932 loci was able to detect individuals with a high-risk of DED. 298 A
limitation of this study was the use of self-reported DED only and that the findings were
not replicated in an independent cohort. This lack of replication is also a major limitation
for several, mostly small, candidate gene studies that have found a link between genetic
variations and DED. 352-358 Further studies are warranted before genetic testing
becomes clinically useful in DED.

6.6.8 Risk factors for MGD

There is debate about the impact of MGD on risk factors and prevalence of DED. It is
recognised as a risk factor for the disease and recent estimates of DED prevalence
suggest that MGD may be present in 50-70% of cases of DED. 359,360 Nonetheless,
there have been attempts to establish demographic risk factors for MGD which may or
may not be independent of those for DED more broadly. Age is frequently reported as a
risk factor in MGD, (Arita et al., 2019) particularly in elderly populations 188 and where
MGD is diagnosed using meibography to determine meibomian gland loss or drop out.
11,361
Ethnic background (Asian compared with other backgrounds) is consistently
reported as a risk factor. The impact of sex in MGD is however equivocal, in contrast to
its impact in DED more broadly where female sex predominates.

In examining population-based studies, there is a higher rate of non-obvious MGD in

Caucasian males 175 and a higher risk of MGD in males based on gland plugging and
telangiectasis in the Singapore Malays study.(Siak et al., 2012) In a smaller population-
based study from Japan where symptomatic disease was diagnosed using lid margin
irregularities and gland plugging, males similarly had a higher risk of MGD compared
with females. (Arita et al., 2019) Male sex was not an independent risk factor for MGD in
a population-based study of adults in Iran.(Hashemi et al., 2017) There is wide
variability in the impact of sex on MGD depending on how the disease is diagnosed
using individual or combined lid signs (gland drop-out, orifice plugging, altered meibum
secretion, degree of gland expressibility, lid telangiectasia, either presence/absence or
presence above a certain level), whether symptoms are present, study design and
whether the study is based on a clinical sample, specific disease group or is population-
based.

41
For the meta-analysis reported above (Section 6.3.7), large population-based studies
only have been included and two analyses report either all MGD, irrespective of
definition and clinically significant MGD signs with symptoms, broadly based on the
TFOS MGD Report. For population-based studies reporting any lid changes with or
without symptoms (Any MGD – Figure 3, F), MGD is more prevalent in older men
(above 70 years of age) than older women but there is no difference between sexes in
the rates of clinically significant MGD (Figure 3, G).

6.7 Morbidity and impact

DED severely affects the lives of sufferers. It negatively impacts quality of life (QoL)
including physical, psychological and emotional well-being, social functioning, daily
living activities and independence. General and mental health, social functioning,
physical, emotional states, bodily pain and vitality are significantly poorer in those with
DED compared with those without 362-365 and that health status worsens in those with
more severe disease. 364,365 In individuals with mild and moderate disease (n =217)
blurred vision, productivity loss, and visits to eye care practitioners were increased
compared with age-matched normal individuals (n=67). 366

Adverse QoL effects appear to be consistent over time, irrespective of geography and
with variations in ethnic background 367,368 and will likely increase with population aging.

The economic cost of DED can be measured in direct resource utilisation (service
provision, medication costs), out-of-pocket costs, cost of lost productivity and reduced
QoL (quantified as a utility). There are limited studies which include an associated utility
algorithm or that meaningfully evaluate productivity impacts of DED.369-371

6.8 Summary and outstanding questions

This update has considered the prevalence of DED in studies that have shown disease
rates by age and sex. Eight major diagnostic groups were identified and meta-analyses
reported here include DED diagnosed using the Womens Health Study criteria,
symptom report; claims data from health or insurance databases; signs and symptoms;
diagnosis according to the TFOS DEWS II criteria; clinical diagnosis; any MGD or
clinically significant MGD (grade 2 or above). Prevalence varied with diagnostic criteria
where not all disease increased with age or showed a female predominance. Broadly,
using the Womens Health Study criteria, DED increased with age and was more
common in women. A clinical diagnosis of DED showed a female preponderance but
not an age-related effect. Symptoms and signs were more common in women with
higher rates in younger and older adults. Any or severe MGD was age-related, with
males more likely to show any MGD. Some studies included in the ‘Any MGD’ analysis
reported non-obvious MGD and the age and sex effects here have not been stratified.

42
These findings are perhaps not unexpected given the multifactorial nature of DED, the
specificity of ocular symptom measurements and differences in the etiology of different
subtypes of DED 1. Prevalence data in some recent studies may be confounded by the
impact of the COVID-19 pandemic. Previous meta-analysis showed that both the
pandemic and mitigating factors (mask and screen use) were associated with greater
ocular symptoms and signs of ocular surface disease. 26,145,191,197

Studies reporting rates of DED in those under 20 are limited. Rates are lower than
adults for clinically diagnosed DED, DED with signs and symptoms (under 10 years)
and any or significant MGD. High rates of symptom-reporting are evident in those under
20 however, although it is recognised that symptom report alone is not specific for DED
and childhood anterior or posterior blepharitis, Demodex and allergy may be common
co-morbidities or contributors to ocular symptoms. Most studies reporting symptoms did
not report signs although they may be present.

Risk factors are reported as consistent or probable, and potentially modifiable or non-
modifiable. New risk factors related to environment, climate and lifestyle are included.
There is some evidence for an increase in prevalence in DED over time which may
conceivably be due to the impact of new risk factors including changes in the digital
environment. Given the differences in prevalence and age/sex associations with
different diagnostic criteria for DED, it may be important to disaggregate risk factors
particularly for DED and MGD where possible. Given the high prevalence of symptom-
reporting in childhood, appropriate triaging for other conditions and hypothesis-driven
and appropriately powered studies to explore risk factors in children would be valuable.

Outstanding questions include:

1. Disease severity. A limited number of studies explored the prevalence, risk factors or
natural history by disease severity, which could help to triage and manage those
more likely to experience more severe DED.
2. Geographical mapping was not considered as part of this update.
3. The generalizability of prevalence measures for DED in children and adults under 40.
These mostly originate from studies in Asia and there are limited studies in other
regions.
4. The need for appropriately powered studies to determine risk factors in those under
40.

7. Pathophysiology

7.1 Introduction

43
The consensus view of the 2017 TFOS DEWS II report envisioned a tear film centric
model of the pathophysiology of DED, 6 broadly classified by compromised quantity;
Aqueous-Deficient Dry Eye (ADDE) or quality; Evaporative Dry Eye (EDE) of the tear
film. In DED, tear hyperosmolarity is considered to set up a cascade of signaling events
within surface epithelial cells, that leads to the release of inflammatory mediators and
proteases. Such mediators, together with the tear hyperosmolarity itself, are conceived
to cause goblet cell and epithelial cell loss and damage to the epithelial glycocalyx.
Damage is reinforced by inflammatory mediators from activated T-cells, recruited to the
ocular surface. The net result is the characteristic punctate epitheliopathy of DED and a
tear film instability which leads at some point to early tear film break-up. This break-up
exacerbates and amplifies tear hyperosmolarity and completes the vicious circle events
that ultimately lead to ocular surface damage and self-perpetuation of the disease.
Epithelial injury and a defective glycocalyx, loss of tear volume and of goblet cell mucin,
lead to increased frictional damage and friction-related symptoms. The tear
hyperosmolarity and epithelial injury caused by DED, stimulates corneal nerve endings,
leading to symptoms of discomfort, increased blink rate and potentially, a compensatory,
reflex increase in lacrimal tear secretion.” The TFOS DEWS II report also highlighted
causes of ADDED that included lacrimal gland infiltration and dysfunction,
neurosecretory or reflex blocks, androgen deficiency or aging-related downregulation of
secretion, obstruction of the lacrimal ducts in cicatricial disease, and iatrogenic causes
such as prescription medication and surgical damage to trigeminal nerves. For EDED,
conditions that affected the ocular surface included MGD, anterior blepharitis,
xerophthalmia, ocular allergy, androgen deficiency as well as iatrogenic causes such as
topical preservative use, contact lens wear and certain anti-glaucoma drugs.

Subsequent research has led to a more nuanced understanding of the disease. Given
the inconsistent relationship between symptoms of DED and signs such as measurable
inflammation, hyperosmolarity, ocular surface staining, low tear volume, low TBUT, or
MGD, 360,372 the data suggest that the associated signs help identify subtypes of the
disease, with symptoms providing no predictive ability as to which etiology is active in
that patient. 360 The implications of the lack of correlation of signs and symptoms in DED
are particularly important in relation to pathophysiology, because while most historical
literature assumes that inflammation and inflammatory pathways are the common
effector of DED, evaporative subsets show a muted if any increase in inflammatory
mediators in the tear film, 372-376 and thus inflammation cannot be assumed to be active
in every patient with DED. For example, one study based on mass spectrometry found
that evaporative patients exhibited no increase in proteins associated with the
inflammatory response compared to normal controls, while 51% of differentially
upregulated proteins in the aqueous cohort were associated with inflammation. 372
Similar data showed that while aqueous- deficient subjects (Schirmer value ≤ 5 mm)

44
had elevated lipid peroxides in tears compared to healthy controls, subjects with 6–10
mm wetting were not significantly different from normal controls. 373 Another study
based on multiplex bead analysis found that evaporative patients tend to exhibit much
higher levels of epidermal growth factor, but no difference in the IL-6 and IL-8 cytokines
compared to healthy controls and essentially no detectable TNF-α in tears, 375
reinforcing the idea that not all forms of DED are driven by the same underlying
pathways. In mouse models, bilateral lacrimal gland excision showed dramatic
increases in IL-1ß, IL-6 and TNF-α protein concentration in the tear film over four weeks,
while topical benzalkonium chloride and environmental chamber-induced DED showed
no significant difference from controls in those tear film cytokine proteins, despite
concurrent increases in tear osmolarity and cytokines measured from corneal mRNA
transcripts. 377 Immune-mediated etiologies, represented by immunological diseases
that include Sjögren disease, Stevens-Johnson’s syndrome, and ocular graft versus
host disease (GVHD), are widely regarded as being more severe than evaporative
counterparts. 378 At the extreme end, severe forms of dry eye found in ocular GVHD
tend to be accompanied by fibrotic processes, 379,380 and Sjögren-related DED is
associated with lymphocytic infiltration of the lacrimal gland, neither of which is evident
in common evaporative DED 380 In a rabbit model, cauterization of the meibomian
glands results in only moderate increases in hyperosmolarity and loss of goblet cells as
compared to models in which lacrimal excretory ducts and accessory glands of rabbits
are sealed. 381 These data may help explain why anti-inflammatory medications have a
dichotomous effect, with some patients reporting improvement, 382,383 while others report
high failure rates of cyclosporine and lifitegrast in the general population. 384-387 As many
patients are still poorly served by the available therapeutic options, it may be that our
understanding of the pathophysiology of DED is incomplete.

Note that a variety of the pathophysiological aspects of the disease are discussed in
more detail elsewhere. For example, androgen deficiency is discussed in the Sex,
Gender, and Hormones section, neuropathic damage is covered in the Pain and
Sensation section, the tear proteome is outlined in the Tear Film section and iatrogenic
causes are described in detail in the TFOS Lifestyle Report (Gomes et al., 2023) and in
the Iatrogenic section of the digest report.

7.2 Initiating triggers of disease

Except in certain situations of injury, 176 surgery 256,328 and therapy 388, establishing
causality is difficult when diagnosing DED. Metabolic disease induces mitochondrial
stress 389-394 and advanced glycation end products in the lacrimal gland 395,396, hormonal
changes that alter glandular production and fatty acid metabolism 3,397-400, and
biophysical stresses of friction 401-405, hyperosmolarity 406-414, and swelling pressure 415.
These effects are compounded by the biological activity of dysregulated or self-reactive

45
immune cells 416-419, cellular stresses from cytokines 377,420-426, proteases 427-429, reactive
aldehyde species430, extracellular deoxyribonucleic acid (DNA) and neutrophil
extracellular traps 431-433, exogenous toxins 390,411, damage or danger-associated
molecular patterns 408, gut dysbiosis 434,435, as well as neurogenic inflammation. 436-441
Typical pathways of initiation of evaporative processes include phenotypic alterations in
corneal epithelial cells that lead to a compromised glycocalyx 442, keratinization of the
meibomian gland that alters the lipid profile of the tear film, 443,444 and incomplete
blinking or reduced blink rate during screen use that exposes the ocular surface to
desiccating stress. 445 On the aqueous deficiency side, inflammatory ingress into the
lacrimal gland can be driven by androgen deficiency or autoimmunity, causing a
cascade of protease release, cytokine expression, inflammatory cell recruitment,
dendritic cell maturation, and an adaptive T-cell mediated response. 3,418,446 As disease
severity increases, the evidence suggests there is a progressive accumulation of these
mechanisms. 227,418,447,448

7.3 Hyperosmolarity

When exposed to hyperosmolar conditions, epithelial cells begin to change their

morphology and lose their microplicae. 449 Increasing levels of hyperosmolarity (excess
salt) on the ocular surface cause an increase in epithelial apoptosis, 450
transglutaminase mediated cornification of epithelial cells, 451 and increased secretion of
IL-1ß, IL-6, IL-8, TNF-α, and MMP-9 from ocular surface epithelial cells. 452-455
Hyperosmolarity-induced epithelial cell stress results in desquamation, revealing the
immature glycocalyx and microplicae-free cells beneath. 449 These immature cells are
hydrophobic and contribute directly to tear film instability. When patches of hydrophobic
cells are adjacent to normal hydrophilic cells, the differential surface tension causes the
tear film to breakup and evaporate within a few seconds, dramatically increasing the
local osmolarity and exposing cells to a toxic environment. 456,457

In addition to the hypothesis that inflammatory cells infiltrating either the lacrimal gland
or conjunctiva are a source of oxidative damage at the ocular surface, 458,459 In vitro
evidence suggests that hyperosmolarity directly induces reactive oxygen species
(hydroxyl and peroxyl activity) in human corneal epithelial cells, along with reductions in
the anti-oxidant superoxide dismutase-2, vitamin D, Notch ligands Dll3 and Jag 1, and a
tripling of the pro-apoptotic Bax/Bcl2 ratio. 460 Reactive oxygen species undergo lipid
peroxidation and cause reactive aldehyde species to be produced, causing a cascade
of protease release, cytokine expression, inflammatory cell recruitment, dendritic cell
maturation, and an adaptive T-cell mediated response 430,461. In vitro data have shown
that exposure to hyperosmolarity causes mitochondrial DNSA to leak into the cytoplasm
of human corneal epithelial cells, activating the cGAS-STING pathway and increasing
cytokines such as CXCL10 and IFN-β, 462 while human conjunctival impression cytology

46
samples from evaporative, short TBUT patients (2.9 s) confirmed an increase in STING
proteins compared to more normal controls. 463 Evaporative hyperosmolar patients (327
mOsm/L, 7.0 s TBUT, 10.4 mm average Schirmer value) exhibit specific upregulation of
IFN-γ, without concomitant increases in IL-2, IL-6, IL-10, TNF-a or IL-17A. 464 IFN- γ,
which is strongly associated with dose dependent CD40, MICA and MHC II expression
along with esithelial cytotoxicity in corneal epithelial cells, 465-467 increases NLRP3
oxidative stress and contributes to pyroptosis in human corneal epithelial cells under
hypertonic conditions. 468 These data mirror earlier findings that hyperosmolarity
disrupts the balance of oxygenases and antioxidant enzymes such as SOD1 and
PRDX4, stimulates lipid peroxidation (e.g., 4–hydroxynonenal & malondialdehyde),
increases COX2, and damages corneal mitochondrial DNA. 469,470 Hyperosmolarity also
potentiates the negative effects of the oxidizing blue light in corneal and conjunctival
epithelial cells, increasing H2O2 production, phototoxicity and altering the mitochondrial
membrane potential in these cells compared to those in normal media. 471 Downstream
of hyperosmolarity, a TLR-4 dependent upregulation of Dual oxidase 2, a member of the
NADPH oxidase family that regulates the production of intracellular reactive oxygen
species, increases alongside high mobility group box 1 production in human corneal
epithelial cells. 408,472 This result was consistent with data that MyD88-/- mice lacking
TLR-4 signaling exhibited significantly less corneal fluorescein staining, cytokine and
protease expression following 5 days of hyperosmolar stress and scopolamine
administration, 473 implicating TLRs and damage or danger-associated molecular
patterns (e.g., HSPs, HMGB1, S100A, tenascin-C) as essential agents in evaporative,
hyperosmolar DED pathophysiology. 473-475 Although inflammatory ingress may be an
eventual result of hyperosmolar exposure, 476 clinically, it is likely a matter of time and
severity that governs which patients progress to the inflammatory phenotype as a result
of hyperosmolarity. 381 For example, at low levels of hyperosmolarity, protective
responses such as the increase in IGF binding protein-3 are observed, but at increasing
levels of osmolarity and length of exposure, cell viability is challenged as respiration and
glycolysis are decreased alongside endoplasmic reticulum stress and caspase-3
activation, leading to unchecked mitophagy and eventual cell death. 393,477,478 Similarly,
short term exposure to low humidity environments showed an increase in damage or
danger-associated molecular patterns such as HSP-60 and a small 2.4-fold increase in
MMP-9 mRNA expression in patients with evaporative DED, but no statistically
significant increase in IL-6 or IL-8 expression. 474 These data suggest that the severity
of tear hyperosmolarity should help inform the underlying pathophysiology; wherein a
strongly elevated osmolarity will lead to oxidative stress and inflammation, while a
middling or low osmolarity is suggestive of a subclinical inflammatory state 479 and may
mitigate against prescription of anti-inflammatory therapy.

7.4 Proteases

47
Another aspect of the pathophysiology of aqueous deficient disease that separates it
from evaporative disease is the disrupted balance of protease and anti-protease activity
on the ocular surface. The normal tear film exhibits an equilibrium of protease and
protease inhibitors. 427 Excess protease is observed in Sjögren disease, where
cathepsin S catabolizes anti-proteases such as cystatin C, 428 MMP-9 is highly
expressed while thrombospondin-1 lags behind, 480 and plasmin activity is increased
ten-fold in Sjögren compared to normal tears. 481 In non-autoimmune DED, protease
activity is stratified by disease severity and is strongly associated with aqueous
deficiency. For example, mild evaporative-type subjects with low TBUT (5.0–7.2
seconds) and normal Schirmer value (13–19 mm) exhibited only slightly higher tear
MMP-9 activity (35-66 ng/mL), compared to the tears of more severe aqueous-deficient
cohorts, (Sjögren disease, Stevens-Johnson, 6 mm Schirmer value) with levels in the
101–381 ng/mL range. 378 These data are consistent with observations that 11-14% of
early stage, patients with EDE (4.9 second TBUT, 312 mOsm/L, 14.0mm Schirmer
value) presented with ELISA-validated MMP-9 positivity. 482 Increases in neutrophil
elastase, MMPs 483 and neutrophil extracellular traps are seen in severe aqueous-
deficient subjects (chronic ocular GVHD, Sjögren disease and ocular cicatricial
pemphigoid) that are not commonly observed in non-autoimmune DED and healthy
controls. 431 Similarly, the anti-protease Cystatin S was found to have a significant,
inverse relationship with aqueous severity in non-autoimmune DED subjects, falling
from about 2,000 ng/mL in controls (16 mm Schirmer value) to 400 ng/mL in moderate
aqueous-deficient DED (4 mm Schirmer value), while MMP-9 was only mildly
upregulated to the 20–40 ng/mL range in those subjects. 484 The increase in cathepsin
S, a serine protease that is known to degrade a variety of essential components of the
normal tear film and glycocalyx (lactoferrin, sIgA, proteoglycan 4), 428,485 is also stratified
by inverse tear volume; increasing monotonically as tear volume decreases. 376 Of note,
although cathepsin S is dramatically enhanced in Sjögren disease compared to non-
autoimmune subjects on average, non-autoimmune subjects with 0–5 mm wetting on
the Schirmer strip exhibited equal or higher levels of cathepsin S than patients with
Sjögren disease with > 15 mm of wetting. 376

These data invoke the question of causality, whether aqueous deficiency leads to
inflammation and protease release, or vice versa. In an animal model of severe
aqueous deficiency, excision of the lacrimal gland resulted in a significant upregulation
of serine proteases in corneal tissue including tryptase, urokinase plasminogen activator
receptor and protease activated receptor 2 expression, which upon activation, is able to
induce MAPK / ERK-1 & 2 signalling with downstream NF-κβ, ICAM-1 & cytokine
expression, 429 suggesting that aqueous deficiency can precede a measurable protease
burden at the ocular surface. In contrast, MMP-9 positivity was found in 84% of GVHD
as compared to 33% of non-autoimmune DED patients, even as the GVHD patients

48
trended towards increased Schirmer test values, lower OSDI and higher TBUT than the
DED controls, 486 suggesting that immune infiltration into the lacrimal gland precedes
the protease release in GVHD.

Ultimately, in non-autoimmune DED, the transition from low levels of protease release
to inflammatory DED with excessive protease concentrations is likely related to
progressive alterations in the lacrimal gland and ocular surface. These might result from
androgen deficiency, 3,6 accumulation of advanced glycation end-products in the
lacrimal gland, 395 diabetic keratopathy and nerve degeneration, 487 or other factors that
lead to significantly less aqueous flow, thereby tipping the homeostasis towards a
severe hyperosmolar state and associated excessive, pathogenic protease release. It is
recognised that in marked EDE or lipid deficiency, tear and ocular surface homeostasis
may be sufficiently disrupted to cause inflammation and that in advanced disease,
features of both are displayed.

7.5 Sub-functional or absent glycocalyx

The glycocalyx lubricates, 488 retains water at the ocular surface, 489 provides wetting to
the ocular surface, 490 acts as a barrier and helps remove debris from the ocular surface.
491
The hydrophilic epithelial surface provides 82.5 sec/cm of specific resistance
compared to 12.9 sec/cm for the lipid layer, 489 establishing the glycocalyx as one of the
components responsible for preventing evaporation. Electron micrographs of
intracellular, pre-expression glycocalyx showed that “the increase in numbers of
subsurface vesicles that occur in some external eye diseases may reflect an attempt to
increase the binding of the mucus to the eye surface… In late keratoconjunctivitis sicca,
however, the subsurface vesicles are absent. 492” These foundational observations
identified the difference between evaporative DED and other ocular surface conditions,
where exogenous stresses lead to a compromised epithelial glycocalyx – the proximal
cause for an increase in evaporation, hyperosmolarity and its downstream
complications in EDE.

Evidence that the glycocalyx plays a central role in disease pathogenesis has begun to
accumulate; for example, knockout of MUC4, the most abundant membrane associated
mucin in the conjunctiva with an ectodomain predicted to extend > 2 µM above the
apical cell surface, resulted in significantly reduced microplicae, tear film disruption, and
increased rose bengal dye penetrance into deeper layers of the ocular surface. 493 In
vitro evidence from corneal and conjunctival co-culture has shown that hyperosmolarity
(caused by a decreased blink rate) directly downregulated components of the epithelial
glycocalyx, increased TLR-4 expression, initiated release of cytokines and proteases
including MMP-9, and eventually, cellular apoptosis. 445 The hyperosmolarity
compromised glycocalyx led to significantly lowered TBUT, lower Schirmer value, and

49
persistently elevated fluorescein staining, whereas replenishment of the glycocalyx
using human recombinant lubricin (recombinant human proteoglycan 4) was able to
reverse these clinical indicators, as well as inhibit epithelial NF-kB translocation and
normalize IL-8, TNF-a, IL-1ß, TLR-4 and MMP-9 expression within one day of
supplementation. 445 A striking aspect of this study is that the common clinical
expressions of DED were recapitulated without immune cell involvement, establishing
the hyperosmolarity-compromised glycocalyx as a causal, initiating event in evaporative
disease. A related study found that intraperitoneal injections of streptozotocin, used to
induce metabolic hyperglycemia similar to type 1 diabetes in mice, caused dramatic
reductions in the extent of the corneal glycocalyx, with associated reductions in tear film
volume, number of goblet cells and upregulation of TLR-4, MAPK, IL-1, IL-6 and IFN-γ
genes within as little as one week following injection. 494 In a C57BL/6 mouse model,
scopolamine and desiccating stress induced an almost complete abrogation of secreted
(MUC5AC, MUC2) and transmembrane mucins (MUC1, MUC4, MUC15 & MUC16)
along with significant increases in conjunctival IFN-γ, leading to persistent epithelial
defects. 495 Importantly, an in vitro study evaluating the impact of applied hyperglycemia
in media (15 & 30 mM glucose vs. 5 mM control) showed that elevated glucose
exposure had no effect on the amount or distribution of membrane associated mucins in
either corneal or conjunctival epithelial cells, 496 suggesting that the pathophysiology of
streptozotocin-induced glycocalyx loss is more likely due to global metabolic disease
than exposure to hyperglycemia alone. In support of these data, a study of vitreo-retinal
surgery found that conjunctival MUC4 & MUC16 gene expression increased post-
surgery and tear osmolarity was reduced in normal subjects, likely as a protective
response, but older diabetic subjects exhibited a lower goblet cell density, lower
MUC5AC and increased cytokine response by comparison, once again linking systemic
metabolic disease to ocular surface glycocalyx impairment. 497 In a mouse model of
GVHD, allogeneic transplantation of a mixture of spleen and bone marrow cells resulted
in a significant reduction in the area and thickness of the corneal glycocalyx, reductions
in MUC4 and MUC5AC and a coincident reduction of tear film volume and an increase
in fluorescein staining, which were partially abrogated after application of topical
rebamipide. 498

More recent data have shown that components of the glycocalyx also regulate immune
cells499, transduce extracellular environments to intracellular signaling pathways 500, and
actively inhibit proteases such as MMP-9 501, which would further implicate the
catabolism or downregulation of the glycocalyx as an initiating event in DED
pathogenesis. As the glycocalyx is altered in ocular surface pathology, large decreases
in sialic acid 502 and increases in galectin-3 are observed in the tear film that strongly
correlate with disease severity. 503,504 When released into the tear film, galectin-3 seems
to amplify the IL-1ß mediated inflammatory response, 505 which is similar to how

50
degraded low molecular weight hyaluronic acid becomes pro-inflammatory in other
tissue systems. 506,507

In summary, hyperosmolarity, metabolic disease and inflammation impair the ocular

surface glycocalyx and initiate the characteristic clinical signs of DED. The degree of
impairment of the glycocalyx seems to be a fulcrum for transitioning from common
evaporative DED to an inflammatory, aqueous-deficient state, as the balance of fluid
output overwhelms the fluid input.

