Ординатура / Офтальмология / Английские материалы / Basic Sciences in Ophthalmology_Velayutham_2009

Fig 13.2: Cornea—Matrix proteins

collagen forms oblique running lamellae in the stroma. Type V assumes anterior conformation that prevents the attachment of additional fibres and this limits the diameter of type I. This tight control of the diameter prevents light scattering in the corneal stroma. Type V aminoterminal domains project onto the fibril surface and when sufficient numbers have accumulated, they block further accretion of collagen monomers and there by limit growth in diameter. Type VI collagen stabilizes the proteoglycans and keratocytes located between the lamellae.

The inter fibrillar distance is maintained at around 55 nm. Only when the distance between the regions of different refractive indices becomes greater than 200 nm, the light scattering occurs. So the critical factor is the interfibrillar distance for maintaining the corneal transparency.

The parallel arrangement of the central corneal fibrils extends to the periphery to form a concentric configuration and curvature of cornea. The curvature of cornea is important in refracting and focusing light to produce an image on the retina. In the corneal stroma, 2 types of glycosaminoglycans viz, keratan sulphate and dermatan sulphate are found linked to core protein lumican and decorin respectively. These proteoglycans bind to collagen at specific binding sites.

Function

Maintenance of corneal clarity and curvature. The proteoglycans serve as spacer molecules between the collagen fibres of the stromal lamellae. Glycosaminoglycans carry negative charge and hence, attract sodium and water. This is important to maintain the corneal clarity as it generates a level of interfibrillar tension that maintains the interfibrillar distance necessary for light transmission.

Clinical Application

In certain diseases of the cornea, for example; corneal dystrophies-water enters the space occupied by the proteoglycans and increases the distance between the collagen fibers. So, light scattering occurs resulting in corneal opacity.

Mucopolysaccharidoses are diseases associated with degradation of glycosaminoglycans due to degradative enzyme deficiency. The resultant partially degraded glycosaminoglycans deposit in the cornea leading to corneal

opacity, e.g; Hurler’s syndrome: Deficiency of α iduronidase, GAGs containing iduronic acid viz heparan sulfate and Dermatan sulfate are not broken down after the initial removal of sulfate. These GAGs accumulate in cell lysozymes, spill over and get deposited in cornea causing opacity.

Abnormalities in curvature can distort image even with clear cornea, e.g. astigmatism, keratoconus, wound scarring. Keratocytes are corneal fibroblast and are the site for post-translational modification of collagen and also proteoglycans synthesis as they contain the necessary enzymes and hence important for corneal transparency.

Bowman’s Layer

Bowman’s layer is immediately posterior to epithelial basal lamina and is secreted by keratocytes (anterior stromal). It is usually of Type IV collagen. Here, one type IV collagen joins the other type IV collagen by association of its noncollagenous peptide extensions to form a spider web like structure or open mesh and not a fiber (Fig. 13.3). This layer is acellular and anterior keratocytes do not repair, damage to this layer after initial formation during embryogenesis.

Function

It separates epithelial cells from adjoining stroma and supports epithelial cells in their location.

It acts as a sieve or barrier for molecules that may approach its associated cells.

Descemet Membrane

Descemet membrane is secreted by endothelial cells and is made of type VIII collagen which forms geometric patterns resembling a box spring mattress or lattice structure. The non-helical regions of this type VIII collagen can form bonds with type IV collagen (Figs 13.4 and 13.5).

Function

1.Provides elasticity and deformability to cornea.

2.Imparts strength and resilience to corneal stroma.

3.Maintains light transmission.

Fig 13.5: Partial lattice structure

4.It is the main resistance to intraocular pressure.

5.It is resistant to proteolytic enzymes.

Anchoring fibrils extend between the basal epithelial cells and the outer most lamella of the corneal stroma, extending through the Bowman’s membrane. It is of type VII collagen. It is attached from the hemidesmosomes of the epithelial basal cells to stromal type I collagen fibers.

Function

It serves to attach the epithelium to the stroma strongly.

Biomedical Importance/Clinical Application

In diabetes mellitus, the synthesis of anchoring fibrils is reduced resulting in loose adhesion of the epithelium to its underlying stroma.

Endothelium

Functions as a fluid pump to keep the cornea in a clear, deturgesced state. ~ 10 µl/hr of fluid is pumped back into the anterior aqueous by the corneal endothelium. Endothelium is a non-vascular, highly metabolic, single cell layer of hexagonal cells of uniform size and shape, bathed by aqueous humor of the anterior chamber. No mitosis occurs in adult corneal endothelium and so cells decrease in number throughout life.

It is termed “metabolic pump” instead of “fluid pump” because it allows the water to move osmotically down the gradient setup by active transport of ions. pH and osmotic requirements of endothelium are: 6.8–8.2 pH and 200– 400 mosm / kg of osmotic tolerance with 304 mosm/kg as optimum.

Transport of ions in cornea:

1.Epithelial ion transport.

