A brief presentation on corneal physiology (Functions ,cell shapes, histology ,biochemical compositions, transparency, drug permeability and cell turnover and wound healing )

1. PHYSIOLOGY OF
CORNEA
PRESENTED BY-
KAUSTAV GOGOI
B.OPTOM 3RD YEAR
RCOJ

2. LAYOUTS-
 Introduction
 Dimensions
 Histology
 Composition
 Biochemical compositions
 Metabolism
 Corneal transparency
 Drug permeability across cornea
 Effect of contact lens wear on corneal physiology
 Corneal cell turnover and wound repair

3. Cornea is
transparent,avascular,watchglass
like structure.
Forms the anterior one-sixth of the
outer fibrous coat of the eyeball

4. Dimensions-
 Average horizontal diameter(ant.surface)=11.75 mm
 Average vertical diameter(ant.surface)=11 mm
 Average diameter=11.5 mm
 Thickness(center)=0.52 mm
(periphery)=0.67 mm
 Radius of curvature(ant,surface)=7.8 mm
(post.surface)=6.5 mm
 Refractive power(Ant.surtface)=+48 D
(post.surface)= -5 D
 Refractive Index= 1.376

5. Composition-
Water 78 %
Collagen 15%
Collagen type 1 50-55%
Type 3 <1%
Type 4 8-10%
Type 6 25-30%
Other protein 5%
Keratan sulphate 0.7%
Chondroitin/Dermatan sulphate 0.3%
Hyaluronic acid and salts 1%

6. EPITHELIUM
 Stratified squamous type cells present
 50-90 µm thick and consists of 5-6 layers of cells
(deepest basal layer is of columnar cells
(middle 2-3 layers of wing or umbrella cells)
(most superficial two layers are flattened cells)
 Entire epithelium is replaced in a period of 6-8 days

7. BOWMAN’S MEMBRANE
 Consists of acellular mass of condensed collagen fibrils
 8-14 µm thick
 Shows considerable resistance to infection and injury
 Once destroyed,it does not regenerate

8. STROMA
 Constitutes most of the cornea
 0.5 mm in thickness
 Consists of collagen fibrils(lamellae) and cells embeded in hydrated
matrix of proteoglycans
 Lamellae are arranged in many layers(200-250)
 In each layer,lamellae are parellel to each other and corneal plane also
and become continuous with scleral lamellae at the limbus
 Cells present are- Keratocytes,wandering macrophages,histocytes and
few lymphocytes

9. DESCEMET’S MEMBRANE
 Strong homogenous layer which binds stroma posteriorly
 Represents basement membrane of the epithelium
 Made up of glycoprotein with no elastic fibres
 Thickness – 3µm at birth
10-12µm in young adults
 Resistant to chemical agents,trauma,infection and pathological
processes
 Can maintain the integrity of eyeball even after whole stroma is sloughed
off
 When destroyed,it can regenerate

10. ENDOTHELIUM
 Consists of single layer of flat polygonal(mainly hexagonal cells)
 Cell density – 6000 cells/mm2 at birth
2400-3000 cells/mm2 in young adults
 Polymegathism fills the defect left by the dying cells and hence,these
cells vary in diameter from 18-20 µ in early life and 40 µ or more in aged
 Cells are best evaluated by specular microscopy
 Cells are attached to Descemet’s membrane by hemidesmosomes and
laterally to each other by tight junctional complexes

11. Biochemical composition of Epithelium
 Contains 10% of the cornea
 Protein synthesis is in high number
 Water-70% of wet weight
 Contains Phospholipid and Cholestrol
 Contains enzymes
 Acetylcholine and ATP,Glycogen and Glutathione
 Electrolytes like Na+,K+,Cl-

12. Biochemical composition of Stroma-
 Contains 80% water and 20% solids
 Collagen(I,V,XII,XIV
 Soluble proteins like albumin,immunoglobulins,glycoprotein
 Proteoglycans(GAG fractions – keratan sulphate50%,chondroitin sulphate
25% and chondroitin 25%
 Glycolytic and krebs cycles enzymes
 Matrix mellanoproteinases :MMP (collagenase-I),MMP-2(Gellatinase
A,MMP-3(Stromelysin I)
 Electrolytes and salts

13. Biochemical composition of Dua’s layer
 Collagen Type I,type IV,type VII more than stroma contains
 Type V collagens are also available
 Proteoglycans like Lumican,Mimecan and Decorin (equal amount both in
stroma and Dua’s layer
 No keratocytes present