7.6 Meibomian gland dysfunction

Meibomian gland dysfunction (MGD) is a chronic, diffuse abnormality of the meibomian

glands, commonly characterized by terminal duct obstruction and/or qualitative and
quantitative changes in the glandular secretion. 237 Although it is a disease distinct from
DED, when the severity of MGD is of a sufficient degree, it may give rise to EDE. 323
While a widely cited study is often misquoted to suggest that 86% of all DED subjects
have MGD, the denominator of that estimate excluded patients with DED that could not
be classified as either aqueous or evaporative, and when included, found that 60.7% of
those with DED had evidence of MGD. 508 Larger, more recent estimates of the
prevalence of MGD within those with DED estimate a number, from 51.3% to 70.3%.
359,360
Those with pure MGD exhibited the lowest severity of signs of ocular surface
damage, 359 consistent with the assumption that evaporative DED is often a subclinical
inflammatory state. 372 Indeed, in patients with MGD, significant negative correlations
were found between inflammatory mediators such as IL-8, C5a and Schirmer value
results. (Mahajan et al., 2021) One study however has suggested there may be
inflammatory mediators and cytokines present in meibum in patients with MGD 509. As a
causative pathophysiology of DED, it is currently estimated that between 33% to 50% of
DED patients have sufficiently impactful MGD to initiate a dry eye state. 360

By leveraging novel models of primary human meibomian gland epithelial cells, ductal
from human tissue, 510,511 3D cultures and organotypic cultures, 462,512-517 researchers
have further investigated the pathophysiology of MGD. PPARγ agonists, such as
rosiglitazone, may significantly promote cell differentiation, lipogenesis and anti-
inflammation in human meibomian gland epithelial cells. 168,518-522

The meibomian gland is located in a physiologically hypoxic environment 523, that is

impaired in MGD. 524 An increased expression of hypoxia-inducible factor 1α (HIF1α)
plays a role in the beneficial effect of hypoxia on the meibomian glands, 525 and as such,
a low oxygen environment may be important in imitating the in vivo condition in culture.
Animal models include injection of complete Freund's adjuvant in rabbits 526 and mice,
527
electrocauterization of meibomian gland orifices in rats, 528,529 transitory alkali

51
exposure of the rat eyelid margin, 530 Soat1-null mice, 531,532 APOE KO mice, 533 and
Elovl1-deficient mice. 534

Recent studies have indicated that it may be possible to restore gland structure after
atrophy. In mice, fibroblast growth factor receptor 2 gene (FGFR2) knockout could lead
to significant gland acinar atrophy, 535 but this change is reversible if the knockout
condition is removed. Moreover, this recovery relies on the extent of ductal atrophy,
which indicates that ductal epithelia may serve as a reservoir for meibomian gland
progenitor cells for regeneration. 536 The importance of FGFR2 and other FGFRs were
also reported in humans, in which FGFR inhibiting anti-cancer drugs could induce
significant gland atrophy, and patients who use these drugs may develop MGD. 537

The relationship between systemic lipids and MGD has been a focus of several recent
studies. Patients with elevated serum total cholesterol, low-density lipoprotein and
triglyceride levels exhibited significantly higher levels of meibomian gland loss
compared to healthy controls. 538 Mice fed with a high-fat diet developed hypertrophic
meibomian glands, 539 decreased PPAR-γ expression, increased meibomian gland acini
cell apoptosis and mitochondrial damage, and activation of MAPK and NF-κB signaling
within the gland. 540 A high fat diet significantly altered the rhythmicity of meibomian
gland, which may offer new insights into the regulation of the glands by dietary lipids. 541
Patients who have long term dyslipidemia also showed significant meibomian gland
atrophy and changes in meibum quality, even while undergoing statin treatment. 542
Dietary cholesterol has a direct impact on meibum components and meibomian gland
pathophysiology. 543-545 In contrast, dyslipidemia and increased triglyceride levels were
found to be protective factors for meibomian gland atrophy in an elderly female
population. 546 A systematic review has suggested there is moderate evidence for a
beneficial effect of omega-3 supplements on MGD. 547

Meibum proteins also play an important role in ocular surface homeostasis. A

discussion of the role of meibum proteins in MGD is beyond the scope of this review but
these have been described more fully in Jeyalatha et al., 2017. 548 Ectodysplasin A
protein secreted from meibomian glands plays a significant role in regulation the
proliferation of corneal epithelial cells through the epidermal growth factor receptor
signaling pathway. This discovery connects meibomian gland function with corneal
epithelial homeostasis, 549 and suggests future research pathways surrounding the
feedback between the eyelid and ocular surface.

7.7 Inflammatory cell recruitment

By leveraging novel single-cell RNA sequencing (scRNA-seq) methods, CD45+

lymphocyte populations were quantified in mouse corneas before and after short term
oral scopolamine hydrobromide and desiccating stress. T cells (18.6%), resident
52
macrophages (18.2%), B cells (12.8%), type 2 conventional dendritic cells
(cDC2)/macrophage (9.9%), NK cells (9.3%), monocytes (8.9%) and neutrophils (7.9%)
were the most prominent cell types in normal corneas, whereas a striking shift towards
resident macrophages (55.2%), MMP12 and MMP13 high macrophages (11.5%) and
type 2 conventional dendritic cells (cDC2)/macrophage (8.5%) were observed following
the induced hyperosmolar ADDE eye. 550 In the mouse conjunctiva, pharmacologic
suppression of tear secretion followed by desiccating stress recruited monocytes from
the blood and promoted maturation of Ly6ChiMHClo monocytes to Ly6CloMHCIIhi
macrophages. 551 In a hyperosmolar model that exposed mice to a low humidity
environment without the scopolamine administration, scRNA-seq revealed that
conjunctival CD4+ T cells, Mo/Mφs, and DCs were amplified during DED progression,
CD8+ T cells gradually decreased, and epithelial cells exhibited EMT-like characteristics
that created a positive feedback loop of pro-inflammatory MφC3 activation. 552
Importantly, although mouse models tend to echo human DED in terms of innate or
myeloid immune cells (macrophages, dendritic cells, neutrophils), lymphoid cells such
as NK, NKT, or γδ, or adaptive CD4+, CD8+ or Th17 lymphocytes do not necessarily
follow the same trajectories between species. 553 For example, brush cytology of human
conjunctival tissues revealed an increase in IL-17A producing γδ cells from ≈ 4% in mild
evaporative disease (3.3s TBUT, 26 OSDI) to 7–10% of the CD45+ cells in more
symptomatic human samples (2.0s TBUT, 49 OSDI), which was far lower than the
increase from 25% to 33% of CD45+ cells in normal versus induced DED mouse
conjunctivas. 554 Of note, conditional knock down of lymphangiogenesis in a
scopolamine, dessicating stress murine model resulted in the significant reduction of
TNF-α, IL-1ß, IFN-γ and IL-8 within corneal tissues but did not lessen the release of
those cytokines in the lacrimal gland, highlighting the importance of angiogenic immune
cell trafficking in the progression of aqueous DED. 555

7.8 Immune cell dysfunction

Dysfunctional regulatory T cells (Tregs) exhibiting reduced Foxp3, CD24 and CTLA-4
expression lose the ability to suppress the chronic Th17 mediated inflammation in an IL-
6 dependent manner. 556 Blocking the substance P / neurokinin-1 receptor interaction,
439
or the TLR4/MyD88 pathway 557, both of which are upstream of NF-κB, were shown
to normalize the Treg/Th17 balance. In mice colonized with intestinal microbiota from
patients with Sjögren disease, a reduced frequency of CD4+Foxp3+ T regulatory cells in
cervical lymph nodes was observed 558, while in aged mice, CD4+CD25+Foxp3+ T cells
were dysfunctional, lost suppressive ability, and produced significant amounts of
inflammatory cytokines IL-17 and IFN-γ. 559

7.9 Future directions

53
Recent studies exploring the pathophysiology of DED have continued to define the
etiologically distinct subpopulations within the disease, recognizing limited evidence for
inflammation in evaporative DED, and expanding roles for metabolic, hormonal,
physical and molecular impacts on the ocular surface. Neural regulation of the
epithelium is taking on increasing importance, as are factors from the ocular surface
that damage nerves, such as hyperosmolarity, reactive oxygen species, and other
potentially excitotoxic stressors. Yet major questions remain about how to best classify
the subsets of disease within a single patient and to what extent observed pathways are
causal to symptomatology. The natural history of the disease is unclear, when or if
inflammation becomes involved, or when acute inflammation transitions into a chronic
state. While animal models of the pathophysiology have provided valuable mechanistic
insights, they often fail to fully replicate human disease progression. If, as the existing
data show, inflammation is most strongly associated with aqueous deficiency, whether
this due to a relative increase in hyperosmolarity compared to that seen in evaporative
disease, or whether inflammatory mediators and proteases from the diseased lacrimal
gland driving downstream changes in the epithelium underpin these effects. In part due
to these questions about pathophysiology, it is currently not possible to predict which
patients will respond to anti-inflammatory or other types of therapy and which ones will
show positive responses to lubricants. While yet to be established, there may be value
in evaluating human immune cell dynamics and cell populations using advanced
imaging methods such as functional in vivo confocal microscopy. 560 As the diversity of
drugs expands, it will be increasingly necessary to align etiology with mechanism of
action. Hundreds of millions of dollars of development rests on predicting responders
from non-responders. Thus, it is becoming ever clearer that classical indices of DED
such as corneal staining and TBUT are insufficient to support modern drug development
efforts, and that more informative methods of determining disease subtype are critical
for future research.

8. Tear Film

8.1 Introduction

The Tear Film Report of TFOS DEWS II described the biophysical and biochemical
aspects of tears and how these change in DED. 561 The report noted that DED is
characterized by loss of tear volume, more rapid breakup of the tear film and increased
evaporation of tears from the ocular surface. The tear film lipids, proteins, mucins and
electrolytes contribute to the integrity of the tear film but exactly how they interact was
an area of active research. Tear film osmolarity, proteins and mucins could potentially
be used as biomarkers for DED. Some of these themes have been explored by
research since the publication of the TFOS DEWS II report, and these are highlighted in
the current report.

54
For the current report, the authors undertook searches in Pubmed, Scopus and Web of
Science databases for keywords ‘tear film’ and ‘dry eye’ since the publication of the
TFOS DEWS II Tear Film Report in 2017. After reviewing the titles and abstracts of
documents obtained, the authors have concentrated on those areas that have produced
the greatest number of articles. This report focusses on concepts and more detail on
specific components can be found within the cited references.

8.2 Clinical measurements of the tear film

The reproducibility of non-invasive TBUT (measured without fluorescein) obtained with

combinations of several commercially available instruments have been reported.
Instruments generally had good agreement, 562-566 but measurements from different
devices are not interchangeable. Reproducibility tended to worsen for those with DED,
564
Another study showed that NIBUT, whether measured by small-cone (E300) or
large-bowl (K5M), had poor agreement with FBUT, even when precise timing of blinking
was taken into consideration, and Bland-Altman analysis showed limits of agreement
that spanned the entire dynamic range of the instruments (31.4 seconds) with an
average difference of 3.9 seconds between the methodologies, making the techniques
neither interchangeable nor accurate compared to FBUT, and not interchangeable with
each other. 567 One of the challenges in interpreting the reported correlations of signs of
DED is the influence of subjectivity and low inter-rater reproducibility.

Simple correlations between symptoms and tear film or ocular surface characteristics
were sought. Whilst correlations do not imply causation, they can point the way to a
greater understanding of the mechanisms involved in signs and symptoms of DED and
identify areas for further investigation, recognising that there may be confounders in
individual studies and populations may differ. There was no correlation between tear
evaporation and symptoms (OSDI score), nor between tear evaporation and tear
production. 568 However, one study found significant positive correlations between OSDI
and tear evaporation rate scores in subjects with refractive errors. Both OSDI and tear
evaporation rate were lower in those without refractive errors. 569 These findings may
have been confounded by the inclusion of participants with uncorrected refractive errors,
smokers or differences in other demographic factors, but it may be important to adjust
for refractive error in studies of tear film and symptoms.

OSDI score has been negatively correlated with NITBUT, 563,570-574 tear meniscus height
(TMH) 570,571 and meibomian gland area, 574 and positively correlated with meibomian
gland loss. 573,574 NITBUT was positively correlated with TMH, meibomian gland dropout
grade 570,571 and corneal or lid margin staining. 563 Females with refractive errors had
significantly lower tear film lipid layer (TFLL) thickness, TMH, and NITBUT scores than
those without refractive errors.570 A strong correlation was found between TMH and

55
NITBUT, but no correlations between these parameters and OSDI scores.575 These
differences may be due to difference between populations and between techniques.
TBUT has been correlated with meibomian gland irregularity determined from
meibography images taken with a keratograph.576

Tear meniscus height was significantly correlated with the Schirmer value and tear film
breakup time and negatively correlated with ocular surface staining score, but there
were no correlations with any MGD indicator. 571

A study of normal subjects and those with MGD found a positive correlation between
precorneal tear film thickness and thinning rate and negative correlations between
precorneal tear film thickness and TFLL thickness and between precorneal tear film
thinning rate and TFLL thickness. 577. There were no associations between TFLL
thinning rate and any of precorneal tear film thinning rate, precorneal tear film thickness
or TFLL thickness.577 TFLL thickness has been positively correlated with age,
meibomian gland expressibility, and negatively correlated with meibomian gland dropout
572,578
and OSDI score.572

Several studies have been conducted to determine whether tear film osmolarity
correlated with clinical signs and symptoms of DED. Across population-based studies,
tear osmolarity is generally not correlated with tear film breakup time, corneal
fluorescein staining score, lid hyperemia, tear production, blink interval, Ocular
Protection Index, Schirmer I test, meibum expressibility, meibum quality or MGD. 579-583
However, this may depend on how DED was classified, as other studies found
correlations between tear osmolarity and OSDI discomfort subscore, corneal and
conjunctival staining scores 584 OSDI score, ocular surface staining and Schirmer I.585
Data from the DREAM (Dry Eye Assessment and Management) study, whilst
demonstrating some correlations between tear osmolarity and signs and symptoms,
were interpreted as being not indicative of causation as changes in tear osmolarity were
not associated with changes in signs and symptoms of DED. 586 Other recent
longitudinal studies, however, have suggested a relationship between change in tear
osmolarity with treatment and iatrogenic effects changes, suggesting a link in certain
DED subtypes. 587-592 The Diagnostic Methodology Subcommittee report of TFOS
DEWS III 1 gives more background on correlations and the limitations of this approach,
and the hyperosmolarity section of the Pathophysiology section of the TFOS DEWS III
Digest report (Section 7) discusses the role of osmolarity in more detail.

8.3 The tear lipids – composition and function

Several studies have examined the tear film lipid layer (TFLL). These have examined
changes to the lipids in DED and/or MGD patients compared to controls, as well as
which lipids contribute to tear film stability and reduced tear evaporation.
56
Broadly, meibum, combinations of meibum lipids and a thicker TFLL were associated
with a significantly slower evaporation flux, 593-596 and significantly slower tear breakup.
597
Tear film proteins can also reduce evaporation of water, and this is enhanced when
some of the proteins (human serum albumin and lactoferrin) but not others (human
lysozyme and bovine mucin) are used in conjunction with mixture of polar and non-polar
lipids found in meibum. 598 Lysozyme can reduce the surface tension of lipids on water,
by disrupting the order of lipid molecules. 599

Patients with MGD have significantly lower peak height ratios of the CH3/CH2 bands in
NMR studies than normal subjects, and this was hypothesized to be due to changes in
branched hydrocarbon chains, which contain fewer CH2 moieties, and straight chain
hydrocarbons, which contain more CH3 moieties. 600The authors further hypothesized
that, as van der Waals interactions between CH2 moieties were responsible for lipid
ordering, the more CH2 in meibum from MGD patients could contribute to a more
ordered TFLL. A more ordered TFLL can contribute to a more patchy layer resulting in a
deterioration in spreading and decreased elasticity. 600 Another factor that is involved in
TFLL stability is saturation of lipids, with increased saturation resulting in stiffer, thicker,
and more elastic films at high surface pressures. 601 Lipid saturation is a major factor
that contributes to lipid disorder and the phase transition temperature of lipid layers. 602
Meibum of MGD patients has lower surface pressures in Langmuir trough experiments
compared to meibum in those without MGD, and formed more brittle unstable patchy
layers. 603

The cholesteryl ester to wax ester ratio decreases in patients with MGD. 604-606 An
optimal mixture of wax and cholesterol esters in the TFLL may be necessary to disrupt
the ordered packing of pure lipid species, leading to better lipid spreading, 607 a more
stable tear film, 608 and thinner lipid layer patterns. 609 In a model system, mixtures of
cholesteryl oleate (CE) and behenyl oleate (a wax ester mimic) plus phosphatidylcholine
increased the surface pressure of films on phosphate buffered saline. 610 A layer of
behenyl oleate can form a crystalline state on water and reduce its evaporation 611 Iso-
branched wax esters help to reduce evaporation and increase surface pressure. 612 On
the other hand, addition of cholesterol to cholesteryl esters increases film rigidity. 613
People with thin TFLLs (who tended to be those with EDE) and people with irregular
TFLL patterns (who tended to be those with ADDE), had lower levels of cholesteryl
esters and lysophospholipids, but higher levels of glycerolipids and phospholipids in
their tears than those with normal TFLL patterns. 614

Other meibum and tear film lipids that have been examined are the O-acyl-ω-hydroxy
fatty acids (OAHFAs). Ultra-long OAHFAs found in TLLF and meibum are capable of
integrating other species in the polar lipid layer thereby molding its properties and/or
providing a base for the creation of hierarchical structures within the TFLL. 615 In vitro,

57
OAHFAs can spread well on the surface of phosphate buffered saline to form a solid
monolayer with a crystalline structure. 616 Longer chain OAHFAs prevented evaporation
of water at a mean molecular area of approximately 18 Å2/molecule. 616

Mixtures of OAHFAs and wax esters may produce even more evaporation resistant
films. 616 Indeed, wax esters existing in the solid state under physiological conditions
were capable of forming a mixed condensed monolayer with OAHFAs, and such a
monolayer exhibited very high evaporation resistance in vitro. 617 Whilst OAHFAs can
induce stable multilamellar cholesterol ester films on water or phosphate buffered saline,
618
OAHFAs alone reduced evaporation, with cholesterol esters having no effect. 618 An
additional double bond in the hydroxy fatty acid chain of OAHFAs was accompanied by
more disordered molecular organization, which led to a loss of the evaporation
resistance.619 The ultralong chain lengths observed in naturally occurring OAHFAs may
require the presence of double bonds to achieve an appropriate balance between
spreadability and evaporation resistance.619 The observations of ordered lipids resulting
in deterioration of spreading and reduced elasticity, whereas a more disordered
molecular organisation leads to a loss of evaporation resistance require further
exploration of how the ordering of different types of or combinations of fatty acids can
optimise the evaporative resistance, spreadability and elasticity of the film.

Individuals with symptomatic MGD or symptomatic mixed MGD/ADDE had reduced

abundance of several OAHFA species in tears, and to a lesser extent in meibum,
compared to normal subjects. 620 Other meibum-derived OAHFAs had negative
correlations with precorneal tear film thinning rate, and one meibum-derived OAHFA
had a positive correlation with precorneal tear film thinning rate.621 Conversely, tear film-
derived OAHFAs had no association with the precorneal tear thinning rate.621

There have also been several studies examining the contributions of sphingolipids to
the TFLL and meibum. Meibomian gland loss has been associated with loss of some
sphingosines from the TFLL. 609 Individuals with poor meibum quality had changes to
their sphingolipids in tears and meibum.622,623 The presence of both sphingomyelin and
ceramide increases surface tension due to the change their position in the TFLL under
lateral pressure.624 Sphingomyelin at the interface of the TFLL with the aqueous phase
has a role in capturing the protein lysozyme and entrapping it in the TFLL.625

Finally, several experiments have shed important light on how meibum lipids are
synthesized and the consequences of disturbance in their synthesis. Using mice with
specific genes knocked out has shown that fatty acid w-hydroxylase Cyp4f39, acyl-CoA
reductase FAR2, the elongases of very long chain fatty acids- 1i or -3, alcohol
acyltransferases Awat1 and Awat2, and sterol O-acyltransferase 1 are all involved in
various stages of the production of meibum lipids such as OAHFAs, wax esters and

58
cholesterol esters. 626-633 Some of the major findings when genes for these enzymes
were knocked out was shortening of tear film break-up time,626 plugged meibomian
gland orifices, tear film instability and increased tear evaporation, 627,631 increases in
blink frequency and evaporation from the ocular surface, 628, shorter chain, branched
and unsaturated cholesterol esters 629, and changes in the melting temperature of their
meibum.629,630

8.4 The tear proteome

Recent findings continue to emphasize the importance of the tear proteome as a non-
invasive tool for trying to discriminate between types of DED 634,635 and also for disease
monitoring, for instance revealing changes in the levels of proteoglycan 4, in tears of
those with Sjögren disease. 636

One interesting study explored the potential of tear MUC5AC and IL-8 levels to
distinguish between Sjögren and non-Sjögren DED, indicating that lower MUC5AC and
higher IL-8 levels could serve as biomarkers for Sjögren syndrome, aiding in its
diagnosis.637 Similarly, the tear thrombospondin-1/matrix metalloproteinase-9 ratio is
significantly reduced in Sjögren disease compared to non-Sjögren DED, offering
another potential biomarker. 480 Additionally, the MMP-9/lactoferrin ratio was positively
correlated with ocular inflammation and tear film stability in stable controlled patients
with Sjögren disease. 638

Tear biomarkers related to ubiquitination (LMO7 and HUWE1) and the regulation of
intracellular vesicle dynamics (TPD52) are significantly elevated in patients with Sjögren
disease, suggesting a role for these pathways in the underlying mechanisms of the
disease. 639 Other robust protein biomarkers have been identified using LCMS/MS in
multiple cohorts, highlighting several proteases and protease inhibitors plus noting the
relevance of oxidoreductase proteins in Sjögren disease. 640 Oxidative stress
biomarkers serve as indicators for assessing the extent of ocular surface damage, and
research using both laboratory models and individuals with DED underscores redox
imbalance as one of the many pathophysiological factors driving the disease. 641
Sjögren disease may also be characterised by reduced or absent levels of complement
regulators (CD59, CD55, and CD46), alongside upregulation of C3. This imbalance may
drive excessive complement activation in ocular tissues and may suggest a possible
therapeutic pathway. 642

Ongoing research continues to shed light on the role of tear cytokines in the context of
DED. A recent meta-analysis of tear film cytokines in Sjögren disease revealed elevated
levels of inflammatory cytokines, including IFN-γ, TNF-α, IL-1α, IL-1Ra, IL-4, IL-6, IL-8,
IL-10, IL-17, IL-21, and IL-22. Interestingly, IL-23 levels were significantly lower
compared to healthy controls. 643 Additionally, reduced levels of epidermal fatty acid-
59
binding protein (E-FABP), which regulates inflammatory pathways on the ocular surface,
further suggest its potential as a biomarker for epithelial damage and altered lipid
metabolism in the disease. 644 However, despite these advances, a persistent challenge
in the field is the lack of standardization in sample collection and analytical techniques,
645
which hinders the translation of these findings into clinical practice.
Tear proteomic profiling has also revealed altered biological pathways related to corneal
sensitivity and nerve parameters in DED and neuropathic corneal pain. 646,647 (See
section 8.5)

One emerging area of interest in this field involves examining the complex interplay
between the tear proteome and the ocular surface microbiome, and its potential
contribution to the onset of dry eye. Recent data indicate that the interaction between
these two systems is beneficial to combat pathogens and maintain ocular health
through modulation of the inflammatory response. 648 Moreover, the application of
whole-metagenome sequencing has enabled researchers to pinpoint unique microbial
compositions in individuals with DED, with the relative abundances of certain bacteria
being correlated with specific tear proteins in the tear fluid. 649 Understanding the
crosstalk between these two components of the tear fluid may provide opportunities for
the development of personalized therapeutics tailored to individual patient profiles.

8.5 Lipid-protein-mucin interactions

Interactions between the various components of the tear film are important for
maintaining its stability and protecting the ocular surface and the expression of these
components is interconnected. For instance, there is a compensatory mechanism where
the absence of lipid secretion, caused by stearoyl-CoA desaturase-1 deficiency, leads
to increased tear volume and enhanced expression of mucins, along with changes in
the expression of lipid metabolism genes. 632 The knowledge gained from understanding
the interactions between tear film components is also being used to develop dry eye
treatment strategies. Two treatment studies suggest that mucomimetic polymers can
improve the structure and functionality of the lipid layer. 650,651 Cross-linked hyaluronic
acid is particularly effective in promoting tear ferning and the spreading of meibum in
vitro, suggesting therapeutic potential of these polymers for ocular surface health. 652

8.6 Mucins

The realm of ocular mucins has witnessed several scientific discoveries, shedding
additional light on their important role in maintaining ocular surface health. Membrane-
associated mucins act as regulators of transcellular barrier function, tear film stability
and apical epithelial cell architecture. 493,653 Membrane-associated mucins are distinctly
distributed along the conjunctiva of those with and without DED, emphasizing the vital

60
role of these mucins in minimizing eyelid friction during blinking. 654 Advanced modelling
approaches have enabled researchers to establish a relationship between the loss of
membrane-associated mucins and the premature rupture of the tear film, predicting
quantitatively the shortening of NIBUT observed in DED. 655

Through comprehensive genomics analyses, researchers have identified mucin variants

lacking sialylation due to a point mutation in a sialyltransferase gene, St6galnac1, which
plays an important role in synthesizing sialyl-Tn and serves to protect the conjunctival
mucosa against foreign particles. 656 Other investigations have highlighted the rapid
turnover of gel-forming mucins in the healthy ocular surface 657 and the potential
benefits of thymosin β4, resolvin D2 and microRNA inhibition in their ability to stimulate
the secretion of gel-forming mucins. 658-660 A randomized clinical trial comparing
thymosin β4 to placebo in a 28 day study showed no difference to placebo in corneal
fluorescein staining and symptoms. 661

8.7 MicroRNAs (miRNAs)

The altered expression of miRNAs in tear fluid has positioned them as promising
candidates for non-invasive biomarkers in DED. To date, approximately 300 distinct
miRNAs have been identified in tear fluid, many of which are recently discovered and
are poorly understood in terms of their regulatory functions. 662 RNA sequencing
experiments has revealed that extracellular vesicles in the tear film in both non-Sjögren
DED and healthy controls carry distinct miRNA profiles, with 126 differentially expressed
miRNAs between the groups. 663 Among these, nine miRNAs (miR-127-5p, miR-1273h-
3p, miR-1288-5p, miR-130b-5p, miR-139-3p, miR-1910-5p, miR-203b-5p, miR-22-5p,
and miR-4632-3p) were significantly upregulated in DED and were associated with
inflammation, indicating a potential role in disease pathogenesis. Other studies, on the
other hand, have shown an inverse relationship between miRNAs in tears and
inflammation. For instance, miR-223 inhibits hyperosmolarity-induced inflammation
through downregulating NLRP3 activation in human corneal epithelial cells and those
with DED. 412 Comparison of the expression of 43 miRNAs in the tears of those with
Sjögren disease and healthy controls have revealed 14 significantly differentially
expressed miRNAs that may be involved in the pathogenesis of Sjögren disease,
though none were correlated with ocular staining scores. 664 As the precise molecular
mechanisms by which these miRNAs contribute to DED remain largely unclear, and the
lack of standardized methods hampers comparison across studies, further research is
essential to clarify the specific roles of miRNAs within the ocular environment.

