} OFFICE: LEVINE BUILDING, INT’L

STUDENTS OFFICE
} Phone: 020 0330 516
} [email protected]
} Mondays and Thursdays
from 12 –2 p.m.
} And by appointment
1. Biocompatibility and Clinical Applications of Biomaterials

2. Immune response to biomaterials, Inflammation, Anticoagulation &

Fibrinolysis, Bacterial adhesion and infection;

3. Sterilization of biomaterials,

4. Class trip\Video display: surgery*

5. Hydrogels and natural biomaterials,

6. Biomaterials used in drug delivery systems,

7. Angiogenesis in biomaterials,

8. In vitro evaluation of biomaterials,

9. In vivo evaluation of biomaterials; animal models,

10. Clinical applications of biomaterials.

1. For reconstruction
2. For therapeutic purposes

Deep brain stimulation

3. For cosmetic purposes
} A biomaterial is any substance that has been
engineered to interact with biological systems
for a medical purpose - either a therapeutic
(treat, augment, repair or replace a tissue
function of the body) or a diagnostic one.

} Over the years, materials used in medicine

(biomaterials) have made great impact on the
treatment of injury and disease of the human
body.
} Metals are extensively used
in many biomedical
applications and therefore
have a significant economic
and clinical impact on the
field of biomaterials
} metallurgical principles
govern the structure-
property relationship of
metals
} Understanding the structure
and properties of metallic
implant materials require an
appreciation of the
metallurgical significance of
the material’s processing
history
} How are metals mined and
fabricated?
• With the exception of the noble
metals (Au, Ag,…), metals
exists in the earth’s crust in
mineral form such as metal
oxides
• These mineral deposits(ores)
must be located and mined,
separated and enriched to
provide ore suitable for further
processing into pure metal and
or alloys
• E.g. Titanium containing sands
are separated using water flow
and gravity into titanium
containing compounds such as
TiO2 and Fe TiO3
• Electrostatic separations
converts Fe TiO3 to TiO2
Ø A metal supplier further processes
the bulk raw metal product (metal or
alloy) into stock bulk shapes such as
bars, wires, sheets, rods, plates,
tubes or powders
Ø These stock shapes may then be sold
to specialty companies (e.g.. Implant
manufacturers) who need stock metal
that is closer to the final form of the
implant
Ø Example a maker of screw-shaped
dental implants might want to buy
rods of appropriate metal to simplify
the machining of the screws from the
rod stock
Ø What manufacturing techniques or
processes does the metal supplier use
to transform the bulk metal product
into stock shapes?
} CASTING: casting involves
pouring liquid metal into a
mold, which contains a
hollow cavity of the desired
shape and then allowing it to
cool and solidify. The
solidified part is ejected or
broken out of the mold to
complete the process

} Products made in this

manner are called cast
products and the alloys used
to produce them casting
alloys
} Hot working/rolling: it is a
metal working process that
occurs above the
recrystallization temperature
of the material
} Cold rolling/working: it
occurs with the metal below
its recrystallization
temperature
} Products that are
manufactured by hot and cold
working the metal from large
ingots are called wrought
products
} In the forging process the metal
is hammered or pressed into a
desired shape
} Most forging processes are done
with the metal in a hot condition,
however in some cases the metal
may be forged cold
} There are two main types of
forging methods; Hammer and
Press forging
} Press forging can be further
classified as Open-die forging
and Closed-die forging
} Open-die forging: carried out
between two flat dies with very
simple shapes
} Closed-die forging: carried out
using a single pair of dies or
multiple-impression dies
} Important to the understanding of strengthening
mechanisms is the relation between dislocation
motion and mechanical behavior of metals

} The ability of a metal to plastically deform depends

on the ability of dislocations to move

} Virtually all strengthening techniques rely on this

simple principle: Restricting or hindering dislocation
motion renders a material harder and stronger.
} When metal forming processes such as
rolling, forging, extrusion and others are
performed cold the work material becomes
strain hardened and thus its stronger but
less ductile
} Many times the reduced ductility is
undesirable and a softer metal is required
} To achieve this the cold worked metal is
annealed
} Annealing: Involves heating material to
above its critical temperature, maintaining a
suitable temperature for a period of time,
and then cooling. Annealing can induce
ductility, soften material, relieve internal
stresses, refine the structure by making it
homogeneous, and improve cold working
properties
} During annealing the cold-worked metal
structure will go through a series of changes
◦ Recovery
◦ Recrystallization
◦ Grain growth
• Fabrication techniques and steps
required to fabricate a
preliminary device from stock
shapes depends on a number of
factors
• These are the final geometry of
the implant, the forming and
machining properties of the
metal, cost of alternative
fabrication methods
• TYPICAL FABRICATION METHODS
• Investment casting
• CAD/CAM
• Forging
• Powder metallurgy
• Grinding
• Polishing
• Most recently 3D PRINTING
} Computer systems used to
design and manufacture
products