14. Biochemical components of Descemet’s
membrane-
 Composes of collagen(73%)and glycoproteins
 Descemet’s membranes collagens are insoluble and extremely resistant
to chemical and enzymatic action
 Doesn’t contain GAG

15. Biochemical component of Endothelium-
 Single cell layered structure
 Contains enzymes for glycolysis and Krebs cycle

16. Metabolism of Cornea-
 Cornea requires energy for maintainig its transparency and dehydration
 Energy is in the form of ATP is generated by the breakdown of glucose
 Most metabolising layers are epithelium and endothelium
 Sources of nutrients required for the corneal metabolism and metabolic
pathways involved are as follows-
Oxygen
Glucose
Amino acids

17. Oxygen-
 Epithelium derives oxygen mainly from the atmosphere through the tear
film as well through the limbal capillaries
 Oxygen required by the epithelium is about one-tenth from the
atmosphere when the eyes are open and about one-fourth of that
available from the palpebral conjuctiva when eyes are closed
 Endothelium derives most of its required oxygen from aqeous humour
 Mean total consumption(QO2) is approximately about 9.5 ml O2 CM2 hr-
1

18. Glucose-
 Primary metabolic substrate for epithelial cells,stromal keratocytes and
endothelium
 Rate of glucose consumption of cornea is about 100mg/cm2/hr

19. Amino acids-
 Continous supply of the amino acids is required to allow synthesis of
proteins needed for the constant shedding and replacement of the
epthelial cels of cornea
 Amino acids are supplied from the aqeous humour,principally by passive
diffusion
 Amino acids are sufficient for the synthesis of about 10 mg/hour of
proteins by the epithelial cells

20. Metabolic pathways of Cornea-
 Glycolysis pathway

21.  The rate of consumption of glucose by the whole cornea is approximately
100 mg/hr/cm2 with 90 being consumed by the epithelium
 Through glycolysis,glucose is broken down to lactic acid producing
energy in the form of 2 molecules of ATP/molecules of glucose broken
down
 Through Krebs cycle or citric acid cycle 36 ATP molecules are produced
per molecule of glucose oxydized

22. Hexose Monophosphate Shunt
 In corneal epithelium,glucose is also metabolized through the
monophosphate shunt(pentose shunt),without gaining ATP
 Purpose of glucose metabolism through pentose shunt is the production
of NADPH(Nicotinamide adenine dinucleotide phosphate) which is
utilized in the biosynthesis of lipids by corneal epithelium
 Ribose produced by the pentose shunt may be used to build the nucleic
acids,DNA and RNA

23. Corneal Transparency-
 Normal corneal transparency is the result of anatomical/physiological
factors
 Anatomical factors such as uniform and regular arrangement of corneal
epithelium,a peculiar arrangement of corneal lamellae and corneal
avascularity)
 Physiological factors(i.e relative state of corneal dehydration

24. Corneal epithelium and tear film-
 Due to homogenicity of refractive index,normal cornea is transparent
 Basal cells are firmly joined laterally to other basal cells and
anteriorly to the wing cells by desmosomes and macular
occludents
 These tight intercellular junctions account for the epithelium’s
transparency as well as its resistance to the flow of water,
electrolytes and glucose

25. Arrangement of stromal lamellae
 Maurice theory
Maurice proposed that cornea is transparent because of the uniform collagen fibrils are
arranged in a regular lattice so that scattered light is destroyed by the mutual interference
 Collagen fibrils are regularly arranged and separated by less than a wavelength of
light(4000-7000Å)

26.  THEORY OF Goldman et al.
 Applied diffraction theory to the problem and concluded that lattice
arrangement is not a necessary condition for corneal transparency
 He postulated that cornea is transparent because of fibrils are small in
relationship to the light and do not interfere with light transmission unless
they are larger than a wavelength of light(2000A)
 In confirmation,they found some ‘lakes’ areas devoid of collagen with
dimensions greater than 2000 Å

27. CORNEAL HYDRATION
 Normal cornea maintains itself in a relative hydration,which is essential
for its transparency
 Water content of normal cornea is 80%
 The factors which keep cornea hydrated and balanced by some factors
which draws water in the cornea like- swelling pressure of stromal matrix
and IOP and the factors which prevents water flow in cornea like-
epithelial barrier and endothelial pump
 Corneal thickness and hydration are lineally related
 If hydration becomes above 80%,corneal thickness increases