8.8 Translational dry eye models of tear film

8.8.1 In vitro models

61
Translational models of the tear film aim to bridge the gap between basic research and
clinical applications. Significant advances in recreating the ocular surface in vitro have
emerged using organs-on-chips and stem cell-derived organoids. Organ-on-chip
technology employs microfluidic devices to replicate the ocular surface environment,
allowing for detailed study of tear film components such as mucins and inflammatory
cytokines. By utilizing microfluidic platforms with segmented channels, researchers can
simulate dry eye conditions by exposing cultured human corneal epithelial cells to an
air-liquid interface within a chip. 665 Additionally, induced pluripotent stem cells have
facilitated the creation of organoids, offering a new platform to study the molecular
mechanisms involved in DED. These approaches have allowed the generation of
functional conjunctival epithelial lineage cells, including goblet cells. 445,666,667

Biomimetic models that rely on the enzymatic removal of mucins in cell culture have
emerged to more accurately recapitulate the pathological changes in lubrication,
adhesion, and barrier function often observed in mucin-deficient DED. 668 In addition,
hydrophilic and hydrophobic glass surfaces have been used to model the interactions of
the tear film with a healthy cornea or a hydrophobic cornea in the absence of a
glycocalyx. 669 Also noteworthy is the development of miniaturized analogs of a blinking
human eye, which utilize a dome-shaped 3D cell culture scaffold to mimic the dynamic
interface between the ocular surface and the external environment. 445 Other
investigators have used molecular modeling of the tear film and machine learning
models to better understand the contribution of proteins to tear film stability and
proteomic changes in MGD, respectively. 670,671

8.8.2 In vivo models

Compared to in vitro models, there have been relatively fewer innovations in the
development of animal models of DED in recent years. The established models rely on
surgical procedures, such as removal of the exocrine glands, exposure of the eyes to
drugs like benzalkonium chloride, and placement in a dry environment, with or without
scopolamine (See Section 5, TFOS DEWS II Tear Film Report). 561 Additionally, there
are numerous genetically modified models targeting genes involved in tear production,
inflammation, or autoimmune responses. These models continue to generate important
results in the development and approval of therapeutic agents for DED. 672 Almost all
current models mimic a severe aqueous-deficient, inflammatory form of DED (see
Section 7) which is not reflective of EDE, and there is a need to develop alternative
models for designing and testing therapies that better address different subtypes of
DED. More recently, there has been a renewed emphasis on the use of canine models
of evaporative and aqueous-deficient DED to better align with human disease. 673

8.9 Summary and future directions

62
This review has identified several areas for future research that may help in the
understanding of the pathogenesis and subclassification of DED, as well as identifying
better biomarkers to help clinicians classify and monitor DED. Researchers should use
lipids that more closely align with those in meibum, rather than mimics of meibum lipids
for future in vitro experiments. Further research is needed to explore the relationship
between disordered lipids that result in spreading and increased elasticity, compared to
ordered lipids that lead to improved resistance to evaporation. Additionally,
incorporating more accurate models of the muco-aqueous layer in laboratory
experiments may help elucidate the roles of lipids, mucins, proteins, and other
components in stability and evaporation. There is also a need for a more detailed
understanding of whether tear biomarkers can be used to differentiate subtypes of DED
as described in the TFOS DEWS III Diagnostic Methodology report. 1 Analyses of
microbiome changes across individuals of different ethnicities and countries of
residence may provide further insights into its potential role in DED pathogenesis or as
a marker for the disease. Understanding the potential role of different microRNAs in
DED pathogenesis, DED subtype or as biomarkers could be a highly promising area for
future investigation.

9. Pain and Sensation

9.1 Introduction

As described in the TFOS DEWS II Pain and Sensation Report, 40 the ocular surface,
particularly the cornea, is densely innervated with sensory fibers that have important
functions in the maintenance of ocular surface health. The dynamic nature of ocular
surface nerves is often underappreciated, as this system continuously undergoes
remodeling in adults, particularly nerve terminals in the corneal epithelium. Many
common insults can amplify this remodeling, including accidental or surgical injury or
disease processes that lead to chronic inflammation (e.g., Sjögren disease, Steven-
Johnson disease, herpes keratitis). Damage to ocular sensory nerves can result in
corneal nerve loss, a change in nerve architecture, and altered sensitivity to stimulation.
Although nerve regeneration can occur after such damage, it is typically gradual and
incomplete. As a result, it often fails to fully restore the original density, architecture, and
function of the corneal innervation, leading to persistent changes in neural excitability.
This update will consider new evidence in corneal nerve remodeling during normal
physiology and following trauma, surgery or inflammation and recent evidence for the
anatomical and functional status of the corneal nerves in diagnosing and managing
DED.

9.2 Corneal nerve remodeling in adults

63
The development of corneal nerves begins in the fifth gestational month with the
formation of sensory axons around the cornea, followed by their radial extension into
the corneal tissue. Although the specific molecular signals controlling corneal nerve
growth are still unknown, molecules such as nerve growth factor (NGF) and various
neurotrophic factors released by corneal cells contribute to the development and
survival of corneal nerves. The growth of the corneal nerves does not stop once
development is complete but continues to occur continuously. In adults, corneal
subbasal nerves and their terminals undergo continuous morphological rearrangements
throughout life while the stromal nerves present few morphological changes. 674-676

Observations from living human eyes using in vivo confocal microscopy reveal that
subbasal nerves move centripetally at rates of 10–20 μm per day. These nerves
elongate by adding new material near the site of nerve penetration to the epithelium
from the Bowman's layer. Distal nerve segments eventually degenerate or slough into
the tear film due to the turnover of the corneal epithelium. 40,677 Additionally,
intraepithelial nerve terminals undergo spontaneous morphological changes through
long-term reconfigurations and short-term reorganization in response to outward
migrations of differentiating epithelial cells. Notably, corneal nerve remodeling is more
prominent in the central regions of the cornea than in the periphery. Research in living
transgenic and knock-in mice reveals that, over time, there are noticeable changes in
the subbasal nerve fibers and intraepithelial nerve terminals. The presence of
continuous remodeling is supported by the expression of growth-associated protein
43sub in the epithelial nerves of intact corneas. 675,677

Continuous remodeling is insufficient to maintain lifelong corneal innervation. As

mammals age, the density of corneal nerve terminals decreases, leading to reduced
corneal sensitivity and changes in tearing regulation. In older individuals, there is a
noticeable reduction in subbasal and intraepithelial nerve density and increased
tortuosity and disorientation of subbasal nerves. These changes affect all corneal
sensory nerves, whether nociceptive or cold thermosensitive, and result in an
abundance of simple nerve terminals and a scarcity of complex nerve terminals in aging
individuals. The reduced density and dysfunction of corneal nerves, especially cold
thermoreceptors, seems to be responsible for the altered tearing and sensitivity in older
population. 678

9.3 Corneal nerve regeneration after surgery

Corneal nerves may be severed during corneal and anterior segment surgery, such as
photorefractive keratectomy (PRK), laser-assisted in situ keratomileusis (LASIK),
cataract surgery, iridectomy, trabeculectomy, and corneal transplantation. The survival
of corneal nerves relies on transporting essential substances from their parent nerve

64
cells in the trigeminal ganglion. Therefore, surgical procedures that disrupt corneal
nerve fibers can lead to rapid degeneration of the distal axons, reduced corneal
sensitivity, and impaired functional integrity of the ocular surface.

Although corneal nerves can regenerate, this process is slow and imperfect. After most
corneal surgeries, the regeneration of nerves is characterized by decreased nerve
density, changes in nerve structure, and diminished corneal sensitivity. Regeneration is
more delayed and incomplete when nerves are cut closer to their origin. 679
Consequently, surgical disruption of the subbasal and subepithelial nerve plexuses
typically results in less severe and short-term damage to corneal innervation than deep
or penetrating incisions affecting major stromal nerve bundles.

Although corneal sensitivity typically returns to preoperative levels after LASIK within 6-
12 months, 680 a significant proportion of patients may experience long-term dry eye
symptoms due to impaired nerve regeneration. 681,682 Nerve damage in small incision
lenticule extraction surgery is less marked than in LASIK, resulting in a faster nerve
regeneration three months post-surgery. However, no significant difference is observed
at six months. 257 A meta-analysis of corneal sensitivity recovery has shown a more
rapid early recovery of corneal sensitivity with small incision lenticule surgery compared
with femtosecond LASIK. 683(See also TFOS Lifestyle report 684)

In PRK, where the corneal epithelium is removed, and the corneal stroma is reshaped
with an excimer laser without creating a flap, nerve regeneration and recovery of
corneal sensitivity occur more rapidly than in LASIK. 685 Despite this, subbasal nerve
density, architecture, and corneal sensitivity may remain reduced for up to 1-2 years
post-PRK. 686

In cataract surgery, small, perilimbal incisions have minimized the risk of significant
injury to corneal innervation. However, in corneal transplantation procedures such as
penetrating keratoplasty, a full-thickness incision cuts all corneal nerves, resulting in
complete denervation of the transplanted cornea. Nerve regeneration following
penetrating keratoplasty is slow, and even years later, the innervation density of the
transplanted tissue remains lower than that of the host peripheral cornea. 687 Stromal
nerves regenerate poorly, which may be attributed to the misalignment of Schwann cell
channels in the donor cornea with the stromal nerve stumps in the host cornea. This
contrasts with the perilimbal incisions used in cataract surgery, where stromal nerves on
opposing sides of the incision remain closely aligned. Following penetrating
keratoplasty, limited nerve regeneration occurs. A few subbasal nerve fibers elongate
through the epithelium at the graft margin to enter the donor basal epithelium.
Regenerated subbasal nerves may exhibit atypical orientations and morphologies. 687

65
The corneal nerve density and corneal sensitivity remain significantly reduced
compared to healthy corneas even decades after surgery. 688

Although theoretically expected not to alter corneal innervation, other corneal transplant
techniques, such as Descemet membrane endothelial keratoplasty, show a temporary
decrease in nerve density early after transplantation. However, complete recovery of
corneal nerve density and function to preoperative values typically occurs within 6-10
months post-surgery. 689 Studies have also reported similar results with Descemet’s
stripping automated endothelial keratoplasty, where the corneal sensations were noted
to be normal within 6 months following surgery. 690 Another study comparing corneal
sensation between Descemet membrane endothelial keratoplasty and penetrating
keratoplasty showed that corneal sensation improved significantly following Descemet
membrane endothelial keratoplasty but was slightly but not significantly decreased after
penetrating keratoplasty. 691 These findings suggest better preserved corneal
sensations following endothelial transplants compared to PK.

9.4 Nerve regeneration in pathological conditions

The peripheral nervous system has impressive regenerative capabilities following injury.
However, injuries to the afferent axons of trigeminal neurons can lead to significant
morphological and functional changes, which depend on the magnitude and location of
the damage. 679,692 The regeneration of neurons after injury is influenced by a supportive
environment for axon growth and the involvement of non-neuronal cells like Schwann
cells. 693

As described in the TFOS DEWS II Pain and Sensation Report, 40 mechanical trauma or
inflammatory damage to the peripheral axons of corneal trigeminal neurons causes a
complex cellular response and changes to their spontaneous and stimulus evoked firing
rates. The expression, distribution and activation thresholds of the transduction ion
channels changes. These disturbances lead to increased responsiveness to normal
stimuli (allodynia), spontaneous firing without intended stimulation and increased
abnormal or unpleasant sensations from a stimulus that would normally elicit a
response (hyperalgesia).

The mechanisms that stimulate and direct neurite outgrowth from injured and intact
areas of corneal innervation following local nerve injury still need to be fully understood.
Corneal epithelial cells release several growth factors following an injury, which may
play essential roles. Nerve growth factor is upregulated after corneal epithelial wounding
and topical recombinant human nerve growth factor has been used to stimulate corneal
nerve regeneration and recovery of corneal sensitivity 694 in the treatment of
neurotrophic keratitis, where sensitivity is reduced. 695 The role of nerve growth factor in

66
the mechanisms resulting in allodynia or hyperalgesia, however, has not been
established, although restoration of nerve function may be advantageous.

Ocular and systemic diseases, including herpes virus keratitis, diabetes, and ADDE,
can negatively impact corneal nerves. 696 Diabetes significantly alters corneal nerve
morphology and function, reducing nerve density and sensitivity. In diabetic patients, the
appearance of subbasal nerves resemble intraepidermal small fiber neuropathy, making
in vivo confocal microscopy of corneal nerves a valuable biomarker for monitoring
diabetic neuropathy. 697

In summary, in healthy adult corneas, the subbasal nerve fibers and intraepithelial nerve
endings undergo continuous remodelling and regenerate rapidly following damage,
whereas stromal nerves maintain their structure over time. The dynamic nature of
intraepithelial nerve endings accounts for their rapid regeneration after injury, while
subbasal nerve fibers regenerate more slowly, and stromal nerve trunks may not fully
regenerate. Corneal nerve morphology is affected by trauma, ocular surgery, infections,
chronic tear deficiency, and various systemic diseases. Although damaged corneal
innervation can regenerate like other peripheral nerves, their morphology and function
are often incompletely restored, leading to reduced sensitivity, abnormal sensations,
and pain.

9.5 Nerve abnormalities and DED

Many manifestations placed under the heading of “dry eye” occur because of
morphological and functional changes in ocular innervation. Changes in ocular sensory
nerves induce symptoms such as unpleasant sensations of different intensities, ranging
from dryness or ocular discomfort to lacerating and burning pain. 698 Conversely,
abnormal nerve functioning can lead to alterations in tissue trophism and in the
regulation of tear production and blinking, which in turn contributes to both symptoms
and signs.699 In other words, sensory innervation can be altered by chronic eye dryness
and can also contribute to DED pathogenesis.

Reduction of the ocular surface moistness either by reduced tearing or increased tear
evaporation is a stressful situation for the corneal epithelium exposed to a hyperosmotic
tear film and the environment (See section 7). Tear hyperosmolarity can independently
affect subbasal corneal nerves in an animal model. 700 As a response to this
hyperosmolar and desiccating stress, corneal epithelial cells and immune resident cells
produce local inflammatory mediators, primarily IL-1 and TNF- α, which stimulate the
production of matrix metalloproteases, activate dendritic cells and local inflammation
occurs, 458 leading to sensitization of nociceptive nerve terminals and development of
discomfort and pain sensations. When dryness becomes chronic, it also leads to
corneal nerve damage and, consequently, the morpho-functional changes of the
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sensory nerves during chronic eye dryness resemble both those produced during
inflammation and those produced by nerve injury. The eyelid movement causes
mechanical friction at the ocular surface when the tear film is thin and does not lubricate
well. Together with the chronic inflammation, this movement damages the epithelium
and the intraepithelial nerve terminals, and eventually the subbasal nerve fibers,
triggering the mechanisms of nerve degeneration and regeneration. 701

In clinical practice, these degeneration and regeneration processes are evidenced by

signs such as the reduction in the density and branching of the subbasal plexus nerve
fibers, and the increase in their tortuosity when explored by in vivo confocal microscopy,
as well as by a reduction in corneal sensitivity to stimulation (shown by increased
sensation thresholds) that often occurs simultaneously with hyperalgesia and
spontaneous pain sensations and is aggravated by tear film instability. 702 A recent in
vivo confocal microscopy of corneal nerves confirmed these nerve and branch density,
and length reductions in 23 participants with DED and showed that osmolarity exhibited
a weak negative correlation to these nerve parameters, whereas other ocular surface
signs were not associated with any nerve parameters. 703 Despite the nerve changes
reported, it does appear however that not all sensory modalities are equally affected
and corneal hypersensitivity to cold stimuli has been reported in DED, 704 in computer
vision syndrome 705 and contact lens discomfort. 706 Repeated evaporative cooling at
the ocular surface may change the excitability of corneal receptors and their subsequent
responsiveness. 40,702,707 Tear film instability can cause changes in suprathreshold
scaling by both cold thermoreceptors and polymodal nociceptors. 708

Most of the alterations of ocular surface sensitivity in chronic eye dryness are due to
functional changes in corneal nerves that resemble those seen in injured nerves. There
is an increased excitability of corneal nerves consecutive to the increased activity and
expression of sodium channels,709-711 resulting in an increased spontaneous firing of
cold thermoreceptors that not only leads to dryness sensations but also to dysregulation
of protective mechanisms driven by thermal sensory input such as tearing and blinking,
contributing to increase and perpetuate the ocular surface disturbances.

In contrast to the pathophysiology of neuropathic pain, defined by pain or altered

sensation due to nerve damage, nociplastic pain occurs in the context of no discernible
tissue or nerve damage. Instead, nociplastic pain is believed to be related to
dysfunctional central sensory processing of pain through a process called sensory
sensitization. Either neuropathic or nociplastic pain can cause chronic ocular pain and
and are particularly associated with a subset of DED patients who report pain of a
magnitude that is out of proportion to ocular surface findings (Table X).c712 Quantitative
sensory testing and functional magnetic resonance imaging studies remain the primary

68
methods of assessing nociplastic pain, but the underlying pathophysiology remains
poorly understood.

Table 4. Mechanistic characterization of pain, reproduced from De Lott et al., 2025.

712

Type Neurobiology Clinical signs & Treatment Non-ocular and

symptoms ocular-related
examples

Nociceptive Actual or Localized to the site  Remove source of Non-ocular: burn

threatened of injury and nociception
tissue damage surrounding area  Promote tissue healing Ocular: corneal
via activation  Topical medications abrasion
of nociceptors  Short term systemic
medications: NSAIDs,
acetaminophen, anti-
inflammatories, opioids

Neuropathic Damaged Localized to a  Peripheral neuropathic: Non-ocular: diabetic

somatosensory dermatome nerve blocks, topical neuropathy
system, either (peripheral) or site medications (e.g.,
peripheral or of injury (e.g., CNS capsaicin) Ocular: Post-
central demyelinating herpetic neuralgia
 Systemic medications:
lesion in the spinal
gabapentinoids, tricyclic
cord causing leg
antidepressants, SNRIs,
paresthesias and
sodium channel blockers
pain)

Nociplastic Altered pain Regional and  Lifestyle: education, Non-ocular:

perception and diffuse sleep, exercise fibromyalgia
processing in pain Multisensory
the central sensitivity COPCs
 Behavioral/Integrative: Ocular: referred
self management, pain from chronic
nervous
psychotherapy, etc. tension-type
system
 Medications: SNRIs headache

9.5.1 Impact of DED on nerve structure and function in animal models

Animal models have been developed to replicate human correlates of DED. While initial
focus examined immune abnormalities, recent studies have found that various insults
that create tear and ocular surface abnormalities also impact upon corneal nerves.

Several mouse models replicate DED in Sjögren disease, including an IL-2 receptor -
chain knockout model. 713 In one study, intraepithelial corneal nerve density of knockout
was compared to wildtype mice using confocal microscopy at 4 weeks, 6 weeks, 8
weeks, and 10 to 11 weeks after birth. After combining data from each time point,
overall density was significantly decreased in the knockout compared to wildtype. These

69
results suggest that corneal nerve alterations coincide with the onset of DED, with
reduced nerve fiber density commonly noted.

Nerve alterations have also been examined in a GVHD murine model, 714 using an
allogeneic bone marrow transplant (case) group (strain B10.D2 to strain BALB/c) and a
syngeneic bone marrow transplant group (strain BALB/c to strain BALB/c, control group)
at 1-, 2-, 3-, and 4-weeks post-transplant. Corneal nerve tortuosity and branching were
increased in the allogeneic bone marrow transplant group at 4-weeks post-transplant
compared to 1-week post-transplant. Conversely, no significant differences were
observed in branching or tortuosity in the syngeneic group at any time point. These
findings support the induction of morphologic changes in corneal nerves associated with
the progression of GVHD-associated DED.

Other protocols have been developed to model DED following ocular surgery, such as
photorefractive keratectomy (PRK). 715 PRK (-9D) was performed monocularly in New
Zealand rabbits while the contralateral cornea was used as a control. Corneal nerve
fiber density was evaluated in post-PRK corneas with acetylcholinesterase
histochemistry staining at 1 day (n = 3), 1 month (n = 3), 2 months (n = 3), 3 months (n
= 3), and 6 months (n = 4) after surgery. Compared to controls (n = 3), nerve density
was significantly decreased in post-PRK corneas after 1 day, 1 month, and 2 months,
but was not different at 3- and 6-months post-surgery. Nerve morphology alterations
persisted however, (increased tortuosity and aberrant innervation) until the final time
point at 6 months. Findings that surgery may result in corneal nerve alterations that do
not fully return to baseline by 6 months post-surgery, consistent with the effects of
refractive surgery in human. 680

Other studies focused on DED prompted by iatrogenic challenges. One murine study
used confocal microscopy with staining using an anti-β3 tubulin antibody to examine
corneal nerve fiber density after repeated topical administration of benzalkonium
chloride for 14 days. 716 There was a significant reduction in density in benzalkonium
chloride-treated mice compared to controls.

Partial or complete lacrimal gland excision has also been used to model DED. 436 In one
study, the number of corneal nerve terminals in the surgical group was assessed
qualitatively by confocal microscopy and compared to a sham-surgery and control group.
The surgical group demonstrated fewer corneal nerve terminals compared to both
sham-surgery and control groups. This study also examined nerve function with
electrophysiological studies and found that post-surgical animals developed time-
dependent corneal mechanical hypersensitivity accompanied by increased spontaneous
ciliary nerve fiber electrical activity. Other investigators have found similar functional
changes after gland excision, with increased activity and expression of sodium channels

70
that lead to increased excitability of corneal nerves. This results in a greater
spontaneous firing of corneal cold thermoreceptors, causing sensations of dryness, and
an enhanced excitability of corneal nociceptors that results in low frequency
spontaneous firing and increased response to stimulation, resulting in sustained
discomfort and hyperalgesia. 710,717,718 These findings suggest that, beyond nerve
density and morphology, corneal nerve function can be impacted differentially in DED
subtypes.

Desiccating stress is often induced using a combination of scopolamine and low

humidity conditions (e.g., humidity: 25%, constant airflow). In one study, dessication
reduced corneal sensitivity over time, 719 linking dry eye onset with corneal functional
abnormalities. In summary, animal models can be used to study ocular surface nerve
anatomy and sensitivity (and their upstream connection) as they relate to various insults
that lead to chronic dry eye. Changes in morphology and sensitivity have been noted
across many models, both reduced abilities to detect stimuli (further aggravating ocular
surface disturbances) and the development of spontaneous discomfort and pain, that
primarily stem from functional alterations in corneal nerves, resembling those observed
in peripheral nerves after injury. These data highlight the intricate relationship between
corneal nerves and dry eye.

9.5.2 Structural and functional nerve alterations in DED in human

In DED, alteration of corneal nerve structure and function is frequently implicated in the
onset, progression, and severity of symptoms. Disruption of the tear film in DED can
result in nerve fiber damage which stimulates regenerative processes that are often
improper or incomplete. The TFOS DEWS II Pain and Sensation Report in 2017 720
determined that while IVCM is a reliable method for detecting abnormal corneal nerve
morphology, nerve density was a somewhat unreliable marker for nerve fiber damage,
but that other morphological parameters were more useful including increased tortuosity,
reflectivity and increased beading. More recently a higher frequency of microneuromas
has been documented. 702 These structural aberrations, in turn, may further impair tear
film production and the blinking reflex resulting in additional nerve damage and
conceivably neuropathic dysfunction. 702 Typical manifestations of neuropathic
dysfunction include ocular pain, loss of sensation, and tear hyperosmolarity. Damage to
nerve fibers in DED can also result in physiologic disruption of the corneal nerve’s usual
neurotrophic functions, including regulation of nerve growth factor and substance P, that
help mediate epithelial growth, local immunoregulation, and nerve fiber regeneration.
702,721
Together, these structural and functional changes likely play a significant role in
the pathophysiology of DED.

71
Interestingly, some individuals with DED report symptoms out of proportion to ocular
surface alterations and others are found to have signs that are more severe than their
symptoms suggest. 158 One possible explanation for the symptom>sign discordance is
the presence of central abnormalities, with inappropriate processing of somatosensory
signals due to changes in the central nervous system referred to as central sensitization.
Central sensitization is associated with generalized somatosensory dysfunction and
cutaneous hypersensitivity in addition to excessive ocular pain.

9.5.2.1 Corneal nerve anatomy in DED

Different manifestations of corneal nerve fiber abnormalities likely have clinical

significance for patients with various subtypes of DED but are not yet fully understood.
In recent years, several studies have improved the understanding of IVCM in DED.
Table 5 summarises the key findings from studies exploring corneal nerve tortuosity.
Nerve tortuosity appears to higher in Sjögren DED compared with non-Sjögren DED 722
723
based on the Oliveira-Soto and Efron (OSE) grading scale, 724. However, greater
tortuosity was seen in corneal neuropathic pain associated with autoimmune disease
compared with corneal neuropathic disease in non-autoimmune DED, MGD or MGD
without pain and controls. 725 726

Table 5. Corneal nerve tortuosity in DED subtypes

Author(s)/year Location Sample Size Methodology Findings

727
Japan GVHD (n = 12) Imaging: multi- ↑ tortuosity in GVHD
image (3), central compared to
Controls (n = 10) cornea + inferior controls.
limbal epithelia

Analysis:
ImageJ/NeuronJ,
OSE grading
scale
722
United States SDED (n = 22) Imaging: multi- ↑ tortuosity in SDED
image (3), central and NSDED
NSDED (n = 12) cornea compared to
controls.
Controls (n = 5)

Analysis:
ImageJ/NeuronJ,
OSE grading
scale
725
France AIDED-NCP (n = 7) Imaging: single ↑ tortuosity in

72
 image, central AIDED-NCP
MGD-NCP (n = 11) cornea compared to
controls. Tortuosity
AIDED (n = 8) not different in MGD-
NCP, painless
MGD (n = 8) Analysis: AIDED, or painless
ImageJ/NeuronJ, MGD compared to
Controls (n = 10) study-designed controls.
tortuosity scale
723
China SDED (n = 22) Imaging: multi- ↑ tortuosity in SDED
image (5), central compared to
NSDED (n = 20) cornea NSDED.