} CAD (computer aided

design): is the use of
computer systems to assist
in the creation,
modification, analysis, or
optimization of a design

} CAM (computer aided

manufacturing): is the use of
computer software to
control machine tools and
related machinery in the
manufacturing of work
pieces.
} Using 3D scanners and
CAD/CAM technology patient
specific implants can be
developed and manufactured
with increased efficiency and
reduced cost

} This approach to orthopedic

implant has obvious benefits
compared to the common use
of off-shelf implants that
generally consist of a limited
range of geometries

} Patient specific implants last

longer, perform better with less
complications
} The market for implants is now
shifting towards gender and
patient specific implants
 } Process
1. Obtain a scan of the joint or
part of interest using CT
scan, x rays radiography,
lasers, ultrasonic probes
2. Use CAD/CAM to recreate the
part using the data obtained
from scanning
3. Use the software to generate
or code the fabrication steps
(milling, grinding, polishing
etc) needed to create the part
• Geometry of femur is influenced by 4. Feed the code to for example
horizontal offsets, neck shaft angle, a 5-axis CNC (computer
femoral head diameter, age, weight, numerically controlled)
height, race and genes grinder to produce the part
} Is a processes for
making a 3-D object
of almost any shape
from a 3D model or
other electronic data
source primarily
through additive
processes in which
successive layers of
materials are laid
down under computer
control
} Due to the patient specific, low run production needs
of the medical industry, 3D printing seems like the
perfect manufacturing solution.
} If a wide range of printable, biocompatible materials
could be created, additive manufacturing could have a
major impact on the global economy.
} Who knows, maybe one day 3D printers will have their
own wing at your local hospital, churning out metal
and even tissue-based replacement parts on a regular
basis.
• Surface modifications are important
to achieve among other things
stable fixation, wear resistance ,
biocompatibility, etc.
• Sintering: powdered particles of the
coating metal are heated to about
half the melting temperature to
allow them to stick to each other
and then sprayed or applied to the
surface of the device
• Nitriding: High energy beam of
nitrogen ions is directed at the
implant surface under vacuum. This
is usually used to improve surface
hardness and wear resistance
• Polishing: to vary surface roughness
by making it smooth
Ø Finally, the manufacturer of a
metallic implant device will normally
perform a set of finishing steps.
Ø These vary with the metal and
manufacturer, but typically include
chemical cleaning and passivation
(i.e., rendering the metal inactive) in
appropriate acid,
Ø or electrolytically controlled
treatments to remove machining
chips or impurities that may have
become embedded in the implant’s
surface.
Ø As a rule, these steps are conducted
according to good manufacturing
practice (GMP) and American society
of testing and materials (ASTM)
specifications for cleaning and
finishing implants.
} Internal Environment of the body
} Highly corrosive?
◦ Could degrade the implant
material resulting in the
release of harmful ions or
molecules
} Local stress field
◦ Average human experiences
1-2.5 million cycles of
stress on his or her hip per
year
◦ This translates to about 50-
100 million cycles of stress
over a 50 year period
} Hence the metallic
implant must be
◦ Biocompatible
:corrosion resistance
and chemical stability
◦ Must be
– Strong
– Fatigue resistant and
– Wear resistant …
in a highly corrosive
environment
} Toxicology – A biomaterial should not be toxic unless it is
specifically engineered for such requirements. E. g. “smart
bomb” drug release system.

} Biocompatibility – is the ability of a material to perform with

an appropriate host response in a specific application.

} Healing – injury to tissue will stimulate the well-defined

inflammatory reaction sequence that leads to healing where a
foreign body is involved.

} Unique Anatomical sites – consideration of the anatomical site

of the implant is essential.

} Ethics – ethical concerns relevant to biomaterials use have been

raised.
} All materials placed in the body have a propensity to
cause some response from the body, and that the
severity of that response could increase with duration
in the body.
} The response from the patient’s tissue is therefore an
important consideration.

} It is necessary to expand this general introduction on

inflammation to lay the foundation for the discussion
about the details of the tissue response to biomaterials
(referred to as ‘biocompatibility’).
Ø For many years, biocompatibility has been defined as ‘the
ability of a material to perform with an appropriate host
response in a specific situation’. (Williams, 1987).

Ø This has been updated recently to a more comprehensive

definition:

Ø ‘Biocompatibility refers to the ability of a biomaterial to

perform its desired function with respect to a medical
therapy, without eliciting any undesirable local or systemic
effects in the recipient or beneficiary of that therapy, but
generating the most appropriate beneficial cellular or tissue
response in that specific situation, and optimizing the
clinically relevant performance of that therapy (William,
2008).
Ø In the simplest sense, a biocompatible material or device does not
harm the patient.