28. Factors affecting corneal hydration-
 STROMAL SWELLING PRESSURE
 60 mm Hg is exerted by GAGs of the corneal stroma which acts like a
sponge
 Electrostatic repulsion of the anionic charges on the GAG molecule
expands the tissue,sucking in the fluid with equal but opposite pressure
called Imbibation pressure(IP)
 Imbibation pressure is equal to SP in vitro not in vivo

29. Barrier function of epithelium and
endothelium
 Both epithelium and endothelium acts as barriers to excessive flow of
water and diffusion of electrolytes into the stroma due to semipermeable
nature
 Corneal transparency is decreased and corneal thickness is increased
when corneal endothelium is damaged and as a lesser extent if the
epithelium is damaged

30. Active pump mechanism-
 Corneal endothelium plays a predominant role in controlling fluid transport
 Pump mechanisms are active process where energy requires
 Dependent upon metabolic activity of cornea
 Na+ K+ ATPase pump (active process)
 It is an important function of endothelial function where Na+K+ activated
ATPase mediates the active extrusion of the Na+ from the tissue

31.  Bi carbonate dependent ATP-ase
 Also important in maintaining corneal thickness
 Bicarbonate transported by the endothelium is generated intercellulary via
the action of carbonic anhydrase

32. Evaporation of water from corneal surface
 Evaporation of water from the pre-corneal tear film concentrates fluid and its
osmolarity
 Hypertonicity of tear film can withdraw water from cornea
 Loss of fluid is replaced by aqueous humour

33. Effect of IOP on corneal hydration
 Swelling pressure of corneal stroma and epithelial oedema will occur
when IOP rises with a value of more than 50mm Hg
 Even with normal IOP and low swelling pressure,corneal oedema also
occur in endothelial dystrophy

34. CELLULAR FACTORS AFFECTING
TRANSPARENCY
 Keratocytes are important in maintaining transparency as they are the
stromal collagens and proteoglycans
 Contains enzymes involved in assembly of stromal matrix
 Collagen turnover in early postnatal life is about 24-50 hours

35. DRUG PERMEABILITY ACROSS
CORNEA
(1) Lipid and water solubility of the drug
 Lipophilic property of corneal epithelium and endothelium that are
crossed readily by the non-polar ( lipid soluble) drug
 Hydrophilic property of stroma is easily crossed by polar (water soluble)
compounds
 Therefore,a drug should be amphipathic

36. (2) Molecular size,weight and concentration of the drug
 Lipid soluble molecules can cross the corneal epithelium easily
irrespective of their molecular size
 Water soluble molecules with the molecular size less than 4Å only can
filter through the pores of the cell membrane
 Molecular weight of less than 100 can pass readily through the cell
membrane and those with more than 500 cannot

37.  Substances with large molecular size,when used in high
concentration,then a small amount of drug can cross the cornea following
laws of mass action
 The rate of penetration through the cornea of the drug depends upon their
concentration in the solution eg.- PILOCARPINE
HOMATROPINE
ATROPINE
STEROIDS

38. (3) Ionic form of the drugs
 Topical drugs must have capacity to exist in both ionized and non-ionized
form for a better penetration through the cornea
 Since non-ionized drugs can penetrate through the epithelium and the
ionized drugs can pass through the stroma
 Due to the barrier of both the epithelium and stroma,Flourescein which is
a negatively charged ion cannot penetrate the intact epithelium

39. Kinsey model of drug
 With Homatropine, KINSEY described a model of drug for both in non-
ionized and ionized form for penetration through the cornea

40. (4) pH of the solution
 pH can also affect the penetration of the solutes by its effect on the
electrical charges and stability of solutions
 pH range between 4-10 doesnot affect permeabilty
 pH of more than this range increases permeability

41. (5) Tonicity of the solution
 Hypotonic solutions(those 0.9% of sodium chloride) increase the
permeability of the epithelium

42. (6) Surface active agents
 Agents that reduce the surface tension,increase corneal wetting and
therefore present more drug for absorption eg.- Benzalkonium

43. (7) Pro-drug form
 Pro-drugs are lipophilic and after absoption through the epithelium are
converted into proper drug which can easily pass through stroma
 For example- Dipivefrin is a pro-drug which is converted into epinephrine
after its absorption into the eye

44. EFFECTS OF
CONTACT LENS
WEAR ON
CORNEAL
PHYSIOLOGY

45.  Contact lenses predominantly affects the function of the endothelium
 Contact lenses reduce the direct availability of oxygen to the
epithelium,thus shifting the balance from aerobic to anaerobic metabolism
 Lactate levels present in the cornea becomes doubled with contact lens
wear and carbon dioxide production is increased
 Increased acidosis has a direct effect on stromal hydration by impairing
stromal deturgescence mechanism