Analysis:
ImageJ/NeuronJ,
OSE grading
726
France SDED (n = 71) Imaging: multi- ↑ tortuosity in SDED
image (5), central compared to MGD
MGD (n = 20) cornea and controls.

Control (n = 20)

Analysis:
ImageJ/NeuronJ,
OSE grading

GVHD=graft-versus-host disease; DED=Dry eye disease; SDED=Sjögren DED; NSDED= non-Sjögren

DED; AIDED= autoimmune DED; MGD= meibomian gland dysfunction; NCP= neuropathic corneal pain.

Corneal nerve fiber density (CNFD) has also been studied in various DED subtypes
(Table 6) and in most studies appears to be significantly decreased in Sjögren DED,
GVHD, non-Sjögren DED and neuropathic corneal pain compared to controls, 722,728
729,730 731
although within studies density is similar between DED types. 732 One small
study has evaluated MGD and found no reduction compared with controls, 725 however
a larger study found a reduction in both ADDE and EDE compared with controls. 733
The reduction in density is greater with increased symptoms. Those with normal-to-mild
symptoms have similar CNFD to controls. 734 These studies reinforce that CNFD values
are generally lower in some DED subtypes, most notably in auto-immune associated
DED and that CNFD may be affected by degree of symptoms severity/ocular pain.

Table 6. Corneal nerve fiber density in DED subtypes

Author/year Location Sample Size Methodology Findings

734
India Normal-mild EDE (n Imaging: single ↓ CNFD in EDE with
= 29) image, central moderate-to-severe
cornea symptoms compared

73
 to controls. CNFD not
Moderate-severe different in EDE with
EDE (n = 23) normal-to-mild
Analysis: symptoms compared
Control (n=43) ACCMetrics to controls.

727
Japan GVHD (n = 12) Imaging: multi- CNFD not different in
image (3), GVHD compared to
Control (n = 10) central cornea + controls.
inferior limbal
epithelia

Analysis:
ImageJ/NeuronJ
722
United SDED (n = 22) Imaging: multi- ↓ CNFD in SDED and
States image (3), NSDED compared to
NSDED (n = 12) central cornea controls.
Control (n = 5)

Analysis:
ImageJ/NeuronJ
730
Japan NSDED (n = 25) Imaging: multi- ↓ CNFD in NSDED
image (5), compared to controls.
Control (n = 25) central cornea

Analysis:
ImageJ/NeuronJ
735
France DED (n = 32) Imaging: multi- ↓ CNFD in DED
image (5), compared to controls.
Control (n = 15) central cornea

Analysis:
ImageJ/NeuronJ
732
Italy SDED (n = 20) Imaging: multi- ↓ CNFD in SDED and
image (3), GVHD compared to
GVHD (n = 19) central cornea controls.
Control (n = 30)

74
 Analysis:
ACCMetrics
736
Turkey GVHD (n = 22) Imaging: multi- ↓ CNFD decreased in
image (3), GVHD compared to
Control (n = 28) central cornea controls.

Analysis:
ImageJ/NeuronJ
729
United NCP (n = 25) Imaging: multi- ↓ CNFD in the NCP
States image (3), and DED compared to
DED (n = 30) central cornea controls.
Control (n = 16)

Analysis:
ImageJ/NeuronJ
731
United ADDE (n = 24) Imaging: multi- ↓ CNFD in ADDE and
States image (3), EDE groups to
EDE (n = 46) central cornea controls.
Control (n = 45)

Analysis:
ImageJ/NeuronJ
725
France AIDED-NCP (n = 7) Imaging: single ↓ CNFD in AIDED and
image, central MGD-NCP compared
MGD-NCP (n = 11) cornea to controls. CNFD not
different in AIDED-
AIDED (n = 8) NCP or MGD
MGD (n = 8) compared to controls.
Analysis:
ImageJ/NeuronJ
Control (n = 10)
723
China SDED (n = 22) Imaging: multi- ↓ CNFD decreased in
image (5), SDED compared to
Control (n = 20) central cornea NSDED.

Analysis:
ImageJ/NeuronJ,
737
India ADDE/EDE + Imaging: single ↓ CNFD in those with
symptoms (n = 10) image, central DE symptoms
cornea compared to controls.
Control (n = 15)

75
 ADDE +/- Analysis: CNFD not different in
symptoms (n = 57) ACCMetrics ADDE (+/- symptoms)
or EDE (+/-
EDE +/- symptoms symptoms) compared
(n = 16) to controls.
Control (n = 15)
738
China DED (n = 155) Imaging: multi- ↓ CNFD in DED
image (≥10), compared to controls.
Control (n = 20) central

Analysis: CS-Net
726
France SDED (n = 71) Imaging: multi- ↓ CNFD in SDED
image (5), compared to controls.
MGD (n = 20) central cornea CNFD not different in
SDED compared to
Control (n = 20) MGD.
Analysis:
ImageJ/NeuronJ
739
China DED (n = 25) Imaging: single ↓ CNFD in DED
image, central compared to controls.
Control (n = 20) cornea

Analysis:
ImageJ/NeuronJ,
ACCMetrics
733
Turkey ADDE (n = 22) + Imaging: multi- ↓ CNFD in ADDE and
EDE (n = 21) image (3), EDE compared to
central cornea controls.
Control (n = 20)

Analysis:
ImageJ/NeuronJ

DED= dry eye disease; EDE= evaporative dry eye; GVHD= graft-versus-host disease; SDED= Sjögren
dry eye disease; NSDED= non-Sjögren dry eye disease; NCP= neuropathic corneal pain; ADDE=
aqueous-deficient dry eye; CNFD= corneal nerve fiber density; MGD = meibomian gland dysfunction.

Newer nerve parameters, such as microneuromas, defined as irregular expansions of

subbasal nerve endings that suggest nerve damage underlying ocular symptoms, have

76
been included in some studies (Table 7). Conceivably, either due to the subjective
determination of microneuromas, or the difficulty of discriminating a turn in the nerve
from an ending, there are inconsistencies in the literature. 740,741 Some studies found an
increase in microneuromas among DED subtypes compared to controls, most notably in
autoimmune DED and MGD, 725,742 with a greater increase where there was co-existing
neuropathic corneal pain. 725 While these studies suggest that microneuroma frequency
or number is increased among several DED subtypes compared to controls, other
studies have not replicated these findings in those with symptoms of DED. 743 These
findings point to the need for additional studies that standardize microneuroma
determination and further explore their utility as a biomarker for DED.

Table 7. Corneal nerve microneuromas in DED subtypes.

Author(s)/year Location Sample Size Methodology Findings

729
United NCP (n=25) Imaging: multi- MN frequency ↑ in NCP
States image (3), compared to controls.
DED (n=30) central cornea MN frequency not
different in DED
Control (n = 16) compared to controls.
Analysis:
ImageJ/NeuronJ
725
France AIDED-NCP (n = Imaging: single MN frequency ↑ in
7) image, central AIDE, AIDE-NCP, MGD,
cornea and MGD-NCP
MGD-NCP (n = compared to controls.
11)

AIDED (n = 8) Analysis:
ImageJ/NeuronJ
MGD (n = 8)

Control (n = 10)
743
United DED (n = 119) Imaging: multi- MN frequency not
States image (3), different in DED or DED
DED + refractive central cornea + refractive surgery
surgery (n = 19) compared to controls.
Control (n = 18)
Analysis:
ACCMetrics
742
India ADDE/EDE + Imaging: single MN frequency ↑ in those
symptoms (n = 14) image, central with DE symptoms
cornea compared to controls.
Control (n = 27)

77
 ADDE +/- Analysis: MN frequency not
symptoms (n = 24) ACCMetrics different in ADDE (+/-
symptoms) or EDE (+/-
EDE +/- symptoms symptoms) compared to
(n = 65) controls.
Control (n = 27)
726
France SDED (n = 71) Imaging: multi- MN frequency ↑ in
image (5), SDED compared to
MGD (n = 20) central cornea controls but not
compared to MGD.726
Control (n = 20)

Analysis:
ImageJ/NeuronJ

DED= dry eye disease; NCP= neuropathic corneal pain; AIDED= autoimmune dry eye disease; MGD=
meibomian gland dysfunction; ADDE= aqueous-deficient dry eye; EDE= evaporative dry eye; SDED=
Sjögren dry eye disease; MN= microneuroma.

Recent work has explored the use of AI technology in nerve analysis, particularly the
use of deep learning AI models to automatically segment corneal nerves.744. Using the
TFOS DEWS II diagnostic criteria, 131 individuals with DED had reduced CNFD
compared with controls and values were similar to that of manual annotation.738
Applying new technologies to IVCM will allow for more consistent and faster
quantification of images that can applied across centers, facilitating comparisons across
different populations.

9.5.2.2 Corneal sensitivity in DED

Corneal sensitivity testing is an important diagnostic tool that can provide valuable
insights into underlying somatosensory abnormalities in DED. The TFOS DEWS II
report highlights that corneal sensitivity to mechanical stimuli tends to be reduced in
patients with ADDE.720 However, studies using air/gas esthesiometers have shown
equivocal findings, with increased 745,746, decreased 747,748, and similar 749 corneal
sensitivity in various DED subtypes. This could be due to differences in stimulus
parameters (e.g., cold, chemical stimulus included in some testing paradigms) or due to
differences in neural responses across various DED subtypes (e.g., ADDE vs MGD).750
Thus, consideration of the type of stimulus, the subtype of DED and patient factors is
critical to interpreting corneal sensitivity results. Table 7 summarizes studies evaluating
sensitivity or pain responses in DED.

78
In a study of corneal mechanical sensitivity in those with DED symptoms tested using
an air jet aesthesiometer, 13% showed hypersensitivity and 11% had hyposensitivity. 749
When grouped by DED subtype, there was an association between ADDE and reduced
sensitivity.726 There is growing awareness that some individuals with symptoms of DED
likely have neuropathic contributors to symptoms and this group, as a whole, display
corneal hypersensitivity. Individuals with hypersensitivity had more severe ocular pain
complaints while those with hyposensitivity had more severe epithelial disruption. 751
These data highlight that differences in corneal sensation and symptom reporting may
align with differences in DED profiles.751 In a study using the Cochet-Bonnet
aesthesiometer, mechanical corneal and pain sensitivity were assessed in individuals
with short tear film breakup DED, defined as TBUT<5secs and Schirmer value >5mm,
and controls (n=46). Pain sensitivity threshold but not mechanical sensitivity threshold
was higher in the DED group. 752 Corneal hyperalgesia was present in 37% of the DED
group and there was a strong relationship between pain sensitivity and subjective pain
scores. 752 Cold sensitivity measured using cooling scores was assessed in a DED and
control group and was compared with mechanical sensitivity threshold using a Cochet-
Bonnet esthesiometer. Treatment with a lubricant containing TRPM8 agonist (0.01%
menthol, a compound that activates cold thermoreceptors) or lubricant alone was
applied bilaterally in a cross-over design. While there was no difference in mechanical
sensitivity, the DED group reported a higher cooling response and score compared to
controls, suggesting mechanical and cold sensitivity may be differentially impacted in
DED. 704 The duration of disease was associated with a greater cooling response,
suggesting that nociceptor activity may change over disease duration. Taken together,
these results suggest abnormal pain and conceivably cold stimulus processing in
different DED subtypes and that different qualities of stimulus processing and other
ocular surface sites, may help to understand neurogenic changes in different subtypes
of DED.

Increased sensitivity (reduced mechanical threshold) has been reported after treatment
of DED or GVHD with autologous serum tears, 753 Sjögren DED with cyclosporine, 754
and short TBUT DED with diquafosol (Table 8).755 This underscores the dynamic nature
of corneal sensitivity, highlighting changes in the short term, with disease, and with
treatment, the implications of which need further study.

The relationship between corneal nerve structure and function is equivocal. Nerve
structure was not consistently associated with gross corneal sensitivity using a cotton
wisp test, where a significant relationship was only noted between corneal sensitivity
and corneal nerve fiber area.756 In a diabetic population, there was a stronger
relationship between anatomy and function. 757 These data highlight that relationships
between corneal nerve anatomy and function may vary with disease type and that
relying on structural findings alone may not suffice in evaluating nerve functional health.

79
Given the importance of understanding nerve function in DED, there have been efforts
to develop new esthesiometers aimed at improving precision, range, stimulus type and
portability, to facilitate routine clinical use. These are reviewed in the TFOS DEWS III
Diagnostic Methodology Report. 1 Briefly these have included non-contact airjet
esthesiometers, 758,759 liquid jet esthesiometers, 705,760,761 and a single-use filament
mechanical esthesiometer (Kerasense, Dompè farmaceutici SPA, Italy). 759,762 While the
initial reports on these esthesiometers are encouraging 705, further research is
necessary to explore their clinical utility, particularly in individuals with DED.

Table 8. Corneal sensitivity in DED

Author/year Location Sample population Methodology Findings

Variation in testing paradigms and DED subpopulations

751
USA DE symptoms (none- Corneal detection Those with
severe) and pain thresholds hypersensitivity had
and relationship more severe ocular
(n=129) with symptoms and pain complaints
signs of DED while those with
(Belmonte hyposensitivity had
esthesiometer) more severe
epithelial disruption
704
USA DED (TBUT5 secs, Cold sensitivity Mechanical
and corneal score by comparing sensitivity was
staining4; n=33) responses to similar between
lubricants with or groups.
Control (n=15) without a cold
receptor stimulator
(TRPM8 agonist:
0.01% menthol) Cold sensitivity
reflected by cooling
(Cochet-Bonnet scores was greater
esthesiometer) in DED than
controls (24.6 vs
12.1, p=0.0005) and
shorter disease
duration (<10 years)
had higher cooling
scores than longer
disease duration
(>10 years)
752
Japan Short TBUT DED Mechanical corneal Detection thresholds
(TBUT<5secs, sensitivity and pain were similar
Schirmer value>5mm; sensitivity (fiber between the groups
(n= 60)) length that elicited
pain) thresholds Short TBUT DED
Control (n=46) (Cochet Bonnet had higher pain
esthesiometer) thresholds,
(26.3±23.1 vs.

80
 6.9±16.4 mm,
p<0.01) which
correlated with
subjective pain
scores (scale 0-4;
r=0.24, p<0.05)
749
USA DED (DEQ ≥6) Corneal detection Mean corneal
thresholds in DED detection threshold
N= 403 (Belmonte of 87 ± 46 mL/min.
esthesiometer) 13% of participants
with hypersensitivity
and 11% with
hyposensitivity

Alterations in corneal sensitivity in response to therapy:

754
France SSDED (primary/ Corneal sensitivity Increased corneal
secondary; n=30) tested after 6 sensitivity over 6
Control (n=15) months of 0.05% months of treatment
cyclosporine use (5.1 to 5.6mm,
(Cochet-Bonnet) p=0.03)
755
Japan Short- TBUT DED Corneal detection DQS treatment
(TBUT <5secs) and pain thresholds significantly lowered
measured after 5 pain sensitivity
DQS group (n=12) weeks of DQS or compared to AT
AT therapy. after 5 weeks of
AT (n=15) (Cochet-Bonnet) therapy.

Neither lowered
detection thresholds
753
USA DED (Ocular GVHD; Corneal sensitivity Mean corneal
n=20) tested after 12 sensitivity increased
weeks of 20% AST at 12 weeks of 20%
use (Cochet- AST in 17 patients
Bonnet) with ocular GVHD
(31.1±23.8 to
51.6±12.6mm,
p=0.001)

DED= dry eye disease; GVHD = graft versus host disease; TBUT = tear film breakup time;
NCP= neuropathic corneal pain; AT = artificial tears; AST = autologous serum tears

In conclusion, sensitivity measurements can examine certain functional qualities of

corneal nerves in DED. There is heterogeneity across and within DED subtypes, with
conditions such as ADDE generally showing reduced corneal sensitivity, and with
symptom severity overall relating to increased sensitivity. Variability in results can also
be attributed to the type of nociceptors stimulated, nature of the stimulus, threshold vs
pain sensitivity, esthesiometer type, and factors such as disease duration and severity,
underscoring the complexity of DED. As research progresses, the integration of

81
sensitivity assessment into routine clinical practice could enhance the management and
therapeutic outcomes for patients with DED. Most studies have focused on corneal
sensitivity only and there is limited evidence for other ocular surface regions including
the bulbar and palpebral conjunctiva and lid margin, which may also contribute to ocular
symptoms. A commercial, quantitative and suitably sensitive, non-contact esthesiometer
is urgently needed along with evidence-based normative and reference values for
application in different subtypes of DED.

9.6 Anesthetic challenge in DED

The TFOS DEWS II report emphasized the role of neurosensory abnormalities in the
etiology of DED and the need to evaluate somatosensory functions in its assessment.
720
The topical anesthetic challenge has emerged as an important tool for assessing
neurosensory abnormalities. This simple and rapid test can differentiate between pain
originating from peripheral nociceptor activation versus central (or non-ocular surface)
mediated pain arising from proximal sensory pathways or the central nervous system.
720
The test involves applying a drop of topical anesthetic (e.g., 0.5% proparacaine
hydrochloride) to the ocular surface and evaluating pain relief. Pain relief typically
indicates peripheral neuropathic or nociceptive causes of pain, as proparacaine
stabilizes nociceptor neuronal membranes, impeding initiation and conduction of nerve
impulses. 763 In contrast, the absence of relief suggests a central or non-ocular surface
etiology of pain, while partial improvement may indicate a mixed component to pain. 764
However, it is important to note that in patients with complete pain relief, the test cannot
differentiate between pain due to peripheral nerve abnormalities or nociceptive causes
like DED. 763 It is further not informative if no pain is present prior to anesthetic
placement (in individuals with pain that waxes and wanes).

Persistent pain after anesthesia can indicate central abnormalities in pain processing
pathways. 765 Specifically, individuals with persistent pain after topical anesthesia often
exhibit other features of somatosensory dysfunction. A study of veterans with DED
symptoms were categorized based on anesthetic challenge response. Those with
persistent pain had greater discordance between signs and symptoms and lower non-
ocular cutaneous cold and hot thresholds (see Section 9.7) compared with those where
ocular pain was reduced following topical anesthesia. 766 These data suggest increased
cutaneous sensitivity both at a site innervated by the trigeminal nerve (forehead) and at
a distant site (forearm), supporting the contribution of central mechanisms in individuals
with persistent ocular pain following anesthesia. Similarly, individuals with DED who
experience with central-dominant pain had higher ocular and non-ocular pain scores
compared with those who had greater reduction in pain with topical anesthesia
(peripheral-dominant).767 These findings have therapeutic implications as treatments for
peripheral vs centrally mediated pain vary. For instance, patients with peripheral

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dominant pain may benefit more from anti-inflammatory and topical neuromodulation,
while those with central pain may benefit from systemic neuromodulation.763 Further
studies are needed, however, that examine whether results of diagnostic tests can be
used to predict therapeutic responses.

The anesthetic challenge is a simple and accessible test that can aid in the evaluation
of somatosensory (dys)function in DED. When used appropriately, it can help identify
the location of pain generation which may assist in formulating a personalized treatment
plan for individuals with ocular pain.

9.7 Quantitative sensory testing in DED

Somatosensory function, including central abnormalities, can be investigated using

quantitative sensory testing, a method of assessing an individual’s response to various
stimuli (thermal, mechanical, vibratory). Similar to ocular sensory testing, non-ocular
function can be evaluated using detection and pain thresholds, measuring degree of
pain to a fixed stimulus, temporal summation (a phenomenon where repeated stimuli of
the same intensity cause a gradual increase in pain intensity), and aftersensations (a
sensation that persists after the external stimulus that caused it has stopped).
Abnormalities in the latter two tests (temporal summation and aftersensations) have
been linked to central abnormalities. Several studies have applied these protocols to the
study of DED (Table 9).

Table 9: Quantitative Sensory Testing in DED

Author/year Location Sample and QST Metrics Results

Size
768
United DED (n=118) Detection threshold HTPS positively
States for vibration, cool, correlated with burning
warm, cold pain, and pain, wind sensitivity,
hot pain at the and ocular pain.
forearm

Pain, aftersensation
and temporal
summation of cold
pain and hot pain
stimuli at the forearm
769
United Concordant Intensity rating of Intensity rating for
States DED (n=25) pressure pain at the pressure pain and
thumbnail auditory tones were not
Discordant statistically different in

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 DED (n=23) discordant DED.

Total DED (n
=48) Intensity rating of
auditory tones in both Pressure pain and
Controls (n= ears auditory sensitivity were
26) not correlated with DED
symptoms.
770
United DED (n=326) Detection threshold Aftersensations of cold
States for vibration, cool, pain and hot pain at the
warm, cold pain, and forehead and forearm
hot pain at the positively correlated
forearm with DED discordance
scores. Intensity ratings
at threshold of cold pain
and hot pain positively
Pain, aftersensation correlated with DED
and temporal discordance scores.
summation of cold
pain and hot pain
stimuli at the forearm
771
United LASIK (n=43) MPT at the forehead MPT at the forehead
States and forearm negatively correlated
with baseline ocular
pain.
Temporal summation
and aftersensation
intensity of MPTS positively
mechanical pain at cordrelated with
the forearm baseline DE symptoms.

Conditioned pain Aftersensations were

modulation at the associated with
forearm increased baseline
DED symptoms and
ocular pain.

Presence of
aftersensations pre-
LASIK was a predictor
for chronic DE
symptoms at 6 months
post-LASIK.
772
United DED (n=235) Aftersensation Intensity of ocular pain
States intensity of cold pain due to light was a

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 and hot pain at the predictor for the
forearm presence of
aftersensations.

DED = dry eye disease; HTPS = hot pain temporal stimulation; MPT = mechanical pain
threshold; MPTS = mechanical pain temporal summation

Hot and cold thermal stimuli (hot and cold) are frequently used to evaluate
somatosensory (dys)function. Despite variation in quantitative sensory testing protocols,
increased responsiveness to hot pain has been noted in individuals with DED where
there are increased symptoms disproportionate to the signs (measured as a
discordance score). 768 DED discordance score positively correlated with hot pain
aftersensation intensity at the forehead. 770 These studies support the finding that
abnormal perception of hot pain at sites near and distant to the eye correlate with DED
symptoms in the absence of ocular surface signs and that the altered perception may
arise from central abnormalities.

Mechanical pain can also be used as a stimulus to evaluate somatosensory

(dys)function. Prior to LASIK surgery, mechanical cutaneous pain thresholds at the
forehead negatively correlated with ocular pain intensity, indicating higher sensitivity in
individuals with more severe pain. The presence of aftersensations to a cutaneous
mechanical stimulus prior to surgery predicted symptoms of DED six month after
surgery. 771 Using a thumbnail pressure test, no significant differences were found in
mechanical pain testing or auditory testing in discordant DED compared with
concordant DED or controls, although the sample size was relatively small. 769 These
discrepancies highlight the need for more standardization in protocols when applied to
the study of DED and perhaps better definition of DED subtypes.

9.8 Brain imaging in DED

Advanced neuroimaging techniques, such as using functional magnetic resonance

imaging (fMRI), have been applied to the study of individuals with chronic ocular pain.
773,774
fMRI is a neuroimaging technique that uses a light stimulus and measures
regional changes in brain metabolism over time and has been extensively utilized in
various studies. 775 In a small study of individuals with and without ocular pain, the
chronic ocular pain group exhibited greater activation in brain regions like the primary
somatosensory, insular, and anterior mid-cingulate cortices compare with controls.
Instillation of topical anesthetic reduced activation in the primary somatosensory and
anterior mid-cingulate cortices. These results suggest different activations patterns to
light in individuals with chronic pain and photophobia, supporting the contribution of
central mechanisms in driving pain in this group. 773

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fMRI studies have also studied the neural impact of various therapies used to treat
ocular pain. In a study examining the impact of botulinum toxin A administered to the
forehead, frontalis, procerus and corrugators, showed that in those individuals who
reported reduced unpleasantness scores when viewing the light stimulus during the
post-injection scan, exhibited activation in the spinal trigeminal nucleus in response to
light stimuli prior to injections and this was not evident in those who did not show an
improved response post-injection. 776 This suggests that photophobia may be driven by
different neural pathways, and that individuals with activity within the spinal trigeminal
nucleus during light stimulation may be the ones more likely to response to botulinum
toxin. Use of the FL-41 spectacle tint to reduce photophobia led to significant reductions
in light-evoked blood oxygen level dependent signals (used as an indirect measure of
neural activity) in the bilateral primary and secondary somatosensory, bilateral insular,
right temporal pole, precuneus, anterior cingulate cortex and paracingulate cortices as
well as bilateral cerebellar hemispheric lobule VI, although the responses in regions
associated with pain processing were not eliminated completely. 777 These findings
indicate that fMRI may play a role in predicting and monitoring responses to therapy and
suggest that more studies are needed to examine central mechanisms to pain in various
DED subtypes.

9.9 Future directions and conclusions

Ocular pain and sensation are intimately associated with the structural anatomy of
corneal nerves. This structure can be altered by trauma, surgery, DED, other systemic
diseases, natural aging, and incomplete or improper neural regeneration. These
alterations are pleomorphic but most commonly manifest as reduced nerve fiber density
and increased nerve tortuosity. Aberrant corneal nerve anatomy generally correlates
with altered sensation, increased ocular pain, and symptoms typical of DED. Functional
abnormalities of corneal nerves are also implicated in altered ocular sensation and
include changes in growth factor activity and increased expression of sodium channels.
Ocular pain, in some individuals can be driven by generalized dysfunction of peripheral
and central nervous systems, with studies supporting activation of the primary
somatosensory, insular, and anterior mid-cingulate cortices in individuals with chronic
ocular pain, which may be mitigated with certain therapies.

Research into this field is ongoing and future developments may include the elucidation
of specific pathways controlling corneal nerve development and regeneration,
exploration of neuroimmune crosstalk in DED, greater implementation of artificial
intelligence networks in processing large bodies of data, more sophisticated
assessments of peripheral and central nerve function, and targeted treatments to
address dysfunction in an individual patient.

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10. Iatrogenic

10.1 Introduction

Iatrogenic disease is an adverse clinical condition resulting from diagnosis or medical

treatment performed by a health professional. It affects many patients worldwide in all
fields of medicine, including ophthalmology. 778 In the eye, the ocular surface and the
tear film are probably the most affected. This assumption is related to the fact that this
anterior interface represents the first target for the effect or penetration of the first line of
treatment for most eye diseases and is involved directly or indirectly in the most
common surgical and non-surgical procedures for the eye. Among all ocular surface
disorders, DED is the most prevalent. 779 In addition to topical medications and surgical
and non-surgical procedures, systemic drugs and the use of contact lenses are major
causes of iatrogenic DED (Table 9).

The TFOS DEWS II recognized the importance of this topic, and a specific report about
Iatrogenic DED was included.778 The impact of elective medications and procedures
was also highlighted in the "Lifestyle Epidemic: Ocular Surface Disease TFOS
Workshop".684 This review presents an update since these reports of the most common
iatrogenic causes of DED, including a summary of pathophysiology and
recommendations for management.