Ø Arises from differences between living and non-living materials.

Ø Implants are specifically designed depending on applications

Ø No material is universally biocompatible!!

Ø Implants trigger inflammation or foreign body response.

Ø New biomaterials must be tested prior to implantation according to

FDA regulation.
Biocompatibility cont’d
} The “biocompatibility” of a medical device can be
defined in terms of the success of that device in
fulfilling its intended function.

} This is because there are many mechanisms that the

body has available to respond to material intrusions
into the body.

} A material that will not trigger one response

mechanism may be highly active in triggering another
mechanism.

} Bio-implants trigger inflammation or foreign body

response.
} Biocompatibility is primarily a surface
phenomenon …
} The implantation of a biomaterial often creates a wound,
and bleeding generally ensues.
} Blood thus typically makes first contact with the implanted
biomaterial.
} Proteins are built of long chains of only 20 amino acids
that are strung together by peptide bonds.
} They can function as enzymes that catalyze thousands of
important chemical reactions essential to life.
} Cell signaling molecules responsible for cell migration and
proliferation are made of proteins.
} Proteins play an important role in determining the final
nature of the tissue–implant interface.
} Biomaterials can promote cell/tissue attachment and activity
by allowing selective protein adsorption or can inhibit tissue
interactions by repelling protein.
} Importantly, changes in microenvironment which can occur
after biomaterial implantation, such as pH and ionic strength,
can alter the conformation of a nearby protein and hence its
function.
} Proteins also can experience structural alterations during
interaction with the solid surfaces of biomaterials and lose
some of their biological activity.
} Albumin is the most common protein in blood, followed by
the protective immune system proteins known as
immunoglobulins.
Features of Biocompatible materials
I. Absence of carcinogenicity (the ability or
tendency to produce cancer)

II. Absence of immunogenicity (absence of a

recognition of an external factor which
could create rejection)

III. Absence of teratogenicity (ability to cause

birth defects)
IV. Absence of toxicity.
} Blood compatibility can be defined as the property of a
material or device that permits it to function in contact with
blood without inducing adverse reactions.

} Thrombogenic material is a material that is NOT blood

compatible.
} Such a material would produce specific adverse reactions
when placed in contact with blood: formation of clot or
thrombus composed of various blood elements; shedding or
nucleation of emboli (detached thrombus); the destruction of
circulating blood components; and activation of the
complement system and other immunologic pathways
(Salzman and Merrill, 1987).

} A particular application in use will inform which of these

blood reactions to minimize. E.gs a haemostatic device to
promote the rapid induction of clotting (thrombogenic
material) and a coronary stent (blood compatible).
 Biocompatibility Testing
} No one material will be appropriate for all
medical device applications.

} The material, its composition and

degradation products may affect host cells
and tissues.

} The host environment may also affect

material properties and device performance.
 In vitro Testing
Types of in - Vitro tests for estimating biocompatibility
Cytotoxicity – Elution or extract test
Agar overlay test
Direct contact test
Haemocompatibility - Haemolysis assay
Clotting and complement activation
Mutagenecity – Ames test
Hypersensitivity – Lymphocyte transformation test
Leukocyte migration inhibition test
 Cytotoxicity
} Cytotoxicity is the ability to cause death or damage
at the cellular level by direct cell lysis (disintegrate)
or by fatally altering cellular metabolism, inhibition
of enzyme activity, changes in cell membrane
permeability and other sub lethal effects.

} It is distinctly different from physical factors that

affect cellular adhesion (surface charge of a material,
hydrophobicity, hydrophilicity, etc.).
 CYTOTOXICITY TESTS
Ø Elution test – the aim of this test is to determine toxic
doses and changes in cell growth or proliferation and to
compare to non-treated cells over 24-78-hours period
using hemocytometer or electronic cell counter.
Ø Agar overlay – the material is placed on an agar layer
covering the cells for 24 hours, components from test
material are allowed to diffuse through the agar to the
cells, cytotoxicity of the diffusible components is
determined by staining the cells with a viability dye and
then measuring the zone of dead cells surrounding the
test materials.
Ø Direct contact tests – changes in cell growth or
proliferation are measured in a similar manner as in the
elution tests.
 Hemocompatibility Tests
} This test is used to evaluate the effect of a material on blood
coagulation processes, thrombus formation and haemolysis
(destruction of red blood cells).
} Materials or their extracts are incubated with red blood cells,
isolated from rabbits, mice, or rats for three hours with intermittent
shaking to keep samples mixed and in contact with blood.
The amount of haemoglobin released into the supernatant from
the cells is determined spectrophotometrically and reported as
percentage haemolysis with respect to negative controls.