46.  Hard(rigid) contact lenses are usually made from
polymethylmethacrylate(PMMA) which have the greatest effect on corneal
function,in addition to restricting oxygen permeability and deplete
glycogen stores
 Hard lense induced inhibition of aerobic enzymes such as hexokinase
reduces direct glucose utilization of cornea and ultimately disturbs the
metabolism
 On the other hand soft contact lenses are made from polymers of HEMA
,silicone or other materials and permit extended wear of lens owing to
their permeability to oxygen and carbon dioxide

47.  Oxygenation of anterior cornea underneath a contact lens is brought
about by- (a)diffusion of atmospheric oxygen through the contact lens
(b)influx of oxygenated tear fluid underneath the
contact lens as a result of blinking
 Tear pump is the only source of oxygen in case of PMMA lenses

48. Tear pump supplies about
14-20 % tear exchange
underneath a RGP lens
Soft lens exchanges tears
from 1-5 % only

49.  Contact lenses manufactures nowadays,continually producing newer
biomimetic type lenses(hydrogel lenses) with increased water content
(upto 59%) in attempt to support normal corneal transparency
 Gas permeable lenses,which combines the reduced toxicity of PMMA
material with high gas transfer capability
 The wide variety of lens types and materials has led to their being
characterized on the basis of their Oxygen Flux defined as the DK value
 Oxygen Flux= DK/L × DP
where D=diffusion coefficient,K=solubility,L=thickness of the material
DP= change in the partial pressure of oxygen across material

50.  HEMA and PMMA have a low oxygen flux,while hydrogels and silicones
have a high oxygen flux
 Both DK value and thickness of the lens determines the suitability for use
in terms of gas permeability
 Contact lenses may have deleterious effects on the epithelium,which can
cause thinning,reduction in the hemidesmosomes and number of
anchoring fibrils and can reduce the adhesion of epithelium to the
basement membrane
 Extensive wearing of contact lenses produces epithelial oedema and
keratopathy in the form of punctate epithelial erosions

51. CORNEAL EDEMA
CORNEAL BASEMENT MEMBRANE
Hemidesmosomes and collagens

52. BAND KERATOPATHY
BULOUS KERATOPATHY
PEE
KERATOCONJUNCTIVITIS SICCA

53.  Rigid lenses also can produce tear film instability by causing damage to
the epithelium in the mucin layer

54. CELL TURNOVER AND WOUND
HEALING
EPITHELIUM
 Epithelium is constantly being regenerated by mitotic activity in the basal
layer of cells
 After epithelial debridement,the initial response of epithelium is to migrate
as a flattened sheet of single cells across the stroma to close the defect
 Hemidesmosomes and intercellular contacts then reform and gradually
the single layer is restored to its six layered structure by mitosis in the
peripheral basal cells
 Migration of epithelial cells is achieved by marked cytoskeletal and cell
shape changes involving redistribution of actin-myosin fibrils

55.  Fodrin and E-Cadherin proteins precede the actin distribution in the cell
 Migration of cells also dependent on intracellular signaling via
components such as fibronectin,laminin and collagen peptides
 Adhesion of epithelium to the basement membrane and Bowman’s layer
is achieved normally via hemidesmosomes,lamina densa and anchoring
type VII collagen fibrils
 Most of the mitotic activity in the epithelium takes place at limbus

56. STROMA
 Incisional wounds of the cornea that involve the stroma may be accidental
or intentional
 Deposition of fibrins within the stromal wound occurs
 Rapid epithelization of the wound incision
 Activation of keratocytes to divide and synthesize collagen and GAG’s
 Loss of specialization in the keratocytes such that they revert to a
fibroblast like function
 Production of corneal matrix to restore clarity in small wound

57. ENDOTHELIUM
 Corneal endotheium doesnot normally undergo mitosis even after direct
injury as in a perforating corneal wound
 With age,there is a decrease in the number of cells with an increase in
size and morphology
 If sufficient amount of cells are lost then the cell layer cannot perform its
pumping action and cornea decompensates water and becomes opaque

58. VASCULARIZATION
 Vascularization occurs when vessels from the conjunctiva or the deep
scleral plexus invade the periphery of the cornea during healing of the
wound or corneal ulcers
 Cytokines,Macrophageinflammatory proteins and granulocytes-
macrophage colony stimulating factor liberated from the inflammatory and
local cells stimulate further ingress of inflammatory cells and intiate a
vascularisation response