Table 10. Classification of iatrogenic DED (Reproduced from 778)

I. Drug-induced

A. Topical

B. Systemic

II. Contact lens-induced

III. Ophthalmic surgery*

A. Refractive surgery

B. Keratoplasty (Penetrating, lamellar and endothelial)

C. Cataract surgery
D. Lid surgery
E. Other surgeries

1. Conjunctival surgery
2. Glaucoma surgery
3. Vitreoretinal surgery
4. Strabismus surgery
5. Intrastromal corneal ring segment implantation

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6. Others

IV. Non-surgical ophthalmic procedures

A. Botulinum toxin
B. Crosslinking (CXL)
C. Cosmetic procedures
D. Others

V. Non-ophthalmic conditions

A. Graft-versus-host disease (GVHD)

B. Others

*Ophthalmic surgeries were extensively reviewed previously and are not included in this
update. 684,778

10.2 Topical drug-induced DED

10.2.1 Prevalence

As mentioned in TFOS DEWS II, evaluating DED caused by topical medications is

challenging, as clinical trials often exclude patients with ocular surface diseases, which
may lead to an underestimation of symptoms of DED.778 Most epidemiological studies
evaluate the effect of anti-glaucoma topical medications on the ocular surface. Given
glaucoma and DED are two common conditions that can occur concurrently in the same
individual, these results may be confounded. Furthermore, patients undergoing
treatment for glaucoma frequently show signs of deterioration in the ocular surface. 780

In a multicenter study conducted across four European countries with 9,658 patients,
over 40% of individuals treated for glaucoma reported symptoms of DED, such as pain
or discomfort during the application of eye drops, foreign body sensation, dry eye, and
burning.781. Furthermore, more than 20% of patients exhibited signs of blepharitis,
conjunctival hyperemia, or keratitis. These findings were more prevalent in patients
treated with preservative-containing eye drops. Similarly, a recent cross-sectional
comparative study with 320 patients concluded that glaucoma patients are more
affected by DED than non-glaucoma patients, showing a lower TBUT and greater
corneal staining in eyes with glaucoma using multiple eyedrops and daily doses.782
Another cross-sectional study with 101 patients undergoing anti-glaucoma treatment
detected signs and symptoms of DED in over 50% of the patients and advanced
changes in the ocular surface in 27% of the individuals examined.783 Reduced values on
the Schirmer test were observed in 61%, decreased TBUT in 78%, and staining of the
ocular surface in 22% of the subjects.

88
In a German study involving 20,506 glaucoma patients from 900 centers, the
prevalence of DED was higher in women (56.9%) than in men (45.7%), and this
difference was more pronounced in patients over 50 years old. The prevalence
increases with age and duration of glaucoma, and it occurs more frequently when three
or more medications are used.784 In another epidemiological study including 4,107
glaucomatous patients, the most frequently reported dry eye symptoms were: excessive
discomfort after applying anti-glaucoma eye drops (43%), pressure behind the eyelids
(40%); foreign body sensation (31%); dry eye sensation (23%); excessive reflex tearing
(21%); and eyelid itching (18%) of cases. These symptoms were significantly more
prevalent when using preservative-containing than preservative-free eye drops.785,786

10.2.2 Topical drugs contributing to DED

According to TFOS DEWS II, various topical medications and excipients have been
implicated in DED. 778 (Table 11) Of particular concern is the use of chronic medications,
such as anti-glaucoma, antiallergic, and anti-inflammatory eye drops. However, there
needs to be more specific data on active compounds, as ophthalmic formulations are
often evaluated with preservatives, making it difficult to isolate the impact of the
medications, preservatives, and excipients. Also, common excipients in ophthalmic
preparations, such as solutions, ointments, suspensions, and emulsions, may cause dry
eye symptoms. 778 The chemical properties of the formulation, including
isotonicity/hypotonicity and pH, can influence the tear film and local tolerance after
application. 778

Topical drug-induced DED has mainly been studied in individuals in long-term treatment
for glaucoma and ocular hypertension. 778 Other than the presence of benzalkonium
chloride, the main risk factors for iatrogenic DED in anti-hypertensive topical drug users
are the treatment duration, higher IOP, and glaucoma severity. 787,788 The frequency of
mild or greater DED symptoms tended to increase with an increasing number of anti-
glaucoma medications. 788 Interestingly, patients on brimonidine performed the worst.
Patients on timolol reported pain induced by light, and those on latanoprost complained
of stinging. 788

Table 11. Topical drugs implicated in DED 778

Agents used to treat glaucoma

Betablocking agents (eg: Betaxolol, Timolol)

Adrenergic agonist drugs (eg: Apraclonidine, Brimonidine)

Carbonic anhydrase inhibitors (eg: Brinzolamide, Dorzolamide)

Cholinergic agents (eg: Pilocarpine)

89
Prostaglandin analogs (eg: Bimatoprost, Latanoprost, Travoprost)

Agents used to treat allergies (eg: Emedastine Olopatadine)

Antiviral agents (eg: Aciclovir, Idoxuridine, Trifluridine)

Decongestants (eg: Naphazoline, Tetryzoline)

Miotics (eg: Dapiprazol, Pilocarpine)

Mydriatics and cyclopegics (eg: Cyclopentolate Tropicamide)

Preservatives (eg: Benzalkonium chloride)

Topical and local anesthetics (eg: Proxymetacaine, Tetracaine)

Topical ocular NSAIDs (eg: Bromfenac, Diclofenac, Ketorolac, Nepafenac)

10.2.3 Mechanism

Topical medications can affect the ocular surface through various mechanisms, causing
allergic, toxic, and/or immune-inflammatory effects or through chemical interactions with
different components of the ocular surface. 778 These effects can result from disruption
of the lipid layer of the tear film due to the detergent properties of the compounds,
reduced aqueous secretion, damage to the ocular surface epithelium, neurotoxic effects
on the corneal nerves, and injury to the eyelids, including the skin or meibomian glands.
789-792

Indirect effects may also arise if chronic inflammation induced by the topical medication
stimulates the precursors of the keratinized envelope, leading to the entrapment of
mucous cell contents and squamous metaplasia. 790 Keratinization of the eyelid margins
can further worsen MGD. 789 Additionally, destruction and/or dysfunction of goblet cells,
which are increasingly recognized as having an important role in immunomodulation of
the ocular surface, can exacerbate chronic inflammation. 793

As reported in TFOS DEWS II, distinguishing between spontaneous changes in ocular

surface disease and medication-induced effects presents a clinical challenge. 778
Conjunctival allergic reactions caused by eye drop use may be indicative, but both
conjunctival congestion and papillary conjunctivitis can occur with or without atopy.
Additionally, delayed allergic reactions may arise, often mimicking blepharitis with mild
inflammation. 778 Similarly, determining whether corneal staining is due to pre-existing or
induced DED, or caused by toxic epithelial damage and corneal melting, such as that
induced by overuse of anesthetics or non-steroidal anti-inflammatory agents, can be
complicated. With many factors to consider, the relationship between eye drop use and
ocular inflammation, tear film instability, or ocular surface staining is often difficult to

90
establish, especially when treatment is essential for severe or vision-threatening
conditions. This is particularly true in glaucoma, where treatments are usually prolonged.

10.2.4 Role of preservatives and excipients

Preservatives used in topical ophthalmic medications are derived from different

chemical families, such as mercury derivatives, alcohols, parabens,
ethylenediaminetetraacetic acid, chlorhexidine, and quaternary ammonium. They act as
a surfactant to solubilize ionic components, facilitating stabilization of medications, and
inhibit microbial activity. 792 Benzalkonium chloride is a quaternary ammonium
compound widely used as a preservative in eye drops that has its toxicity to the ocular
surface well-documented. Benzalkonium chloride concentrations as low as 0.0001%
can cause damage to corneal epithelial cells, with more severe toxic effects observed at
higher concentrations, such as 0.01% and 0.2%. 792,794

The impact of benzalkonium chloride on the ocular surface involves multiple

mechanisms of action. One main effect is through mitochondrial dysfunction, which
leads to dysfunction and subsequent cell apoptosis. 390,790 Additionally, exposure
increases the production of reactive oxygen species, inducing oxidative stress in
corneal epithelial cells. 390,790 This stress directly contributes to the apoptosis and
damage to ocular surface and trabecular cells.

Benzalkonium chloride can activate various inflammatory pathways, increasing the

production of the inflammation marker HLA-DR and the expression of pro-inflammatory
cytokines such as IL-6, IL-8, and CCL2, resulting in inflammation of the ocular surface
and worsening of DED symptoms. 790-792 Furthermore, benzalkonium chloride causes
DNA damage, contributing to cytotoxicity and reduced cell viability. It also affects gene
expression in trabecular and ocular surface cells, impacting genes related to apoptosis
and inflammation, such as Fas and caspase-3. 795,796

Corneal neurotoxicity is another harmful effect of benzalkonium chloride, which

decreases corneal nerve fiber density and reduces tear production while also causing
nerve inflammation and degeneration. 790,797 These combined effects result in significant
damage to ocular surface cells, exacerbating conditions such as DED. 798 Recent
research suggests that benzalkonium chloride may affect both the mucin and lipid
layers of the tear film, as damage to goblet cells and Meibomian glands has already
been documented, leading to tear film instability. 326,786,799

Increased tear osmolarity has also been observed in patients using preserved eye
drops compared to preservative-free topical medications. 262 Once the tear film loses its
protective properties, the compromised tear film not only leads to symptoms of DED and
corneal damage but can also spread cytotoxic inflammatory mediators across the ocular

91
surface. Consequently, increased corneal epithelial permeability has been observed in
DED with more significant deterioration when using artificial tears containing
benzalkonium chloride compared to preservative-free drops. 800 Changes in the tear film
may, therefore, initiate a series of biological alterations on the ocular surface, leading to
subsequent neurogenic inflammation and further tear film impairment, creating a vicious
cycle. New preservatives, such as Polyquad, Purite, and SofZia, have significantly
reduced cytotoxic effects compared to benzalkonium chloride. However, their impact on
DED patients needs to be further investigated. 262,684

Preservative-free eye drops are associated with better ocular surface health and higher
tolerability. 800 A systematic review of randomized clinical trials comparing beta-blockers
with and without preservatives in patients with glaucoma or ocular hypertension found
that, although the difference in intraocular pressure reduction was not clinically relevant,
preservative-free eye drops performed better in tear film breakup time and Schirmer test.
800
Additionally, in patients with DED, preservative-free eye drops demonstrated a
significant reduction in ocular inflammation symptoms and increased antioxidants in the
tear film compared to preserved eye drops. 801 Another observational study in patients
with DED who switched from preserved drops to preservative-free drops containing
hyaluronate showed significant improvement in the OSDI and reduced frequency of
superficial punctate keratitis. 802

The use of preservative-free prostaglandins has also shown significant benefits for
ocular surface health compared to preservative-containing versions. The PRAMOS
study demonstrated that patients using preservative-free prostaglandin analogs had a
lower prevalence of conjunctival hyperemia and corneal staining compared to those
using preserved eye drops. 803 A prospective study showed that switching from
preservative-containing prostaglandin-timolol fixed combinations to a preservative-free
formulation resulted in a significant improvement in ocular surface disease symptoms,
and a reduction in conjunctival hyperemia. 804 Another randomized controlled trial
revealed that transitioning from preservative-containing glaucoma therapies to
preservative-free formulations improved both ocular surface health and intraocular
pressure (IOP) control. 589 These findings are supported by studies indicating that the
absence of preservatives, such as benzalkonium chloride, prevents chronic ocular
surface toxicity, leading to improved treatment adherence and long-term efficacy. 805,806

10.2.5 Recommendations for management

The first step is to investigate which medication is causing DED and try to stop its use.
This subtraction can be challenging when discontinuing the treatment, which presents a
risk to the eye's health. Sometimes, multiple drugs and components are involved, or
adverse effects appear long after treatment initiation, making identification of which is

92
causing DED even more difficult. 778 In some cases, eye drops may be necessary to
treat dry eye symptoms, but adding preserved drops to eyes already suffering from
dryness caused by other drops may be ineffective and worsen the condition. Once the
responsible drug is identified, efforts should be made to discontinue using the
preservative or medication. Considering that toxicity is dose-dependent, reducing the
number of preserved eye drops can minimize adverse. 778 In glaucoma patients, more
invasive and definitive options such as laser trabeculoplasty or surgery may replace or
diminish the use of topical medication when the ocular surface and QoL are very
compromised. 778

Conceivably the adverse effects of toxic preservatives should reduce with time. There is
a significant shift in first-line primary open angle glaucoma therapy in most countries,
from topical therapies towards laser therapies. 807 Preservative-free therapies are
increasingly available and there is good evidence for their ocular surface benefits. A
significant issue related to preservative-free eye drops is their cost, as they are
generally more expensive than preserved drops. This is primarily due to higher
production costs and patent fees associated with preservative-free drops. Allocation of
healthcare and out-of-pocket costs for patients vary with region and insurers or health
system payers, and preservative-free alternatives may not be available in certain
jurisdictions.

10.3 Systemic drug-induced DED

Systemic medications can contribute to DED through different mechanisms, including

reduction of tear production, disruption of nerve input and reflex secretion, inducing
inflammatory responses in secretory glands, or directly irritating the ocular surface
through their presence in tears.262,808 Some systemic medications can exacerbate
immune responses, causing further harm to the ocular surface and its innervation,
decreasing sensitivity or increasing pain, which worsens DED symptoms.262,808 This
section reviews how some of the most common systemic medications can negatively
impact the ocular surface.

10.3.1 Prevalence

At least 1/5 of the best-selling systemic drugs in the US have been associated with the
development of DED. In the elderly population, 62% of DED cases are related to
systemic medications, including nonsteroidal anti- inflammatory drugs (NSAIDs),
diuretics, vasodilators, analgesics/antipyretics, antiulcer agents, sulfonylureas, cardiac
glycosides, anxiolytics/benzodiazepines, anti-infectives, antidepressants/antipsychotics,
hypotensive agents, and antihistamines. 809

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In a secondary analysis of the Dry Eye Assessment and Management Study to evaluate
whether systemic medication use is associated with DED severity, the authors found
that 160 (30%) of the 535 participants used medications for hypertension, 129 (24%)
used statins, 118 (22%) used antidepressant medications, 117 (22%) used
antihistamines, and 16 (3%) used systemic corticosteroids. 810 A multivariable analysis
demonstrated that antihistamines and corticosteroids were associated with the highest
OSDI score among these systemic medications. Users of seizure medications had a
higher composite signs severity score compared to non-users. Compared to non-users,
antihistamines, aspirin, and vitamin D3 users had significantly worse average TBUT.
Unexpectedly, MGD scores were worse in users of vitamin D3, although this could be
confounded if those users of vitamin D3 had vitamin D deficiency. Users of diuretics had
significantly better scores for MGD compared to non-users. 810

In a Military Health System (MHS) beneficiaries database, newly diagnosed DED ( 40

000 beneficiaries) and prevalent DED (> 285 000 beneficiaries) patients were compared
with matched non-DED patients. 811 In both the newly diagnosed and prevalent DED
patients, comorbidities were significantly higher in the DED vs non-DED groups.
Systemic medication use was also significantly higher in the DED than in the non-DED
groups. The most prescribed medications were narcotic analgesics/strong pain killers,
decongestants/vasoconstrictors, b-blockers, antidepressants, diuretics, and anxiolytics.
811
Risk factors are described in the Epidemiology Section 9 above.

10.3.2 Medications and mechanisms

10.3.2.1 Tamsulosin

Tamsulosin, a medication that blocks alpha-1 receptors, is widely prescribed for treating
benign prostatic hyperplasia, but is also utilized in managing ureteral stones, prostatitis,
and female voiding dysfunction. Epidemiological research suggests a link between DED
and benign prostatic hyperplasia itself, as well as the medications used to treat it,
including tamsulosin (See Section 5.5.1).

10.3.2.2 Antihistamines/anticholinergic drugs

A large class of systemic medications that lead to the signs and symptoms of DED are
those with anticholinergic activity. The anticholinergic class covers a wide range of
therapeutic drug categories, including antidepressants, antipsychotics or neuroleptics,
antiparkinsonians, H1 antihistamines, decongestants, and antispasmodics. 812

Anticholinergic medications, such as those used for overactive bladder, have been
linked to a reduction in tear film breakup time and an increase in DED symptoms,
including burning and foreign body sensation. Studies have shown that medications like

94
solifenacin can significantly worsen dry eye symptoms and signs compared to
placebo.813,814

Oral antihistamines and anticholinergic drugs are commonly used to treat allergies and
rhinitis. These medications, especially in combinations like pseudoephedrine and
cetirizine, can increase the sensation of dryness in the eyes and mouth. 815

Results of the comparison of DED symptoms, measured by the OSDI, between users
and non-users of medications showed that patients who used antihistamines had worse
TBUT and OSDI scores compared to non-users. Anticholinergic drugs, such as
antihistamines, contribute to DED by acting on the G-protein coupled muscarinic
receptors of the lacrimal gland acini and mucus-producing conjunctival goblet cells,
affecting the production of the aqueous and mucous components of the tear film.
Furthermore, functional cholinergic receptors have been identified in the epithelial cells
of the meibomian glands, leading to decreased tear film stability. Thus, despite being
commonly used to treat allergy-related eye symptoms, antihistamines confer a high
anticholinergic burden, contributing to the development and worsening of DED signs
and symptoms.268,810,816

10.3.2.3 Isotretinoin

13-cis-Retinoic acid or isotretinoin, is a vitamin A derivative widely used in treating

moderate-severe acne due to its atrophic effects on the sebaceous glands. This
medication acts as a pro-drug that, upon conversion, induces apoptosis in various cell
types, including those of the sebaceous and meibomian glands. Toxicity may manifest
as blepharoconjunctivitis, presenting symptoms such as crusting at the eyelid margins
and conjunctival redness, and is often associated with dry eyes, light sensitivity, and
contact lens intolerance. 261,271,817

Recent studies utilizing standardized tools, such as the OSDI, indicate that DED
symptoms are more commonly found in patients treated with Isotretinoin. The drug can
lead to glandular atrophy, ductal keratinization, and small fiber neuropathy, which may
reduce corneal sensitivity.818,819 Other studies have reported severe DED symptoms
and decreased tear film stability among patients who start using Isotretinoin, associated
with reduced TBUT, lower Schirmer test value, and decreased central corneal
thickness.275,820 Additionally, a few studies show that even after discontinuation of the
medication, these signs and symptoms can persist for months post-treatment.275,820 The
adverse effect of Istotretinoin on meibomian gland lipid production is likely to be through
suppression of the peroxisome proliferator-activated receptor γ pathway which inhibits
meibocyte differentiation and meibum characteristics. 821Topical isotretinoin is also
recognised as a risk factor for iatrogenic DED (see Section 6.6.4). 822

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 10.3.2.4 Chloroquine / Hydroxychloroquine

Hydroxychloroquine is a medication commonly used to treat Sjögren disease and other

rheumatologic diseases, primarily aimed at relieving symptoms related to joint pain and
fatigue, with a well-recognized anti-inflammatory effect. Research on the
ophthalmological effects of hydroxychloroquine is highly controversial, as studies from
the early 21st century have shown that the substance can be secreted into the tear film,
potentially worsening DED symptoms. 823 On the other hand, other studies indicate that
patients with Sjögren disease did not experience significant clinical improvement in
xerophthalmia. 824,825 Likewise, other studies have shown no improvement in symptoms,
tear production, corneal staining, or inflammatory markers after up to 24 weeks of
treatment, with patients not experiencing any symptom relief. 824,826 The divergent
results regarding its efficacy may stem from the common use of hydroxychloroquine in
the treatment of autoimmune disorders, which are often associated with DED,
highlighting the need for further studies on this medication.

10.3.2.5 Corticosteroids and non-steroidal anti-inflammatories

Research on the impact of non-steroidal anti-inflammatory drugs on the ocular surface

is less extensive, and more recent studies should be conducted. However, selective
cyclo-oxygenase-2 inhibitors, such as celecoxib and rofecoxib, have been associated
with ocular side effects, primarily conjunctivitis and blurred vision. These effects
disappear within 72 hours of discontinuation. Although other ocular side effects are less
common and lack clear evidence of a direct link, the consistent findings of blurred vision
and conjunctivitis suggest a potential connection. The occurrence of these ocular
symptoms should lead physicians to consider discontinuing the medication, as this
action generally resolves the symptoms without long-term effects. 827

Aspirin, a non-selective and irreversible inhibitor of cyclo-oxygenase-1 and -2, is

expected to reduce the inflammatory component of DED, leading to an improvement in
signs and symptoms. 828 However, the findings of previous studies on the effects of
aspirin on DED are contradictory. For instance, a 2018 prospective cross-sectional
study involving 106 individuals without a prior diagnosis of DED showed that aspirin
users had lower tear osmolarity, increased TBUT, and lower OSDI scores. 828 However,
no significant difference in corneal staining was found despite greater symptomatic relief.
In line with these results, other older studies were unable to find an association between
aspirin and a higher prevalence of DED. 829 This underscores the need for future studies
to provide a clearer understanding of aspirin's effects on DED.

The use of corticosteroids is a common practice in medicine for treating inflammatory

disorders and has been associated with worsening DED severity, affecting both signs
and symptoms.810 However, it is important to consider the route of administration, as it

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can significantly impact the efficacy and potential side effects of these drugs. For
instance, when administered topically, corticosteroids are effective in treating DED. 830
Additionally, many users of systemic corticosteroids have underlying autoimmune
conditions that are independently associated with DED, making it challenging to isolate
whether the worsening of DED signs and symptoms is due to medication use or the
progression of the underlying disease. 96 More prospective studies are needed to
determine the effects of systemic corticosteroid use on DED. 831

10.3.2.6 Antibiotics

Antibiotics are commonly used to treat various infections, but few studies have been
conducted regarding their impact on the ocular surface. Antimicrobials, particularly
tetracyclines and macrolides, are generally used in the treatment of MGD and have
shown promising results. 832,833

The correlation between DED and antimicrobial use is sometimes indirect, as antibiotics
can cause Stevens-Johnson syndrome. Some studies have shown that antibiotics are
notable risk factors for this condition, potentially leading to severe damage to the ocular
surface. 834,835

10.3.2.7 Antidepressants / anxiolytics / mood stabilizers

The use of antidepressants and antipsychotics is strongly associated with an increased

risk of DED. A systematic literature review suggests that both depression and
antidepressant use independently contribute to the development of DED. Various
clinical and population-based studies have linked the diagnosis and severity of
depression to DED and its symptoms. Additionally, other extensive population studies
have also found an association between antidepressant use and DED. 836

Possible mechanisms for the effects of antidepressants on DED might be related to

their anticholinergic properties. Another plausible theory is that antidepressants disrupt
the corneal epithelial barrier and promote an inflammatory response on the ocular
surface by increasing serotonin levels in the tears, which in turn induces an
inflammatory response and apoptosis of corneal epithelial cells by activating NF-κB
signalling. 837,838

The prevalence of DED in patients using antipsychotics compared to the general

population, with higher prevalence also noted in those on multiple medications
compared to those on a single medication. Patients treated with clozapine
(monotherapy) and those on a combination of clozapine and quetiapine (polytherapy)
showed the highest prevalence of DED. 839

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Selective serotonin reuptake inhibitors can impact the ocular surface by impairing tear
film stability. 840 Conversely, serotonin-norepinephrine reuptake inhibitors, effective in
treating chronic pain syndromes, may help alleviate DED symptoms. Patients using
these inhibitors often have lower OSDI scores, suggesting they may be safer for the
ocular surface compared to other antidepressants. Thus, while DED is common among
antidepressant users, it may be better managed with serotonin-norepinephrine reuptake
inhibitors. 841

Lithium, widely used for the maintenance and acute treatment of bipolar disorders, has
a narrow therapeutic index and various side effects, making its management
challenging. Although the adverse effects of lithium are well-documented, its ocular
impacts are less understood, which may affect patient compliance. Ocular side effects
associated with lithium use include exophthalmos, abnormal eye movements, ocular
myasthenia gravis, papilledema, photophobia, and alterations in the tear film, which can
lead to DED. 842

Given the complexity of these interactions, co-management of individuals with DED and
medicated for mental health conditions is advisable.

10.3.2.8 Hormone replacement therapy

DED is common among postmenopausal women, with estrogen hormone imbalance

being identified as a potential causative factor (see Section 6.6). 310,843 Hormone
replacement therapy may be used to relieve menopausal symptoms, using either
estrogen alone or in combination with progesterone or progestin. However, estrogen
replacement therapy after menopause has been associated with an increased
prevalence of DED (See Section 6.6). 129 Each additional three years of hormone
replacement therapy resulted in a significant 15% increase in the occurrence of dry
eye.843 The same study showed that the onset of DED was linked to initiating estrogen
therapy.843 Conversely, the use of androgens, both topically and systemically, have
been reported to improve the signs and symptoms of MGD and DED. 3,48,844,845

10.4 Contact lenses and DED

The relationship between contact lens wear and DED is complex. Contact lens wear is
recognised as a consistent risk factor for DED (Section 6.6). Ocular symptoms occur
more commonly in contact lens wearers than non-wearers and many large
epidemiological studies have used ocular symptom reporting as a surrogate for DED. 149
Signs such as corneal staining and tear film instability, due to the partitioning of the tear
film during wear occur. 846 Signs of meibomian gland alterations are also more common
in contact lens wearers than in age-matched non-wearers. 847 Contact lens discomfort is
characterized by episodic ocular symptoms of discomfort and dryness which resolve

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when the contact lens is removed, as distinct from those in DED, 284 however this
distinction is not consistently considered when reporting symptoms in contact lens
wearers. A further complexity is that other complications of contact lens wear may also
lead to very similar ocular symptom-reporting. 847 Contact lens wearers may also have
existing or subsequently develop, DED. This has led to the use of series of descriptors
which are often used interchangeably, including contact lens discomfort, contact lens-
induced dry eye and contact lens-associated dry eye. With these caveats, this section
will endeavour to summarise the evidence for prevalence of, mechanisms underlying
and remedial strategies for DED associated with contact lens wear.

10.4.1 Prevalence of DED in contact lens wear

The prevalence of symptoms of DED in soft contact lens wearers in a series of recent
prospective and retrospective studies in select population groups ranged between 31%
to 77%. 848-850 A clinical diagnosis of DED by an eyecare professional was made in
24% of office workers in a Chinese sample 850 and in 14% of University students
wearing contact lenses in Thailand. 851. The frequency of DED in scleral lens wearers
has been more difficult to determine. These are mostly used for medical and therapeutic
indications, and an estimate of 56% of scleral lens wearers had severe DED symptoms
based on OSDI. 852,853 There is a lack of large-scale epidemiological studies on the
prevalence of DED associated with orthokeratology lenses. In a small retrospective
study of contact lens complications conducted at a tertiary hospital in China, 24% of
orthokeratology lens wearers presented with DED. 854 Understanding the pathogenesis,
and provision of mechanism-oriented treatment of DED in contact lens wearers is
important.