} To evaluate the effect of materials and their surfaces on blood

clotting, materials are exposed to whole blood serum.
} Turbulent flow of blood may increase haemolysis and or clotting –
the number of adherent platelets may be determined per unit area
after exposure to whole blood.
 Mutagenecity and Genotoxicity
} Mutagenes – accesses the effect of biomaterial on a cell genetic
material. It is widely accepted that carcinogenic behaviour proceeds
via a mutation in the genome.

} Ames test – uses a mutant bacterial cell line (Salmonella typhimurium

or E coli) that must be supplied with histidine to grow. The cells are
cultured in histidine-free environment and only those material that
mutate the cells back to state of histidine independence will allow the
cells back to grow.

} Hypersensitivity Tests – the leukocyte migration inhibition and

lymphocyte transformation tests have been used as in-vitro models
to estimate delayed hypersensitivity reaction to implant materials
and their released components.
In- vivo animal testing is necessary prior to human clinical testing.

} Short-term implantation tests – subcutaneous, intramuscular

and intraperitoneal implantation tests to evaluate general tissue
necrosis, fibrosis and inflammation.
} Long-term functional tests – device or compositionally identical
prototypes are implanted in appropriate animal models to
replicate/simulate intended end-use in humans. Functionality of
device and pathological evaluation of tissues /organs are
performed.
} Sensitization - Guinea pig, Occluded patch test, Open
epicutaneous test
} Irritation – Skin, Ocular, Mucosal
} Other – Genotoxic, carcinogenic, reproductive, cerebrospinal,
hemocompatible
} Nonfunctional tests - the first study a interaction of the implant with
physiological environment, nonspecific acute toxic or inflammatory
reaction, specialized sites such as the cornea and cerebral cortex are used
for materials.
Histological analyses may be scored or graded based on degree of tissue
necrosis/degeneration, fibrosis and types and amount of inflammatory
} Functional testes - evaluation in soft tissues, much greater complexity,
also specific in-vivo physiological assessment tests (genotoxicity,
carcinogenityreproductive toxicity, etc)
} Genotoxicity testing – alteration in DNA or chromosomal structure or
other DNA or gene damage that result in permanent inheritable changes
in cell function

} Cancerogenity tests – determine, whether chemicals or compounds that

may be released from biomaterials elicit sensitization reactions – result of
immunologically medicated reaction resulting in redness (erythema) and
swelling (edema).
} Irritation test - a localized inflammatory response without involvement of
an immunological mechanism
Fresh extract is injected intracutaneously at multiple sites – tome 24, 48,
72 hours control.
Ocular irritation – ophtalmological materials (rabbit)

} Systemic effects – chemicals released from implant materials are

distributed by the blood and lymphatic system and damage organs and
tissues. Categories: acute (within 24 hr), sub-acute (in 14 – 28 days) sub-
chronic ( 10% of an animal´s life span) and chronic (longer than 10% of an
animal´s life span.

The selection of evaluation in in-vivo depend on the animal used (mice,

rabbit, dog, etc.) and the implant material
} Standard practice – new drug to employ a double-blind study in which
placebo is randomly administrated .
} Implant testing – it is not possible to pair implanted patient which a
placebo-treated patient, because it is not possible to conceal the
implanted site from surgeon and or patient.
} Clinical trials – divided into three phases
Phase I (early trial) – biomaterial is tested on a small group of people
(ca 60-80
Phase II – large group (ca 100-300)
Phase III – comparison of the effectiveness of the new
treatment with a standard of management, (ca 1000 – 3000)
} The reports of clinical trials should discuss the accuracy and precision of all measurements as
well as define a minimum confidence level for all statistical measures of date (usually
p<0,05).The reports must also include confidence intervals or other measures of significance
associated with all derived parameters and must indicate the significance of any conclusion
arrived at by analysis of the trial.
} Stents
} Heart valves
} Electrodes for
pacemakers
} Dental implants are
grouped into these
categories
◦ Subperiosteal:are typically
placed on mandibular bone
but under gum tissue and
does not penetrate the bone
◦ Endosseous : are implants
surgically inserted into jaw
bone
◦ Transosseous implant: are
implants surgically inserted
into jaw bone totally
penetrating and emerging
on the opposite site in the
lower part of the chin
} Fillings (amalgams)
} ORTHOPAEDIC APPLICATIONS
} Hip and knee replacements
} Plates, screws and rods that
hold fractured bone
together during healing
} Fuse segments of the spine
together when the disk has
degenerated
} Look at the clinical applications with
respect to their design and materials
selection (What
properties/characteristics of these
materials make them biocompatible?)

} Research and prepare a 20-minute

presentation on your chosen
application area!

} Presentations are scheduled for next

week Wednesday and probably
Thursday!