10.4.2 Mechanism

Contact lens use causes a variety of biophysical and biochemical changes in the tear
film, as well as alterations in the ocular surface structure and function which are
summarized in Table 12. These mechanisms collectively lead to reduced tear film
stability, increased tear evaporation, and ocular surface damage, 846,847 and likely
contribute to DED in contact lens wearers.

Rigid and soft contact lens wear increase tear film evaporation,855 although they interact
differently with the ocular surface. Mechanical interactions are different between rigid
and soft contact lenses due to material and fitting characteristics. 847,856,857 Upper lid
wiper epitheliopathy is more common in rigid than soft lens wear. 858 There is a higher
frequency of incomplete blinking, 859 leading to uneven tear distribution and
exacerbation of dry eye symptoms. Corneal staining in the 3 to 9 o'clock positions is
common among rigid lens wearers,860,861 attributed to thinning of the tear film 862

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adjacent to the lens edge, whereas soft contact lens wearers more often exhibit inferior
arcuate cornea staining.861

It is conceivable that mechanisms in contemporary scleral lens use might resemble

those observed with rigid corneal contact lenses, including friction between scleral
lenses and ocular tissues, 856 increased mechanical stimulation of the ocular surface, 855

and tear film instability.863

Orthokeratology lenses share some mechanisms for DED with soft and rigid contact
lenses, but the unique mechanism of these lenses is related to their reverse geometry
design, 854 which not only alters corneal shape but also significantly affects the density
and distribution of corneal nerve fibers, 864,865 which may not fully recover shortly after
cessation of wear. 866,867

The evidence for changes in corneal sensitivity underpinning sensation report in contact
lens wear is limited. Contemporary rigid corneal and soft lenses do not appear to alter
corneal sensitivity, although a reduction in central corneal mechanical sensitivity is
observed in orthokeratology lens use 866,868 Reduced corneal sensitivity may reduce tear
secretion and blink frequency, thereby exacerbating symptoms of DED. 867 There is a
substantial augmentation in tear inflammatory mediators in orthokeratology lens wear,
such as IL-17A, IL-6, and prostaglandin E2. 864 The TFOS Contact Lens Discomfort
neurobiology report described the multifactorial effects of contact lens wear, include
mechanical, cooling, drying, change in osmolarity and chemical impacts on ocular
surface neurosensory processes. 869 Altered sensory processing of cooling stimuli
delivered to the cornea has recently been reported in symptomatic soft contact lens
wearers suggesting nerve sensitisation or maladaptation in the absence of sensitivity
changes, 870 which is intriguing but causality for DED has not been demonstrated.

Subclinical inflammation at the ocular surface is reported in several modalities of

contact lens wear, evidenced by recruitment and activation of inflammatory cells to the
cornea, conjunctiva and lid margin and the presence of mediators in tears 846 for review).
In naïve wearers, epithelial immune cells are recruited to the central and peripheral
cornea within 2 hours of soft contact lens wear and morphological changes suggest
enhanced antigen capture capacity. 871 Increased density of inflammatory cells is seen
at the lid margin in reusable hydrogel and silicone hydrogel wearers but not in daily
disposable wear. 872 The link between subclinical inflammation and DED in contact lens
wearers, however, has not been confirmed.

Table 12. Effects of contact lens wear on the tear film and ocular surface

Biophysical tear film alterations Effect

Division of the tear film into pre- and post-lens layers

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Tear meniscus volume Reduced
Lipid layer spreading Reduced
Tear stability Reduced
Structural ocular surface alterations
Conjunctival goblet cell density↓ Reduced
MG expressibility ↓, Reduced
MG obstruction and atrophy Increased
Basal corneal nerve density↓, nerve tortuosity↑: Reduced
Ocular surface sensitivity Corneal sensitivity unchanged in SCL use;
altered sensory processing; conjunctival
sensitivity increased
Corneal sensitivity reduced in OK use
Ocular surface friction and lid wiper epitheliopathy Increased
Biochemical tear film alterations
Tear cholesterol Increased
Malondialdehyde & 4-hydroxy 2-nonenal Increased
Beta-2 microglobulin Reduced
Proline rich protein 4 Reduced
Lacritin Reduced
Lipocalin 1D1 Reduced
Secretory IgA Effects equivocal, may depend on duration
of wear and wear modality
Albumin Increased
Deleted in Malignant Brain Tumors-1 Increased
Prolactin inducible protein Increased
MUC5AC Reduced
Inflammatory tear film/ocular surface changes
Epithelial immune cells Increased at ocular surface in SCL wear,
increased antigen capture capacity. Density
reduced at central cornea in OK
Tear cytokines IL-7,8,13,15 Increased

SCL = soft contact lens; OK = orthokeratology

10.4.3 Recommendations for management of DED in contact lens wearers

Therapeutic and device-related management of DED in contact lens wearers broadly

follows that described in the TFOS DEWS III Management and Therapy report, (Jones
et al., 2025) with caveats about the use of preserved medications during contact lens
wear and noting that many products are not specifically registered for use in contact
lens wearers. Given the potential interdependencies between DED, MGD and contact
lens discomfort, non-specific strategies might include switching to daily disposable
contact lenses, attention to the lens fitting relationship, avoiding preserved care systems
with reusable contact lenses, improving blink completeness, attention to environmental
triggers, ruling out ocular or systemic co-morbidities and recommending the use of

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unpreserved lubricants, with or without lipid additives. 847 873 874 Discontinuing from lens
wear or reducing wear time may be successful 855 unless there is underlying DED.

In contact lens wearers with MGD and consequential evaporative DED, there is
evidence for the benefits of treatments including IPL 875, microblepharon exfoliation 876
or thermal pulsation, 877 including improved TBUT, and symptoms of DED. Low level
light therapy or photobiomodulation therapy has been used in the treatment of MGD and
evaporative DED 2, but there is no high-level evidence for benefit in contact lens
wearers, although an observational study without a control group exists in wearers with
discomfort. 878 While there are no pharmacological treatments approved for MGD in
contact lens wearers, a phase 1 879 and phase 2 880 clinical trial both demonstrated
benefit in improving comfortable wear time and meibomian gland signs in a population
of symptomatic wearers with MGD, using a topical selenium sulphide treatment.

10.5 Procedures

10.5.1 Botulinum toxin

Paradoxically, botulinum toxin A has been used in treating DED. 881 In the literature,
BTX-A was injected into the periorbital area including the medial portion of the
orbicularis muscle and not solely into the lateral canthal region, for the treatment of
existing DED. The mechanism of action of botulinum toxin on lacrimal drainage in such
instances has been suggested to be due to a paralysis of the orbicularis oculi muscle
around the canaliculi with a decreased compression as well as weakness of apposition
of the puncta during blinking. 882,883

10.5.1.1 Mechanism

Dry eye due to botulinum toxin type-A injection for treatment of blepharospasm or after
blepharoplasty and peri-orbital surgery has been previously reported in literature. The
mechanisms by which DE is induced are multifactorial:

10.5.1.2 Direct inhibition of tear secretion:

Botulinum toxin blocks acetylcholine release within the lacrimal gland, disrupts
neuromuscular junctions, and inhibits parasympathetic nerves, thereby suppressing
both basal and reflex tear secretion. 884 This leads to a significant decrease in TBUT
and Schirmer test results. 882,883,885

10.5.1.3 Impact on meibomian gland function:

Botulinum toxin can also diminish lipid production by affecting neuro-muscular

transmission around the muscles of the meibomian glands. 886 Additionally, botulinum

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toxin-induced paralysis of the preseptal orbicularis oculi and Riolan's muscle weakens
eyelid closure and reduce meibum secretion, thereby decreasing lipid layer thickness
and tear film stability. 887 However, further studies are needed to examine the long-term
effects on the meibomian glands.

10.5.1.4 Regulation of inflammatory responses:

Botulinum toxin injections may trigger inflammatory responses at the ocular surface and
in the lacrimal glands, such as increased release of cytokines (e.g., TNF-α and IL-1β),
888
which may further impact tear stability and tear secretion.

10.5.1.5 Chemodenervation of orbicularis oculi:

Botulinum neurotoxin induces chemodenervation of the orbicularis oculi muscle which

can lead to poor blinking, lagophthalmos, and ectropion that may result in corneal
dryness with symptoms of irritation, foreign body sensation, and epiphora. Epiphora is
likely due to a combination of factors including DED causing reflex tearing, hypotonicity
of the medial pretarsal fibers causing decreased outflow of tears, and malposition of the
eyelids, causing impaired retention of tears.884

10.5.1.6 Recommendations for management

1. For patients experiencing DED due to botulinum toxin treatment for other conditions,
individualized management plans should be developed based on specific symptoms
and tear volume.884
2. Patients undergoing botulinum toxin treatment should be regularly monitored for
tear secretion levels, ocular surface condition, inflammatory responses, and
changes in orbicularis muscle function. Patients should be educated about
symptoms of DED and management, including maintaining good eye hygiene
practices and avoiding excessive use of electronic screens.
3. Grading eye lid muscle tone and eye lid laxity and documenting any eye symptoms
or findings prior to injection is of importance. It is recommended that patients who
are to undergo botulinum toxin injections should undergo the snap-back test which
measures muscle tone and the lower lid distraction test that measures lid laxity
resulting primarily from the stretching of canthal ligaments. Patients with severe test
results are more prone to dry eye development after injection of the toxin into the
lateral canthal region. Patient should be asked about any early symptoms including
eye irritation, foreign body sensation, or tearing that were not present previously
and/or by noticing any change in snapback and distraction tests during the
treatment course. In presence of positive test findings and dry eye symptomatology,
one should temporarily discontinue repeated BTX-A injections into the lateral
canthal rhytids. 889

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10.5.2 Corneal collagen crosslinking

10.5.2.1 Mechanism

Corneal collagen cross-linking (CXL) is a treatment that uses ultraviolet A light and
riboflavin (vitamin B2) as a photosensitizer to strengthen the cornea. This technique
reinforces the chemical bonds within the cornea, thereby halting or delaying the
progression of corneal ectatic diseases such as keratoconus.890,891 Both epithelium-on
and epithelium-off CXL have been reported to have a positive effect on DED, and may
improve tear film homeostasis and reduce DED symptoms in patients with
keratoconus.892

Paradoxically, CXL can also induce DED symptoms through multiple mechanisms.
Firstly, epithelial removal and delayed healing may induce DED.893-895 This delay can
affect the uniform distribution of tears and the corneal defences,893,896,897 by
compromising tear film stability708,898 and making the ocular surface more prone to DED
symptoms.

CXL may also reduce corneal nerve density, 899 impairing corneal sensory nerve
function and decreasing corneal sensitivity, which in turn affects the tear secretion. 899
However, studies indicate that in most patients subbasal nerve regeneration occurs
within 2-3 months after CXL,899 reaching preoperative levels in 6-12 months. 900,901
Additionally, although CXL does not directly alter meibomian gland morphology, 902 the
effects on corneal sensory nerves 902, the use of eyelid speculums during surgery, post-
CXL inflammatory responses with elevation of IL-6, and medications used post-surgery
902
can indirectly affect gland function and secretion quality.

10.5.2.2 Recommendations for management

To effectively manage DED that may arise after CXL, a comprehensive approach is
essential. Firstly, bandage contact lenses, due to their material and characteristics, help
protect exposed nerve endings and the corneal epithelium, reducing frictional damage
from blinking and thereby alleviating dry eye symptoms. 903

Secondly, during the postoperative phase, the use of corneal protectants, tear
substitutes, and tear secretagogues is important. These medications should be selected
based on their ability to replace the mucin glycocalyx lost from the corneal epithelial
cells, promote the restoration of the mucin gel on the epithelial cell surface, enhance the
adhesion of the tear film to the corneal epithelium, and aid in the recovery of the lipid
layer of the precorneal tear film. 903,904

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Lastly, advancements in modern CXL technology also help reduce the risk of
postoperative complications. These improvements include transepithelial cross-linking,
localized and personalized techniques, and accelerated procedures, which aim to
enhance surgical outcomes and minimize corneal manipulation time. 898,905-907

10.5.3 Other procedures

10.5.3.1 Eye cosmetics and beauty treatments

The use of eye cosmetic products and procedures represent a lifestyle challenge that
may exacerbate or promote the development of ocular surface and adnexal disease.
This topic has been recently explored in detail. 263

10.6 Non-ophthalmic conditions

10.6.1 Radiotherapy

10.6.1.1 Mechanism

DED is associated with radiation therapy for head and neck cancer, 908,909 temporal
tumors, breast cancer, and Graves' eye disease. This condition often results from direct
damage to tissues around the eye.909-911 The incidence of radiation-related dry eye
depends on the type of radiation therapy, tumor location 912,913 and radiation
dose.909,914,915

Mechanisms of Radiation-Induced Dry Eye are multifactoral:

(1) Damage to the Lacrimal Glands and Tear Drainage System: Radiation therapy can
cause inflammation of the tear ducts or obstruction of tear duct openings,916,917
significantly reducing lacrimal gland function and leading to DED symptoms.
Ultrastructural analysis of post-radiation lacrimal glands reveals extensive tissue
damage, including membrane rupture, loose cell junctions, and nuclear
fragmentation.918 Additionally, the expression of markers such as lysozyme, S-100, and
CD117 is reduced in the lacrimal tissues.918
(2) Corneal and Conjunctival Damage: Radiation therapy can cause direct damage to
corneal epithelial cells and loss of conjunctival goblet cells,914 which can destabilize the
tear film. Additionally, inadequate conjunctival closure during brachytherapy can lead to
conjunctival and scleral necrosis, further affecting ocular surface health and tear film
stability.919
(3) Eyelid Damage: The meibomian glands exhibit acute inflammatory responses to
radiation therapy characterized by cystic dilatation of ducts containing keratin, and
atrophic decrease of the glands and ducts or their complete loss.920

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 10.6.1.2 Recommendations for management

The management of radiation-related DED can be divided into three key approaches:

1. Preventive measures to minimize the risk such as limiting the radiation dose to
the lacrimal glands, ideally keeping the dose below 30 Gy.914,915,921,922
2. Proper patient positioning, eye shielding, and adequate patient immobilization
during radiotherapy sessions to reduce radiation- related toxicity.912
3. Therapeutic interventions for DED. 2

Long-term management is essential for maintaining good ocular health and QoL.
Regular ophthalmologic examinations and assessments to monitor changes in DED
allow for timely adjustments in treatment plans and interventions. A multidisciplinary
team, including ophthalmologists and oncologists, can provide comprehensive care and
personalized treatment plans, thereby maximizing QoL.

10.6.2 Bariatric surgery

292,684
This topic has been reviewed in detail in two recent reports.

10.6.3 Stem cell or bone marrow transplant

10.6.3.1 Mechanism
GVHD is a common complication following human leukocyte antigen (HLA)-matched
allogeneic hematopoietic stem cell transplantation. GVHD can affect various organs,
including the skin, gastrointestinal tract, liver, lungs, and eyes, with dry eye being the
most common manifestation of ocular GVHD. 923,924 Ocular GVHD is seen in 60-90% of
transplant recipients, primarily manifesting as secondary inflammation and fibrosis of
the lacrimal and meibomian glands. 925,926 The mechanisms of DED in GVHD include
the following aspects:
1. Lacrimal Gland Fibrosis: Fibrosis of the lacrimal gland is a major cause of GVHD-
related dry eye. Studies have shown the presence of activated CD34+ fibroblasts
in the lacrimal glands of GVHD patients, with T-cell infiltration around the ducts,
thickening of blood vessels and duct basement membranes correlating with the
severity of fibrosis. 927,928
2. Renin-Angiotensin-Aldosterone: Angiotensin 1 type 1 receptor antagonists, such
as valsartan, can reduce fibrosis, decrease inflammatory cell density, and
increase tear secretion. 929 Angiotensin-mediated fibroblast activation and
upregulation of TGF-β expression via the angiotensin 1 type 1 receptor are the
primary mechanisms of fibrosis. 930

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 3. Immune-Mediated Inflammation: In GVHD patients, increased density of T cells
and dendritic cells 931 in the lacrimal glands, conjunctiva, and cornea contribute to
inflammation and fibrotic responses. 932
4. Cellular Senescence: Chemotherapy and radiotherapy-induced DNA damage
and the inflammatory environment activate senescence-related molecular
pathways. 933 Stress-induced cellular senescence promotes the secretion of
various cytokines (e.g., IL-1β, IL-6, IL-8), forming the "senescence-associated
secretory phenotype", which exacerbates inflammation. 934

GVHD patients often exhibit significant abnormalities in the meibomian glands, 935
leading to alterations in the tear film lipid layer and increased tear evaporation. 936
Corneal and conjunctival changes commonly include punctate keratopathy, filamentary
keratitis, and epithelial defects, 937 accompanied by decreased corneal nerve fiber
density, increased dendritic cell density, conjunctival epithelial squamous metaplasia,
and reduced goblet cell density. These changes collectively contribute to the onset and
progression of DED. 931,936

10.6.3.2 Recommendations for management

GVHD-related DED can be potentially managed through a combination of preventive

and treatment strategies. Preventive measures primarily include ensuring high-quality
HLA matching between the donor and recipient, and the use of immunosuppressive
agents such as cyclosporine, tacrolimus, and methotrexate to inhibit donor T cells.938,939
Preservative-free and phosphate-free eye drops are recommended for tear retention
and lubrication. 940
Topical steroids, 941,942 and systemic oral immunosuppressants such as cyclosporine 943
are recommended to control inflammation. Biological tear substitutes like autologous
serum eye drops provide significant anti-inflammatory and nutritional benefits. 944-946 In
severe cases of DED, temporary tarsorrhaphy may be necessary to limit ocular surface
exposure, and surgical interventions may be required to treat cicatricial eyelid diseases.
947
These comprehensive measures aim to maximize symptom relief and enhance the
QoL for patients suffering from GVHD-associated DED. (Cross reference with
management report

10.7 Future directions and conclusions

Iatrogenic causes underpin a significant proportion DED and may arise from topical and
systemic medications and a range of ophthalmic and non-ophthalmic surgeries.
Lifestyle choices such as contact lens wear, cosmetics and cosmetic procedures also
contribute to iatrogenic DED. Improved understanding of the range of causes and how
these contribute to different subtypes of DED and risk factors for more severe disease is

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likely to help reduce their impact. Evaluating and managing underlying DED pre- and
post-intervention will also better support these patients. A challenge in iatrogenic
disease is understanding the impact of an intervention compared with progression of the
primary condition. This report has also identified the need to better disaggregate the
effects of topical medications alone from their preservatives and other excipients. In
DED following use of systemic medications, there is greater certainty in the adverse
effects of isotretinoin in DED and the persistence of effect once the medication ceases.
While depression and antidepressants broadly are independent risk factors for DED, it
is recognised that some classes, such as serotonin-norepinephrine reuptake inhibitors
may improve ocular symptoms and there should be further exploration of the impact of
systemic corticosteroids on the ocular surface. The natural history of DED associated
with lifestyle choices such as contact lens wear and particularly orthokeratology remains
unclear.

11. Clinical trials design

11.1 Introduction

The major international markets for medicines are the US, EU and Japan. These three
regions have consistent regulatory frameworks for approval of medicines following the
International Council for Harmonisation of Technical Requirements for Pharmaceuticals
for Human Use guidelines. 948 There are some variations in evidentiary standards
between these jurisdictions in clinical trials designs and features. As an example, for
new DED treatments, the EU requires a six-month time frame for primary efficacy
studies, rather than the three-months required by the FDA. In addition, EU typically
requires standard of care as one of the trial arms, in addition to vehicle as control,
where using artificial tears as standard of care. Artificial tears are considered devices in
the EU but not in the US or Japan. As far as possible, this report will provide examples
of treatments approved from of these three regions.

The development of novel treatments for DED accelerated in the US and EU following
release of both the TFOS DEWS II report in July 2017 205 and the FDA draft guidance
for industry in December 2020. 949 Five new treatments for DED were approved
between August 2018 and June of 2024 in the US, one new treatment was approved in
the EU and one new treatment was approved in Japan. Eight treatments were approved
before July of 2017, two in the US and six in Japan. 950-952 While prior TFOS DEWS
reports focused on challenges hampering the design and success of trials in DED, 953
this report will focus on how recommendations from the TFOS DEWS II report
translated into innovations in trial design, an independent analysis of trial failures
released in 2012, across ten therapeutic areas, 198 and how flexibility in application of
the FDA draft guidance have improved success rates for bringing novel treatments to

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patients. Innovations in trial design include better matching of treatment mechanism of
action to patient sub-populations and more stringent control on concomitant treatments
and sources of trial error. 951,952,954-958

These innovations in trial design require a deeper understanding by both researchers

and clinicians of their potential impact to support both continued approvals and to
promote the continued introduction of novel treatments for patient sub-populations who
may be underserved by currently approved treatments. The second point could impact
the clinical utility of new market entrants as the potential consequence of designing
approval programs for novel DED treatments that exclude patients with signs of DED
that don’t align with a drug/device mechanism of action may limit efficacy across the
spectrum of patients that present with DED. For example, limiting the inclusion of
patients with MGD in drug trials supporting currently approved products, knowing MGD
is present in 51.3% to 70.3% of patients with DED, 359,360, may partially explain the
apparent dissociation between the regulatory approval success of compounds in
enriched populations and the subsequent high dropout/discontinuation rates for
treatments in clinical practice. 953,959,960 Peer-reviewed literature reports discontinuation
rates up to 40% within the first year of a filled prescription while certain ophthalmological
practices can see estimated discontinuation rates over 60% with some anti-
inflammatory treatments. 959 Newer treatments and associated clinical trial designs with
enriched populations of patients will likely only expand as treatments tailored to newly
identified subpopulations (e.g., Ocular Neuropathic Pain) continue to be identified over
time. 205,953

In addition to searching the published, peer-reviewed literature for information on

existing methodology, detailed reviews on development programs and outcomes that
are publicly available were utilized as a primary source of information for this report. The
FDA website (accessdata.fda.gov) and ClinicalTrials.gov which is a publicly available
registry of clinical trials maintained by the US National Library of Medicine both list
publicly available information, including clinical protocols, statistical reports and
outcomes. The European Medicines Agency (EMA) also issues assessment reports on
products that were accepted or not approved (withdrawal assessment report) for use.
These can be found at https://www.ema.europa.eu/en/documents. The PMDA publishes
review reports on approved drugs and other products in English
(https://www.pmda.go.jp/english/review-services/reviews/approved-
information/0001.html). Beyond the major markets, global regulatory agencies generally
publish assessment reports on products submitted for their review which are also
readily available online (e.g., the Australian Therapeutic Good Administration (TGA)
published report on a lifitegrast ophthalmic preparation:
https://www.tga.gov.au/sites/default/files/auspar-lifitegrast-191107.pdf).

109
 11.2 Approvals for DED

The recommendations for trial design from TFOS were first published in the TFOS
DEWS report 961 and were followed by the TFOS DEWS II report in 2017. 953 A
cyclosporine ophthalmic emulsion 0.05% was approved in 2003 in the US and is
indicated for an increase in tear production in patients whose tear production is
presumed to be suppressed due to ocular inflammation associated with
keratoconjunctivitis sicca. Sodium hyaluronate 0.1% and Mini 0.1% and 0.3% were
approved in 1995 in Japan for conjunctival epithelial disorders including sicca syndrome.
The TFOS DEWS report described the approval period followed by interest in the
development of additional anti-inflammatory and immunomodulatory drugs for the
treatment of DED. This time was marked by the failure of numerous follow-on
compounds and exploration of how the better control of trial design, endpoints and
operational excellence could potentially lead to a greater success rate in DED trials.

Lifitegrast ophthalmic solution (5%) was approved in the US in 2016 and is indicated for
treating the signs and symptoms of DED in adults over the age of 17 years. In Japan a
sodium hyaluronate ophthalmic solution (0.3%), Diquafosol ophthalmic solution (3%),
and Rebamipide ophthalmic suspension (2%) were approved between 2010 and 2012
for conjunctival epithelial disorders including sicca syndrome (dry eye) and
subsequently just dry eye. The TFOS DEWS II report described the approval followed
by interest in exploration of how matching the mechanism of action of a proposed
therapy to a potentially responsive population in which the treatment is likely to
demonstrate efficacy; the inclusion of biomarkers and/or surrogate markers of disease;
and recognition of strong placebo/vehicle effects that if properly controlled could
potentially lead to greater success rates in DED trials. 953 These concepts were
captured in the TFOS DEWS II report, trials for novel compounds and the FDA’s
guidance document on trial design for DED; following which a period of successive
approvals for additional DED treatments was observed (Figure 4 and Table 13).

110
Figure 4: Approved treatments for Dry Eye Disease

111
Table 13. Approved products in major markets

Generic Name Brand Name Year Country MOA Approved Indication

Approved

Cyclosporine RESTASIS® 2003 USA calcineurin inhibitor Indicated to increase tear production in patients
ophthalmic emulsion, immunosuppressant whose tear production is presumed to be suppressed
0.05% due to ocular inflammation associated with
keratoconjunctivitis sicca.
TM
Lifitegrast XIIDRA 2016 USA lymphocyte Indicated for the treatment of the signs and symptoms
ophthalmic solution, function-associated of DED
5% antigen-1 (LFA-1)
antagonist

Cyclosporine CEQUA™ 2018 USA calcineurin inhibitor Indicated to increase tear production in patients with
ophthalmic solution, immunosuppressant keratoconjunctivitis sicca (dry eye)
0.09%

Loteprednol EYSUVIS™ 2020 USA corticosteroid Indicated for the short-term (up to two weeks)
etabonate treatment of the signs and symptoms of DED
ophthalmic
suspension, 0.25%

Varenicline solution, TYRVAYA™ 2021 USA Nicotinic Indicated for the treatment of the signs and symptoms
0.03 mg nasal spray acetylcholine of DED
receptor agonist

Perfluorohexyloctane MIEBO™ 2023 USA semifluorinated Indicated for the treatment of the signs and symptoms
ophthalmic solution alkane of DED

Cyclosporine VEVYE 2023 USA calcineurin inhibitor Indicated for the treatment of moderate to severe
ophthalmic solution, immunosuppressant DED (VEVIZYE in Europe)
0.1%

Cyclosporine IKERVIS 2023 EU calcineurin inhibitor Treatment of severe keratitis in adult patients with
ophthalmic emulsion, immunosuppressant DED, which has not improved despite treatment with
0.1% tear substitutes

112
 Purified sodium HYALEIN® 1995 Japan increasing tear film Indicated for the treatment of corneal and conjunctival
hyaluronate, 0.1% ophthalmic solution stability / corneal epithelial disorders (Keratoconjunctival epithelial
0.1% healing disorder resulting from the following diseases:
Intrinsic diseases such as Sjögren's syndrome,
Stevens-Johnson syndrome and sicca syndrome (dry
eye).
Extrinsic diseases caused by surgery, drugs, trauma,
contact lens wearing, etc.)

Purified sodium HYALEIN® 2010 Japan increasing tear film As above for 0.1% solution
hyaluronate, 0.3% ophthalmic solution stability / corneal
0.3% healing

Purified sodium HYALEIN® Mini 1995 Japan increasing tear film As above for 0.1% solution
hyaluronate, 0.1% ophthalmic solution stability / corneal
(preservative free) 0.1% healing

Purified sodium HYALEIN® Mini 1995 Japan increasing tear film As above for 0.1% solution
hyaluronate, 0.3% ophthalmic solution stability / corneal
(preservative free) 0.3% healing

Moistear/Diquafosol DIQUAS® 2010 Japan P2Y2 purinergic Treatment of DED

sodium ophthalmic receptor agonist
solution, 3.0% (6x
Daily)

Rebamipide MUCOSTA® 2012 Japan upregulates the Treatment of DED

ophthalmic gene expression of
suspension, 2.0% MUC1, MUC4, and
MUC16

Diquafosol sodium DIQUAS® LX 2022 Japan P2Y2 purinergic Treatment of DED

ophthalmic solution, receptor agonist
3.0% (TID)

Cyclosporine RESTASIS® 2010 India calcineurin inhibitor Not known

ophthalmic emulsion, immunosuppressant
0.05%

113
 Cyclosporine CEQUA® 2021 India calcineurin inhibitor Not known
ophthalmic solution, immunosuppressant
0.09%

Lifitegrast SECA® 2023 India lymphocyte function Not known

ophthalmic solution, associated antigen
5% 1(LFA-1) antagonist

Diquafosol sodium DIQUAS® 2017 China P2Y2 purinergic Indicated for DED diagnosed with corneal and
ophthalmic solution, receptor agonist conjunctival epithelial injury accompanied by tear
3.0% (6x Daily) abnormalities

Cyclosporine Zirun Cycloome® 2020 China calcineurin inhibitor Indicated to increase tear production in DED where
ophthalmic immunosuppressant tear production is presumed to be suppressed due to
nanoemulsion, ocular inflammation associated with
0.05% keratoconjunctivitis sicca

Diquafosol sodium RUNLIMING 2022 China P2Y2 purinergic Indicated for DED diagnosed with corneal and
ophthalmic solution, receptor agonist conjunctival epithelial injury accompanied by tear
3.0% (0.4mL:12mg) abnormalities

Varenicline solution, TYRVAYA™ 2024 China Nicotinic Indicated for the treatment of the signs and symptoms
0.03 mg nasal spray acetylcholine of DED
receptor agonist

Perfluorohexyloctane HENGQIN 2024 China Semifluorinated Indicated for the treatment of the signs and symptoms
alkane of DED

DED-dry eye disease; EU-European Union; lymphocyte LFA-1-function-associated antigen-1; TID-Three Times Daily; USA-United States of
America
Disclaimer- This is not necessarily a complete list of approved prescription medications approved in China and India

114
Globally, the availability of treatments for DED targeted at a sub-population of patients
who present predominately with signs of inflammation or its impact (e.g., hyperemia,
corneal staining, and conjunctival staining) and ADDE (e.g., decreased tear production)
speaks to a potential pathway for approval of additional products. 962-966

To date, compounds with immunomodulatory, anti-inflammatory, tear stimulatory and

tear conservation mechanisms of action have all targeted DED patients. Approvals for
these products in the major markets are generally followed by approvals in significant
rest of world markets. However, a number of these compounds have failed repeatedly in
clinical trials in other countries.

11.3 Defining a pathway toward approval

11.3.1 Solving industry-wide failure rates

Pfizer completed a review of its portfolio and determined that in the 2010’s the
pharmaceutical industry, in general, faced declining R&D success rates which reached
a low in 2016 when only 10% of Phase 2 trials succeeded while Phase 3 success rates
were only 30%. 198,967 By 2020, this had improved to a Phase 2 trial success rate of 53%
and a Phase 3 trial success rate of 80%. 198

Improvements in Phase II success rates were enabled by better understanding of

disease heterogeneity, which led to better selection of patients for clinical trials and
longitudinal sampling of mechanistic biomarkers (e.g., advanced imaging techniques
and blood-borne biopsies). 198,967

Three fundamental elements were identified to significantly improve trial success rates:
1) Exposure at the site of action (Pillar 1); 2) Binding to the pharmacological target
(Pillar 2); and 3) Expression of pharmacological activity from the site of action (Pillar 3).
Successful trials could achieve all 3 Pillars, achieve Pillars 1 and 3, or achieve Pillars 2
and 3. This was often combined with the inclusion of biomarkers and/or surrogate
markers of disease to achieve an early proof of mechanism and/or early signal of
efficacy. 198,967

11.3.2 Overcoming disease heterogeneity with targeted sub-populations

Ocular surface diseases including EDE, 953,961,968 ADDE, 953,961,968 MGD 237,969 and
Contact Lens Discomfort (CLD) 284,970 represent ocular surface disease heterogeneity
that has been accounted for in more recent DED trial designs by matching the
mechanism of action of a proposed therapy to a potentially responsive population in
which the treatment is likely to demonstrate efficacy. 82,951,954-956,962,963,965,966,971-979

Trials targeting patients presenting with ADDE and inflammation have consistently
linked decreased tear production with more severe disease that disrupts the ocular

115
surface, exemplified by ocular surface staining, through the concept of the lacrimal
functional unit. 951,954-957,962,963,965,966,971,972,979 The lacrimal functional unit is defined as
an integrated system comprising the lacrimal glands, ocular surface (cornea,
conjunctiva and meibomian glands), lids, and the sensory and motor nerves that
connect them. These trials have inclusion criteria and associated primary endpoints
focusing on increased ocular surface staining, decreased tear production, more severe
symptoms (e.g, VAS score ≥ 40 at screening and baseline and an OSDI score of ≥23,
and inflammation. 951,952,954-958,962,963,965,966,971,972,974,976,977,979-981 As a group, many of
these trials also excluded patients with signs of MGD, EDE or CLD. 951,955-957,962-966,971,972

The products targeting patients presenting with ADDE and inflammation can be broken
down into two main categories based upon their mechanism of action. The first group of
compounds includes immunomodulatory and anti-inflammatory agents 951,955-957,962-
966,971,972
and the second can be broadly categorized as tear stimulatory or conservation
agents for the various layers of the tear film (i.e., lipid layer, aqueous layer, and mucin
layer) (see 13X). 952,954,958,973,974,976,977,979-981 Studies involving three of these compounds
further restricted their inclusion/exclusion criteria and primary endpoints to closely
match their products’ proposed mechanism of action. 954,955,973,974,976,977,979

Table 14. Mechanism of action and approval pathway

Approved Drug, Global Regulatory Agencies Approve Drugs for DED Irrespective of Pathophysiological Subtype
Mechanism of Action
Signs of Evaporative DED Signs of Aqueous-Deficient/Inflammatory Symptoms of DED
DED
Meibomi Meibum TBUT Corneal Conjunctival Schirmer Eye Ocular Total
an Gland Quality Staining Hyperemia Responder Dryness Discomf OSDI
Obstruct VAS ort VAS
ion Scale Scale
Meibomian Gland
Dysfunction &/or
Evaporative DED
None
Aqueous-Deficient &/or
Inflammatory DED
Calcineurin inhibitor IKERVIS RESTASIS®, IKERVIS
immunosuppressant CEQUA™,
VEVYE
lymphocyte function- XIIDRATM XIIDRATM
associated antigen-1
(LFA-1) antagonist
corticosteroid EYSUVIS™ EYSUVIS™
Tear
Replacement/Stabilser
(Non-Imunomodulatory)
Cholinergic agonist TYRVAYA™
Semifluorinated alkane MIEBO™ MIEBO™
Water retention and Sodium
binding to fibronection HyaluronateƗ
possibly promoting
adhesion and elongation
of corneal epithelial cells
P2Y2 purinergic receptor DIQUAS®,
agonist DIQUAS® LX
Quinolinone derivative MUCOSTA®
Ɨ HYALEIN® ophthalmic solution 0.1%; HYALEIN® ophthalmic solution 0.3%; HYALEIN® Mini ophthalmic solution 0.1%; HYALEIN® Mini ophthalmic solution 0.3%

Studies using a semifluorinated alkane to target the lipid layer included patients
presenting with ADDE, inflammation and MGD defined as a total MGD score ≥3 [range
0-15] at screening and baseline (secretion of 5 central glands on the lower eyelid was

116
evaluated, and each was scored from 0-3: 0=normal; 1=thick/yellow, whitish, particulate;
2=paste; 3=none/occluded) and a TBUT ≤5 seconds at screening and baseline. Thus,
the inclusion criteria were tailored toward the mechanism of action of the compound
even though the primary endpoints were consistent with patients presenting with ADDE
and inflammation. 973,974,976,977

Studies involving a cholinergic agonist additionally targeted patients presenting with a

baseline Schirmer score response to cotton swab nasal stimulation of at least 7 mm
greater in the same eye that qualified as ADDE. Thus, patients with a marked response
to mechanical activation of the trigeminal parasympathetic pathway in the nose were
subsequently tested to determine if nicotinic acetylcholine (nACh) receptor activation of
the same pathway results in increase basal tear production. 954,979

A new topical corticosteroid additionally targeted patients presenting with bulbar

conjunctival hyperemia at Screening and Day 1 of ≥ 2 as assessed using the Cornea
and Contact Lens Research Unit Grading Scale. Corticosteroids cause adrenergically
mediated vasoconstriction and non-competitive antagonism of vasodilation due to
prostaglandin E and bradykinin. 982 Thus, targeting patients presenting with aqueous-
deficient DED, inflammation and hyperemia would be consistent with a mechanism of
action for a corticosteroid to reduce dilation of conjunctival blood vessels which can
present as hyperemia associated with inflammation (see Table 15). 955,983

117
Table 15. Approved drugs and criteria for defining DED
Criterion for Defining DED
Signs Symptoms
Approved Drug

Corneal Staining
Meibum Quality

Ocular Dryness
Eye Redness
Studies

Conjunctival

Schiermer's

NEI-VFQ-25
Expression
Meibomian
Functional
Number of

Staining

SANDE
Glands

SPEED

Facial
Score
TBUT

Scale
OSDI

VAS
192731 -002, Sum of corneal and conj. Staining ≥ +5 in (without anaesthesia) ≤ 5 At least 9 Score ≥ 3
-003, and - the same eye with corneal staining ≥ +2 mm/5min, if 0 mm/5min responses
501 (Oxford scheme; 0-15) then nasal stimulation ≥ on the OSDI
RESTASIS®

3 mm/5 min other than

N/A and a
minimum
score at
screening
and baseline

Phase 2 and Active lid margin disease Corneal fluorescein (without anaesthesia) ≥ 1 Conjunctiveal ODS ≥ +3
Opus-1 not included staining score ≥ 2 in and ≤ 10 mm/5min redness score at 2
at least one region ≥ 1 (0-4 scale) consecutive
of either eye (0-4 in at least one time points
scale) at Screening eye (any eye intra-CAE
and Baseline; for OPUS-1)
Change in Inferior
Corneal Staining
(pre to post CAE ≥
XIIDRATM

+1)
Opus-2 and Active lid margin disease Corneal fluorescein (without anaesthesia) ≥ 1 Conjunctiveal Eye Dryness
Opus-3 not included staining score ≥ 2 in and ≤ 10 mm/5min redness score Score ≥ 40 at
at least one region ≥ 1 (0-4 scale) Screening and
of either eye (0-4 in at least one Baseline
scale) at Screening eye
and Baseline;
Inferior Corneal
Staining ≥ 0.5 at
screening and
baseline)

Significant eyelid Lissamine green Global symptom

irregularity not included conjunctival score (based on
staining sum symptoms of
CEQUA™

score of 3 or dryness, irritation,

more to 9 or or both) rated by
less of a total the patient of 40 or
possible score more (range, 0-
of 12 in the 100)
same eye

Meibomian gland Corneal fluorescein Uanesthetized Schirmer Bulbar A score of >

dysfunction (MGD) not staining score at Test score at Screening conjunctival 50 mm on
included Screening and Day of ≤ 10 mm hyperemia at Modified
1 of ≥ 6 NEI Screening and SANDE
(National Eye Day 1 of ≥ 2 Severity at
Institute) as assessed Screening;
EYSUVIS™

using the > 40 mm on

Cornea and Modified
Contact Lens SANDE
Research Unit Severity on
(CCLRU) the day
scale prior to Day
1

118
 Onset-1 Blepharitis not requiring Corneal fluorescein baseline Schirmer’s Test
(OPP-002) treatment and mild staining score of ≥2 Score (STS; with topical
Meibomian gland disease in at least one anesthesia) of ≤10 mm/5
that are typically corneal region OR minutes with a cotton
associated with DED have a sum of ≥4 swab nasal stimulation
were allowed. for all corneal STS at least 7 mm
regions greater in the same eye;
<20 mm difference from
the study eye Schirmer
TYRVAYA™

Test and the fellow eye

Schirmer Test Score
Onset-2 Blepharitis not requiring Corneal fluorescein baseline Schirmer Test OSDI score
(OPP-101) treatment and mild staining score of ≥2 Score (STS; with topical of ≥23 with
Meibomian gland disease in at least one anesthesia) of ≤10 mm/5 ≤3
that are typically corneal region OR a minutes with a cotton responses of
associated with DED sum of ≥4 for all swab nasal stimulation “Not
were allowed. corneal regions STS at least 7 mm Applicable”)
greater in the same eye; at the
<20 mm difference from Screening
the study eye Schirmer Visit
Test and the fellow eye
Schirmer's Test
GOBI MGD defined as total TFBUT ≤5 Total corneal unanesthetized Schirmer OSDI score
MOJAVE MGD score ≥3 at seconds at fluorescein staining test I score ≥5 mm at ≥25 at
Screening and Beseline Screening score between 4 Screening and Baseline Screening
(secretion of 5 central and and 11 (i.e., sum of and
glands on Baseline inferior, superior, Baseline.
the lower eyelid was central, nasal, and
evaluated, and each was temporal) according
MIEBO™

scored from 0-3: to the NEI scale at

0=normal; 1=thick/yellow, Screening and
whitish, particulate; Baseline
2=paste;
3=none/occluded). Total
score ranged from 0-15

CYS-002 Meibomian gland Total corneal Total lissamine Schirmer Test I score Score of ≥ 40 on
dysfunction (MGD) not fluorescein staining green between ≥ 2 mm and ≤ 8 the dryness VAS
included score of ≥ 6 (e.g. conjunctival mm at Screening and at Screening and
sum of inferior, score (sum of Baseline Baseline
superior, central, temporal and
nasal, and nasal) of ≥ 2,
temporal) according based on the
VEVYE

to the NEI grading Oxford grading

at Screening and at Screening
Baseline and Baseline

119
 ESSENCE -1 Total corneal Total lissamine Schirmer Test I score Subjective
(CYS-003) fluorescein staining green between ≥ 1 mm and ≤ complaints of
score of ≥ 10 (e.g. conjunctival 10 mm at Screening and poor near vision
sum of inferior, score (sum of Baseline that impact
superior, central, temporal and activities of daily
nasal, and nasal) of ≥ 2, living, as defined
temporal) according based on the by at least a
to the NEI grading Oxford grading moderate impact
at Screening and at Screening (score ≥ 3) on at
Baseline and Baseline least 1 question
on the NEI VFQ-
25 Questions 5
to 7 in the main
questionnaire or
Near Vision
Subscale,
Questions A3 to
A5 in the
Appendix of
Optional
Additional
Questions, at the
screening visit

ESSENCE 2 Meibomian gland Total corneal Total lissamine Schirmer Test I score Score of ≥ 50 on
(CYS-004) dysfunction (MGD) not fluorescein staining green between ≥ 1 mm and ≤ the dryness VAS
included score of ≥ 10 (e.g. conjunctival 10 mm at Screening and at Screening and
sum of inferior, score (sum of Baseline Baseline
superior, central, temporal and
nasal, and nasal) of ≥ 2,
temporal) according based on the
to the NEI grading Oxford grading
at Screening and at Screening
Baseline and Baseline
Siccanove Severe blepharitis and/or Tear break- Corneal fluorescein Lissamine green Schirmer tear test OSDI score NOT VAS: At least
(NVG06C103) Meibomian gland disease up time staining ≥2 and ≤4 staining >4 (Van without anaesthesia of ≥23 one moderate to
(MGD) not included (TBUT) ≤8 (modified Oxford Bijsterveld ≥2 mm/5 min and <10 severe symptom of
seconds scale, scale 0-5) scale, scale 0-9) mm/5 min dry eye with a
score ≥2 (severity
graded on a 4-
point scale) i.e.,
burning/stinging,
foreign body
sensation, itching,
IKERVIS

eye dryness, pain,

blurred vision or
sticky feeling and
photophobia
Sansika Severe blepharitis and/or Corneal fluorescein Schirmer test without OSDI score
(NVG10E117) Meibomian gland disease staining score of 4 anaesthesia scored ≥2 ≥23
(MGD) not included on the modified mm/5 min and
Oxford scale

N/A not listed not listed Corneal fluorescein not listed not listed not listed
HYALEIN®
ophthalmi

staining score
c solution

≧2+(0~3?)
0.1%

"moderate or
above"

N/A not listed not listed not listed not listed not listed not listed
"refractory or
solution 0.3%

severe"
ophthalmic
HYALEIN®

N/A not listed not listed not listed not listed not listed not listed
m

%
M
®
H

N
Y

E
L

o
p
h

u
ti
o
n
a

c
s

"moderate or
t
I

120
 above"
N/A not listed not listed not listed not listed not listed not listed
hal

sol

0.3
HY
AL

op

uti
on
mi
Mi

"refractory"
ht
EI

%
ni
®
N

≤5 seconds Corneal fluorescein staining score: ≥3 (a Schirmer I test: ≤5 mm Subjective symptoms (including visual disturbance)
maximum score of 9); Rose Bengal
staining score: ≥3 (a maximum score of
DIQUAS®

9); Lissamine green staining score: ≥3 (a

maximum score of 9) [Staining of the
temporal conjunctiva, cornea, and nasal
conjunctiva, with each graded on a 0-3
scale. The scores of the three regions are
summed.]

A corneal fluorescein staining score of 4 Schirmer I test: ≤5 mm Score of 2 or more for 1 or more dry eye–related ocular symptom(s)
MUC

or more and a LGCS score of 5 or more

OST
A®

≤5 seconds Corneal fluorescein staining score ≥ 1 Schirmer I test: ≤5 mm Dryness score ≥ 1 using the DEQS questionnaire
DIQ
UA
S®
LX

OSDI = Ocular Surface Disease Index

121
Obstructive MGD presents as lid telangiectasia, gland orifice capping, gland dropout,
altered gland expressibility, and low TBUT). 6,984,985 This distinction from ADDE is
important as newer products in development like selenium sulfide ophthalmic ointment
975,978
and devices for treating eyelid glandular abnormalities, 986 have inclusion criteria
and associated primary endpoints focusing on glandular morphology and associated
evaporative DED. 987-993 Such trials may exclude patients with more severe symptoms,
signs of inflammation and significant ocular surface staining. 975 The inclusion/exclusion
criteria and endpoints were modelled on the clinical studies performed using a thermal
pulsation system. 994 These trial designs targeted at obstructive MGD and associated
EDE illustrate a population-targeted trial design aimed toward a group of patients who
may be underserved by some of the existing approved drug treatments.

11.3.3 US FDA clear guidance for industry and flexibility for dry eye approvals
The US FDA Draft Guidance for Industry on Dry Eye: Developing Drugs for Treatment
949
recommends the sponsor of a new DED treatment demonstrate efficacy and safety
in at least two adequate and well-controlled, multicenter independent studies. The
guidance covers trial design, comparator(s), trial population, demonstration of efficacy
and safety database requirements in detail. The following section will summarize the
key guidelines issued by the FDA. 949

11.3.3.1 Trial design

The FDA will consider both traditional environmental exposure trials and an
environmental challenge-model trial (utilizing a controlled chamber with regulated
temperature, air flow, humidity, etc.) to support approval of a new drug. The FDA
recommends parallel, randomized by patient, double-masked trials in which the
investigational drug group demonstrates superiority over the control group (control
agent can be the vehicle). Equivalence and non-inferiority trials are generally
discouraged due to difficulty in defining limits for equivalence and cross-over trials will
only be evaluable for the first treatment period if any carryover from one period to
another is detected. Future work to establish good assay validation (sensitivity) methods
(inclusion of both a positive and negative concurrent control), could change the
recommendation on equivalence or noninferiority trials.

The FDA guidelines are applicable to trials designed to demonstrate the efficacy and
safety of a new product in support of a new drug application and do not preclude the
use of such trial designs by companies or sponsors for the purposes of supporting
internal decisions related to a compound’s ability to progress in development based
upon measures of safety or efficacy such and proof of mechanism endpoints,
biomarkers and alternative thresholds for significance (e.g., 0.1% not 0.05%).

122
Daily exposure to environmental factors can impact DED endpoints: cigarette smoke,
smoke from wildfires, air pollution, dust, topically instilled drugs, and allergens are just a
few examples. 995-997 A model has been developed to standardize climatic, air quality
and visual conditions in a controlled environment. 996,998,999 Aspects of the environment
such as humidity, air flow and temperature can be manipulated to impact the ocular
surface, to identify individuals who may be more responsive to external environmental
factors and to create a reproducible environmental challenge. 999 This model is an
attempt to reduce the number of patients, sites, and time to demonstrate the efficacy of
an intervention. 996,998,999 Effects can be tested over one day in a controlled environment
or longer-term natural exposure trials of 2-weeks or longer. The FDA recommends that
safety trials be conducted at least 6 weeks in duration if efficacy trials are of shorter
duration. Trials in which the investigational drug is used as an add-on to a standardized
treatment regimen are also acceptable. 949

While confirmatory efficacy trials have used CAE exposure to enrich study populations
with patients who can change in severity secondary to environmental stress, the pre-
specified primary endpoints supporting approval were environmental. 964 Trials have
also used the controlled adverse environment to test an intervention’s protection against
the effects of an environmental challenge. 1000

11.3.3.2 Comparator(s)
The comparators and methods for controlling variance have evolved from the first
studies of cyclosporine in DED, 951 to the recent introduction of newer formulations and
concentrations of the active. 951,956,957,971,972 The first studies of cyclosporine in DED
evaluated cyclosporine plus artificial tears as needed to Month 4 and then less than 8
times daily after Month 4 to Month 6. 963 Numerous trials tried to replicate this
methodology and failed due to large placebo effects, lack of efficacy and regression of
signal which could have resulted from a therapeutic effect for the artificial tears. 963,1001
The subsequent approval of a novel integrin antagonist (lifitegrast) used only the vehicle
and excluded the use of artificial tears during the treatment period (See Supplementary
Figure 1 for the 0.05% ciclosporine trial design, Figure 2 for the lifitegrast controlled
environmental trial design, and Figure 3 for a lifitegrast environmental exposure trial
design).

Supplementary Figure 1: Cyclosporine 0.05% (192371-002: Phase 3):

Environmental Exposure Study Design

Supplementary Figure 2: Opus 1 (Lifitegrast: Phase 3): CAE Study Design

Supplementary Figure 3: Opus 2 (Lifitegrast: Phase 3): Environmental Exposure

Study Design

123
The FDA acknowledged the advancement in trial methodology and encouraged trial
designers to consider that even water is known to be an effective component of topically
applied treatments for DED. Vehicle responders may be 1 in 5 of the trial population
suggesting this is a significant confounder in a study without a mechanism to screen for
vehicle response. 1002 In general, comparative clinical trials should use only the
investigational drug’s vehicle as a control agent. Trials should demonstrate statistical
and clinical superiority over a vehicle control or another treatment regimen. 949

11.3.3.3 Trial population

The FDA defines a DED population as including patients with ocular complaints
consistent with dry eye symptoms. Inclusion criteria should include both objective signs
and subjective symptoms. 949 Examples of inclusion and exclusion criteria for approved
products are included in Table 14. The FDA includes measures of meibomian gland
function, meibum quality and TBUT as both a potential inclusion criterion for a DED
population and a potential sign of DED therapeutic effect which can function as a
primary sign efficacy endpoint. 975,978 Studies should include patients from relevant
demographic subsets, including both men and women and multiple age, ethnic
background, and eye color groups consistent with the US population. Confirmatory
efficacy studies do not need to be completed in the US so long as the study population
contains the relevant demographic subsets found within the US.

DED secondary to cicatrisation (following irradiation, alkali burns, Stevens Johnson

syndrome, cicatricial pemphigoid) or the destruction of conjunctival goblet cells (vitamin
A deficiency) represent a specific, severely affected patient population. In general, these
are considered separate indications, and patients with these conditions should be
studied separately from routine DED.

Severe blepharitis or obvious inflammation of the lid margin can interfere with the
interpretation of trial results. In general, patients with these conditions should be studied
separately from routine DED. 949

11.3.3.4 Demonstration of efficacy

In general, safety and efficacy should be demonstrated in at least two adequate and
well-controlled, multicenter (at least 2 sites), independent trials. Primary efficacy
endpoints include one of three options:

Sign and Symptom endpoint (s): 949,950

1. Signs and Symptoms - A statistically significant difference between the

investigational treatment and vehicle from baseline for at least one objective
prespecified sign of DED (mean group score of test versus vehicle) AND at least one
subjective prespecified symptom of DED (mean group score)

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2. Corneal Staining - A statistically significant difference between the percentage of
patients achieving a complete resolution of corneal staining, or
3. Tear Production - A statistically significant difference between the percentage of
patients achieving a 10-millimeter increase or more in Schirmer score

Achieving any one of these 3 endpoints can support approval of a product for the
treatment of the signs and symptoms of DED. Several marketed products have sought
approval based upon sign and symptom primary endpoint(s) and have been granted US
approval using pre-specified secondary endpoints of a known surrogate endpoint, a 10-
millimeter increase or more in Schirmer score, when the prespecified signs and
symptom endpoints failed in confirmatory efficacy studies. 951,956,957,963,971,972

The “10-millimeter increase or more in Schirmer score” was validated as part of the new
drug application for 0.05% ciclosporine and a description of the methods used to
validate the endpoint are described in the “Validation of the Clinical Relevance of the
Clinical Sign” section authored by Dr. William M Boyd from the US FDA as one of the
medical reviewers for NDA 21-023.

11.3.3.5 Safety database

The FDA specifies the size of a drug exposure database to support a new drug
application based upon Adverse Event detection rates. The clinical program should
include enough patients to identify adverse drug events that occur at a rate of 1 percent
or greater. To accomplish this, FDA recommends that approximately 400 or more
patients using the investigational drug complete treatment with a concentration of the
investigational drug at least as high as proposed for marketing and with a frequency at
least as frequent as proposed for marketing. Before submission of a marketing
application, the sponsor should ensure at least 300 patients have completed at least 6
weeks of follow-up after the initiation of treatment and at least 100 patients have
completed 12 months of follow-up after the initiation of treatment to support chronic use.
Acute use, 6 weeks or less of administration each year, does not require long-term
follow-up of 12 months.

For reformulations of drug substances that are already approved in the same dosage
form, same route of administration, and the same or lower concentration, the FDA
recommends the sponsor ensure that a marketing application has safety information
from at least 100 patients treated for at least 6 months.

FDA recommends that the following evaluations be performed in each eye and reported
separately for each eye (regardless of which eye or eyes are treated):

 Best corrected, distance visual acuity (4 meters in distance or more) at every visit.
 A patient comfort examination before and after drug administration at every visit.

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 A slit lamp examination of the anterior segment that includes the cornea,
conjunctiva, anterior chamber, iris, lids, and lashes. At a minimum, examinations
should be performed at baseline, midway through the trial, the end of treatment, and
2 weeks after treatment discontinuation.
 Endothelial cell count, systemic clinical and laboratory evaluations, and dilated
fundus examinations at baseline and at the end of trial or at month 3 (whichever is
later) in at least one trial.

DED also occurs in pediatric patients. 1003 Sponsors may consider a pediatric
assessment waiver request when submitting their required pediatric study plans under
the Pediatric Research Equity Act. 949

11.3.4 EMA guidance for industry and flexibility for DED approvals
EMA guidelines recommend demonstration of efficacy for 6 months, as DED is often
chronic and comparison against artificial tears, which are the standard of care is
recommended. 1004 EMA application submissions for lifitegrast and cyclosporine were
both withdrawn because of data demonstrating insufficient efficacy. 953,1005 The only
EMA-approved treatment is cyclosporine 0.1%, however, the indication is not for DED
but rather “severe keratitis in patients with DED, not improved despite treatment with
tear substitutes.” 1006

Regarding the selection of sign endpoints, the EMA considers both the target population
and the mechanism of action of the compound: Corneal staining, Schirmer score and
TBUT are established clinical endpoints. A composite measure is recommended using a
validated questionnaire for a symptom endpoint (e.g., OSDI) and use of single worst
symptom questions is not recommended. Sponsors are encouraged to consider a
relevant effect size rather than only statistical significance. For anti-inflammatory
products, since inflammation is considered a secondary manifestation, biomarkers may
be required to quantify treatment benefit.

The EMA will generally accept development programs completed in the US assuming
the studies comply with the guidelines outlined above. The EMA only requires
demonstration of safety and efficacy in one large adequately designed Phase 3 trial.
The primary endpoint for a cyclosporine 0.1% Phase 3 trial was a sign and symptom
composite responder rate. Specifically, the primary endpoint was the corneal fluorescein
staining-OSDI composite responder rate at Month 6 (i.e. end of Part 1). A corneal
fluorescein staining-OSDI responder was defined as a patient simultaneously satisfying
the following conditions: 1006

 Improvement of 2 points or more from baseline in corneal fluorescein staining based

on the modified Oxford scale (i.e. change in corneal fluorescein staining ≤-2), and

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 Improvement by 30% or more from baseline in OSDI (i.e. % change ≤-30%).

This is not an endpoint which has been acceptable to the FDA to date. The completed
confirmatory efficacy trials (SICCANOVE and SANSIKA) failed to demonstrate
consistent superiority on the primary endpoint and the product was subsequently
approved on a secondary sign endpoint by showing a statistically significant reduction in
corneal fluorescein staining versus vehicle. 951,1007,1008 No significant improvement in
symptoms was demonstrated along with the observed significant and clinically relevant
improvement in corneal staining and the indication was restricted to “severe keratitis
treatment” instead of a broader DED population.

11.3.5 PMDA guidance for industry and flexibility for approvals

Historically, there has been a considerable time delay between development and
approval in the United States and an ultimate launch in Japan. 1009 The Pharmaceutical
and Medical Device Agency (PMDA) requirements for drug approval in Japan have
been harmonized across regions with toxicology and manufacturing generally not
requiring additional work or rework. 1009

Working with the PMDA involves a multistep process. This process is initiated with the
jizen mendan (“preliminary meeting”) which has no fee associated with it, is shorter in
length and requires less of a briefing package. The intention of this meeting is to confirm
the materials that are to be submitted and the sponsor’s questions that will be discussed
at the subsequent taimen jogen (“full consultation meeting”). At the full consultation
meeting, a more detailed briefing document is submitted in advance and written
comments are provided by the PMDA following the meeting.

The PMDA views ethnic differences as critically important. Phase 1/pharmacokinetics

studies can be completed in the United States, if done in Japanese-American patients
defined as first- or second-generation Japanese Americans of pure Japanese descent.
PMDA generally requires a Phase 1/pharmacokinetics study even in cases where a
global development program may have proceeded into Phase 2 or 3 trials in other
regions, but with a non-Japanese population. Recently the PMDA issued a notice
regarding the basic approach to conducting Phase 1 studies in Japanese patients prior
to global clinical trials for drugs whose clinical development has already started
overseas 1010,1011, stating the following. “In general, it is not mandatory to conduct a
Phase I study in each race/ethnicity or country/region before initiating multi-region
clinical trials. In principle, an additional Phase I study in Japanese is not needed unless
it is deemed necessary after assessing whether the safety/tolerability of the dosage to
be evaluated in the multi-region clinical trials in Japanese participants can be explained
and the safety in clinically acceptable/manageable on the data available prior to Japan’s
participation.” This concept seems to be some progress in the development of

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pharmaceuticals in Japan. The PMDA typically wants to see dose ranging established in
the Japanese population specifically because of potential population differences in drug
activity and practice patterns. Confirmatory Phase 3 study is required to be completed in
Japan in part or whole. The PMDA generally requires a single, properly designed Phase
3 clinical trial to demonstrate safety and efficacy, similar to the EMA and discourages
post-hoc analysis for approval.

Phase 3 trials for the approval of diquafosol sodium ophthalmic solution 952,1012-1014 and
rebamipide ophthalmic suspension 952,1015 both required an active comparator (0.1%
hyaluronic acid ophthalmic solution was used in both) which necessitated a dosing
regimen of 6 times daily. This may have hampered interest in developing product in
Japan due the risk posed by even vehicle effects at such a high dosing frequency. The
introduction of long acting diquafasol sodium ophthalmic solution in 2022 which is only
dosed 3 times daily may have changed this risk profile. Long acting diquafasol also
employed a vehicle placebo in its Phase 3 trial. 1016

All 3 products were tested with primary endpoints at 4 weeks and all measured primary
treatment benefit using staining endpoints:

1. Diquafosol sodium ophthalmic solution: A non-inferiority margin of 0.34 for the study
(between-treatment difference in the mean change in the fluorescein & rose bengal
staining score [diquafosol - HA])
2. Rebamipide ophthalmic suspension: Change from baseline in corneal fluorescein
staining score and the lissamine green conjunctival staining score
3. Long acting diquafosol sodium ophthalmic solution: Change in corneal fluorescein
staining score from baseline to week 4

Similar to those products that were approved in the US targeting a sub-population of

DED patients with ADDE and inflammation, the Phase 3 trials for these products
targeted patients with evidence of ocular surface staining, decreased tear production
and dry eye symptom(s). 952,958,1012-1017 Diquafosol sodium ophthalmic solution and long
acting diquafosol sodium ophthalmic solution also required a TBUT ≤5 seconds which
was also required as part of the development program for perfluorohexyloctane
ophthalmic solution. 974,976,977 In these trials, diquafosol sodium ophthalmic solution and
long acting diquafosol sodium ophthalmic solution were reported to promote the
secretion of tear fluid with aqueous and mucin components and as well as to increase
lipid layer thickness in normal human eyes. 1018 Thus, the Phase 3 trials were enriched
based upon the known mechanism of action of a P2Y2 purinergic receptor agonist. It
should be noted that a P2Y2 agonist and rebamipide failed in multiple clinical trials in
the USA to assess their efficacy in treating DED. 952,958,1012-1018

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The primary endpoint for a hyaluronic acid Phase 3 study was a sign and symptom
composite responder rate. 980 This is not an endpoint which has been acceptable to the
FDA to date. The diquafosol ophthalmic solution Phase 3 also defined a non-inferiority
margin between-treatment difference in the mean change in the fluorescein and rose
bengal staining score (diquafosol compared to hyaluronic acid). 1012,1014 The FDA has
not accepted a non-inferiority margin to support approval of a drug for DED. Thus, the
PMDA has allowed flexibility in defining the type of comparison that will support
approval: superiority or non-inferiority to an active comparator.

The PMDA does not have the same guidance for industry on developing drugs for DED
as the FDA. In addition, the diagnostic criteria for DED were revised in 2016 1019 and no
longer include tear production or keratoconjunctival disorders in the diagnostic criteria.
Two of three approved drugs for the treatment of dry eye, diquafosol ophthalmic
solution 958,1012-1014 and rebamipide ophthalmic suspension 952,1015 were approved before
the revision and the remaining drug, long acting diquafosol sodium ophthalmic solution
1016,1017
had an additional additive that reduced the frequency of eye drops, but the
efficacy itself did not change, so the primary endpoint in trial was the fluorescein corneal
staining score. With the revision of diagnostic criteria (ie., tear production and
keratoconjunctival disorders are no longer included), it may be necessary to have a
robust discussion with regulatory authorities regarding the setting of the primary
endpoint based on the mechanism of action of a new drug.

11.4 Devices

11.4.1 Regulatory pathway (FDA)

The regulatory process for devices is very different from the process for
pharmaceuticals. 953,1020This section briefly describes the process utilized by the FDA,
but there are likely differences in other regions of the world. 953 Similar to its role in the
approval of drugs, the FDA is responsible for protecting the public by assuring the
safety and efficacy of medical devices. Although both drug and device approval have
increased over the last decade, there is an increasing gap favoring new devices in the
USA. 1020-1022 In general, devices have a shorter time for bringing to market and
significantly less cost but may have lower profitability. 1021 The key steps to FDA device
approval start with determining if a product is a device and identify its purpose, and then
to determine its risk classification: low, moderate or high risk (Figure 5).

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Figure 5. Determining the Regulatory Pathway for a Device (from Van Norman
2016). 1021

ProvRisk is assessed to determine the device chance of presenting harm to patients,

including from malfunction or improper use. The risk classification generally indicates
the pathway and type of submission required by the FDA. The type of premarket
submission depends on the risk assessment:
1. Investigational device exemption - devices used to collect safety and
effectiveness data
2. Premarket notification (510k) – the device is safe and effective and substantially
equivalent to a legally marketed device
3. Premarket approval application - for Class III device (high risk) that needs a
clinical trial to demonstrate safety and effectiveness
4. De novo - for devices that have no existing classification regulation
5. Humanitarian device exemption – for devices for a disease that affects less than
4000 US patients yearly and has demonstrated reasonable assurance of safety
and probable benefit
6. Low risk (Class I devices) (e.g., the Meibomian Gland Evaluator, marketed by
Johnson & Johnson Surgical Vision, Inc.) and moderate (Class II devices) (e.g.,
iLux® System by Tear Film Innovations, Inc. and the LipiFlow Thermal Pulsation
System manufactured by TearScience, Inc.) risk devices can be “exempt” or
require a 510(k), while the highest risk (Class III) device requires a pre-market

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 approval. Class III risk devices will ultimately require a clinical trial and an
Investigational Device Exemption so that an unapproved device can be used in
clinical trials.

Trial design for Class III devices: Given that it is often difficult or unethical to have a
sham control, there is more flexibility on what type of trial is needed and discussion with
the FDA will determine what is acceptable. 1021 For the control arm of a device trial, one
may be able to use an active comparator (i.e., compare with an FDA approved device),
use metrics from pre- and post-treatment with a given device, or compare with registry
data. Typically, a double blind randomized clinical trial cannot be done with devices, as
is done with pharmaceuticals. 1022 Mixed devices, pharmaceutical and device combined,
are often reviewed by both divisions of the FDA, with one taking the lead, where
communications with the FDA determine the regulatory pathway. 953

11.4.2 Regulatory pathway (Japan, PMDA)

In Japan, as in the US, the development and approval processes for medical devices
differ depending on the type of device. Medical devices are classified into four classes
according to the degree of risk to the human body. Medical devices in Class I require
only notification, those (except new devices*) in Class II require certification by the
certification body, and those in Class III and IV require approval by government
authority. 953,1023

Medical devices classified as Class III and IV are considered to have a high risk to the
human body and many of them require clinical trials. On the other hand, medical
devices used outside the body that are not implanted in the body and do not have a
high risk to the human body are often classified as Class II. With Class II, if the
probability of benefit is high when considering risk and benefit, it may be judged that
evaluation is possible without the need for clinical trials, as compared to Class III and IV.
Class I medical devices are those that are considered to have an extremely low risk to
the human body.

For medical devices that have already been approved overseas, it is possible to
extrapolate test results from overseas clinical trials, although ethnic differences must be
taken into consideration. Therefore, unlike drugs, some medical devices are approved
without domestic clinical trials by extrapolation to overseas clinical trials.

*New medical devices in Class II require approval by government authority (PMDA

and MHLW: Ministry of Health, Labour and Welfare).

11.4.3 Artificial tears (US, FDA)

Artificial tears contain FDA-approved demulcents as active ingredient(s). Demulcents
are primarily water-soluble, topically applied polymers, which may protect and lubricate

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mucous membrane surfaces relieving ocular dryness and irritation. 953,1021 A list of
1024
approved demulcents and their concentration range has been provided by the FDA:

1. Cellulose derivatives:
a. Carboxymethyl cellulose (CMC),
b. Hydroxypropyl methylcellulose (hypromellose),
c. Hydroxyethylcellulose,
d. Methylcellulose,
2. Dextran 70
3. Gelatin
4. Polyols:
a. Glycerin,
b. Polyethylene glycol (300, 400),
c. Polysorbate 80,
5. Polymers:
a. Polyvinyl alcohol,
b. Polyvinyl pyrrolidone (povidone).

Federal law does not require premarket approval for over-the-counter eye drops,
including artificial tears, but does require eye drops to be sterile for safe use. The
existence of the FDA’s pre-approved monograph which covers artificial tears means
that if a developer lists an active ingredient that is not on the monograph, they will need
to provide clinical data supporting the safety of its use. Manufacturers can list potentially
“active ingredients” under “inactive ingredients” to avoid clinical trials.

While clinical data demonstrate that artificial tears show benefit for the management of
DED and their control is critical during drug/device trials to remove a confounding
source of variance, there are no regulatory standards for study design, clinical end
points, patient inclusion/exclusion criteria, or accepted methods to control for potential
sponsor bias when evaluating comparative efficacy claims. 1024

11.4.4 Artificial tears (Japan, PMDA)

Most artificial tears in Japan are classified as OTC drugs that do not require a
prescription. Over-the-counter drugs require clinical trial results if they contain
ingredients that are not included as active ingredients in any of the drugs specified in
the Japanese Pharmacopoeia, but do not require clinical trials if they do. 953,1021

11.4.5 Devices approved for DED/MGD

Since the first TFOS DEWS report, there have been several devices FDA approved
targeting meibomian gland secretion or mechanical tear stimulation. 993 The first thermal
pulsation device 990,991 was approved by the FDA for the application of localized heat
and pressure therapy in adults with chronic cystic conditions of the eyelids, including
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MGD. It was approved after it met the primary study effectiveness endpoint of
improvement at 2 weeks from baseline in the average number of meibomian glands
yielding clear liquid secretion as compared to a warm compress control. 69 subjects
(138 eyes) were randomized to the thermal pulsation treatment and 70 subjects (140
eyes) were randomized to the warm compress control group. The same system is also
a medical device approved without clinical trials in Japan.

Subsequent thermal pulsation devices could be approved through a single non-

inferiority clinical trial that compared the device to the approved system as the predicate
device. One example demonstrated non-inferiority to the original system using the
primary effectiveness endpoints defined as the change from baseline to 1 month for
TBUT and total Meibomian Gland Secretion Score. 993 There are numerous examples of
other devices, including those using intense pulsed light, that have been evaluated as
treatments of evaporative DED or MGD. 987-989

The Warming Moist Chamber Goggle (Dr.eye®), an NMPA-approved therapeutic device,

is indicated for the management of DED and MGD. It facilitates localized thermal and
hygrometric conditions, which may enhance tear film stability. Empirical evidence 1025
has demonstrated that this device alleviates ocular discomfort and improves tear film
parameters, beyond topical sodium hyaluronate.

A neurostimulation device received FDA approval as an electromechanical tear

stimulator that is a non-implantable device intended to increase tear production via
mechanical stimulation. The device underwent a prospective, open-label, single-arm,
multi-center study in 108 subjects and a multi-center, nonsignificant-risk, prospective,
double-masked, randomized, sham controlled, single visit clinical trial that enrolled 60
subjects. For the latter study, the primary endpoint achieved was an increase in the
mean within-subject Schirmer score post- vs. pre-stimulation when compared to a sham
treatment group in subjects with baseline Schirmer score < 10mm.

11.5 Design features to enhance clinical trial data quality and decisions

This section covers ways to enhance clinical trial data quality and data-supported
decisions on whether to proceed with an intervention under development for a particular
indication. (See TFOS DEWS III Management and Therapy Report, Section X, 2)

11.5.1 Biomarkers / proof of mechanism / early signal of efficacy

One of the most challenging aspects of randomized controlled trials for DED is the lack
of objective, minimally invasive metrics for diagnosing DED for patient selection and for
use as efficacy endpoints. The typical use of signs and symptoms of DED, nearly
always comes with the caveat that DED affects ”a heterogenous group of subjects”.
Outcome measures in RCT though called “objective,” such as corneal staining, TBUT

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and Schirmer test value, are all subject to bias from the observer and /or biologic and
environmental variables-reflex tearing, time of day, humidity etc. Ophthalmology does
not have a lot of reliable, validated markers that correlate with clinically relevant findings
in DED. 953,1026 (See 953, DEWS II Clinical Trials Design Report 2017)

Recent examples of FDA cleared biomarkers include a point of care immunoassay test
for the in vitro detection of elevated levels of the MMP-9 protein in human tears, from
patients suspected of having DED 1027 Osmolarity testing is another FDA approved
biomarker.

Biomarkers were used in support of the approval of 0.1% cyclosporine and are
referenced in the EU Assessment report (level of HLA-DR expression). 1028 More
recently a genetic marker (TNFR1 marker) was used in evaluating anti-TNF alpha,
licaminlimab, and demonstrated improved outcome measures in a genetically defined
subgroup analysis. 1029 To date, no biomarker other than 10-millimeter increase or more
in Schirmer scores has supported regulatory approval for a DED treatment with the US
FDA where they could theoretically serve as a primary sign endpoint if validated as
clinically meaningful with an associated improvement in a primary symptom endpoint.

11.5.2 Missing data considerations

See TFOS DEWS II Clinical Trials Design report. 953

11.5.3 Global trial designs

Diagnostic tools and outcome measures have often been highly variable resulting in
inconsistent observations across clinical trials and programs. This has complicated
clinical development in that multiple trials may be required to achieve a successful drug
approval (e.g., lifitegrast ophthalmic solution 5.0%), 962,964-966 but it has also impacted
harmonization of regulatory requirements across regions. Regulatory requirements can
vary considerably across the major DED markets which include US, the European
Union (EU), and Japan.

For the clinical development of drug therapies, the commonly selected sign and
symptom endpoints used in the trials have been similar across the major markets:
corneal or conjunctival staining (although using different scales), Schirmer score, and
symptom questionnaires. For symptom questionnaires there seems to be a trend
favoring overall symptom questionnaires (e.g., OSDI) and Visual Analogue Scales (i.e.,
ocular discomfort or dryness). 951,952,955-957,962-966,972,974,977,979,1012

Across the US, EU and Japan compounds targeted at a similar sub-population of

patients who present predominately with signs of inflammation or its impact (e.g.,
hyperemia, corneal staining, and conjunctival staining) and ADDE (e.g., decreased tear

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production) have been approved. 951,952,955-957,962-966,972,974,977,979,1012 The major difference
between the regions is the duration of follow-up for the primary endpoints. The US
allows multiday, natural exposure trials of 2-weeks duration or longer, EMA allows
natural exposure trials of 6-months duration or longer and Japan allows natural
exposure trials of 4-weeks duration or longer. 949,1004,1009,1023

Taken together, the commonalities between the regions could allow for a single trial with
a common disease population and endpoint to support approval in more than one
region leveraging multiple statistical analysis plans specifying primarily endpoints at
different timepoints. 975,978 This allows a single trial to potentially support approval in
both the US and EU. To include Japan a sub-population of Japanese participants would
need to be included. While this could work for new compounds targeting the patient
populations and endpoints which have already been accepted in the major markets, it is
unclear how alternative endpoints (e.g., meibomian gland morphology-related endpoints)
will be accepted globally.

11.5.4 Compliance monitoring

Adherence to prescribed therapy is essential for quality clinical trials data and reliable
comparison of treatment modalities. Strategies for assessing adherence to treatment
vary in their applicability to ophthalmic trials but may include assessment of amount of
returned investigational product, patient report by regular electronic or other means,
direct observation of drug or device use, systemic or ocular monitoring of drug or
metabolite levels or more technical solutions such as electronic monitoring, for example
app control of reusable devices can map how long and how often they are used or
sensors in topical medication containers.

11.6 Conclusions

Newer market entrants have utilized clinical trial designs with enriched populations of
patients that take into consideration the mechanism of the drug under study, particularly
as it pertains to defining sign endpoints. 973,974,976,977 This has increased their likelihood
of technical success and is aligned with the US FDA Draft Guidance. This guidance
states that a statistically significant difference between the investigational treatment and
vehicle for at least one objective prespecified sign of DED (mean group score for test
versus vehicle) AND at least one subjective prespecified symptom of DED (mean group
score) can be used to support approval. A sign endpoint consistent with a drugs
mechanism of action can be selected to demonstrate that the drug under study has its
intended effect. The significance of the symptom endpoint then establishes the clinical
relevance of the observed effect the drug under study has upon how a patient feels
making it clinically relevant. 949 Thus, the recommendations from the TFOS DEWS II,
the industry report to identify three fundamental elements needed to achieve a

135
significant improvement in trial success rates (Pillars 1-3) and the US FDA Draft
Guidance for Industry on DED all coalesce on the same conclusion for achieving
regulatory and technical success for getting a new drug approved: match the sign
endpoint to the intended mechanism of the drug under study.

In conjunction with better selection of patients, trial designs have significantly evolved
over the 20 years of research. They have gone from little control over confounding
variables like artificial tear use 963 to careful control over comparator arms and run-in
periods within confirmatory efficacy studies. 954,955,966,973,974,976,977,979 Global regulatory
agencies have recognized this evolution with the FDA even encouraging trial designers
to consider that even water is known to be an effective component of topically applied
treatments for DED. Thus, comparative clinical trials should use only the vehicle from
the investigational drug as a control agent.

The agencies have also recognized that trial endpoints could change in sensitivity along
with disease severity and as a result have also evolved the requirements for replication
in confirmatory efficacy studies. The US FDA Draft Guidance recommends the sponsor
of a new DED treatment demonstrate efficacy and safety in at least two adequate and
well-controlled, multicenter independent studies. This is because the agency recognized
that different study designs, populations and timepoints may be needed to support sign
and symptom endpoints in different studies. 949 Global regulatory agencies also allow
pre-specified and alpha corrected secondary endpoints to support approval even if the
primary endpoint miss statistical significance 951,956,957,971,972,1007,1008 While major
differences still remain for trial requirements across the major markets, advances in trial
design, patient identification and statistical methods will undoubtedly continue to
advance allowing for more global confirmatory efficacy studies that can support
approvals in more than one region.

12. Summary
This report has explored key research published since the 2017 TFOS DEWS II
Workshop reports to underpin the evidence described in the TFOS DEWS III Diagnostic
Methodology 1 and Management and Therapy 2 reports and to present how new findings
from each of the topic areas considered, have informed the drivers of disease. Each
topic area has also identified key research needs for the next short and medium time
frame.

Table 16 below shows how each of the sections of the Digest report inform evidence for
the drivers of disease. Figure 6 illustrates the key findings from these interdisciplinary
areas, all of which have informed diagnosis and methodology and pathways towards
new treatment modalities. The importance of disease subtyping and the relevance of
new biomarkers has been consistently recognized.

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Figure 6. Key findings from interdisciplinary reports

137
Table 16. Digest sections and drivers of DED

Drivers of DED

Tear film mucin/glycocalyx

Anatomical juxtaposition

Inflammatory /oxidative
stress abnormalities
Ocular surface cell
Tear film aqueous

Blink/lid closure
Tear film lipid
abnormalities

abnormalities

abnormalities

abnormalities

abnormalities

abnormalities

abnormalities

abnormalities
Lid margin

Nerve
Sex, gender,
✔️ ✔️ ️ ️ ✔️ ️
and hormones
Epidemiology ️ ️ ️ ️ ️ ️ ️

Pathophysiology ️ ️ ️ ️ ️ ️ ️ ️ ️

Tear film ✔️ ✔️ ✔️ ✔️ ✔️ ✔️
Pain and
✔️ ✔️ ✔️
sensation
Iatrogenic ✔️ ✔️ ✔️ ✔️ ✔️ ✔️ ✔️ ✔️ ✔️
Clinical trials
✔️ ✔️ ✔️ ✔️ ✔️ ✔️
design

13. Acknowledgements

Rajendra Gyawali [email protected]; Judy Nam [email protected]; John Everett

Serralta [email protected]; Pragnya Rao [email protected]

The TFOS DEWS III effort was supported by unrestricted donations from Alcon, Bausch
+ Lomb, Azura, AbbVie, CooperVision, Dompé, Espansione Group, Harrow, Laboratoire
Théa, SIFI, SINQI, Tarsus, Topcon and Trukera.

14. Figures

Figure 1. Thyroid eye disease

Figure 2. Prisma flowchart of the literature search outcome for prevalence studies
published between 18 September 2015 and 29 June 2024.

138
Figure 3. Prevalence of DED based on age and sex for different diagnostic criteria

Figure 4. Treatments for Dry Eye Disease (DED)

Figure 5. Determining the regulatory pathway for a device (from Van Norman 2016)

Figure 6. Key findings from interdisciplinary reports

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