2.Endothelial ion transport.

Endothelial ion transport:

Several ion transport systems exist in the corneal endothelium.

1.Sodium Potassium Pump (Na+ /K+ ATPase).

2.Sodium / Hydrogen exchanger.

3.Bicarbonate / Sodium cotransporter.

1.Sodium Potassium Pump (Na+ /K+ ATPase) (Fig. 13.6)

It has two special functions in ocular tissue.

Fig 13.6: Sodium potassium pump

1.Control of corneal hydration.

2.Production of aqueous fluid.

It is located in basolateral membrane of the endothelial cells about 1.5 × 1000000 pump sites / cell. It is an enzyme – sodium, potassium stimulated adenosine triphosphatase, in short Na+ / K+ ATPase. It is membrane bound or an integral protein, spanning the width of the cell plasma membrane. It consists of 4 polypeptide chains 2 α and 2β chains. β chains with small chains of carbohydrate are necessary for membrane insertion, stabilization and orientation of α subunit.

Alpha chains are the actual catalytic molecules for which the substrate is the high energy compound ATP.

ATP Na+/K+ ATPase→ ADP + Pi

–———–

This reaction is energetically coupled to an ion transport process. Pi becomes bound to one of the α subunit and in the process supplies energy necessary to transport 3 Na+ out of the cell and 2K+ inward. The actual transport of ions is assumed to take place either by a conformational shuttle within the α subunit or ions pumped through the pores present in the subunits. The pumping out of Na+ into the narrow channel between the adjacent endothelial cells generates a counter osmotic pressure drawing water also out of the stroma. The other transporters support this enzyme in their action.

2.Sodium / Hydrogen exchanger: It is present in the basolateral membrane moving Na+ into the cell and H+ outward. This is necessary to maintain the intracellular Na+ that is pumped out by Na+/ K+ ATPase. The hydrogen

ions acidify the extracellular fluid, increasing the level of CO2 which diffuses into the cell.

3.Bicarbonate / Na+ cotransporter: It is present on the apical membrane

of the cell. The CO2 inside the cell combines with water in the presence of the enzyme, carbonic anhydrase to form carbonic acid which then

dissociates into H+and HCO¯. Both are transported out, H+ from this reaction

3

exchanges with Na+, so that this can provide the ions necessary for the action of Na+/ K+ ATPase. The HCO3¯ from this reaction goes out through the bicarbonate transporter which carries one Na+ also for 2 HCO3¯ transported. All these transport systems are interlinked or dependent on each other.

Na+ from the lateral space goes into aqueous humor which has negative potential and along the path of least resistance into the stroma. Water follows sodium through osmotic gradient.

Chloride transport also has a role in this. Thus, the deturgescence or corneal hydration is maintained by these endothelial pumps (Fig. 13.7).

Fig 13.7: Endothelial pumps

Epithelial Transport

Sodium potassium pump. Sodium/chloride co-transport pump.

Model of ion transport, ion channels and sympathetic neural control of chloride channels in the corneal epithelium (Fig. 13.8).

The Na+/ K+ ATPase ion pump in the basolateral membrane maintains the Na+ gradient for Na+/ Cl¯ co-transport. Chloride diffuses down its chemical gradient through apical channels which are opened by cAMP. Corneal epithelium is a tight epithelium with low ionic conductance for Na+ through its apical cell membrane (sodium is pumped from tears to stroma while chloride is transported into tears).

Fig 13.8: Epithelial transport system

Sodium from tears enters the corneal epithelial cell through low conductance channels in the apical membrane of the superficial cells. It is then extruded into the stroma by the Na+/K+ ATPase located in the basolateral membrane of the cells, i.e. 3 Na+ out in exchange for 2K+ into the cell. As there develops a Na+ gradient in the stroma, it moves down the gradient into the epithelial cell again of while moving it carries the Cl¯ also along with it. Cl ¯ movement against the concentration gradient is possible because of this Na+ co-transport pump. There is a high conductance K+ channel in the basolateral membrane transporting K+ into the stroma. Once inside the cell, Cl ¯diffuses into the tears through channels in the apical cell membrane. The transport processes in the corneal epithelium result in osmotic transport of water out of the cornea. Epithelial chloride is stimulated by catecholamines through ß adrenergic receptors and second messenger cAMP.

The epithelial transport of water and electrolytes also helps in maintaining the corneal transparency.

Biomedical importance

Chemical burns of cornea: caused by mineral acids and alkali, requires immediate medical attention e.g., NaOH is the alkali, exposure to which causes instantaneous cloudiness of cornea.

Early chemical damage results in cell destruction and disruption of collagen of proteoglycans. With sufficently concentrated alkali, the cell membrane is damaged by saponification or lysis of lipid ester bonds in the membrane resulting in cell death. The OH¯ of the alkali binds to the basic amino acid of collagen and through Vanderwalls forces and hydrogen bonding gets sorbed to the collagen molecules, allowing water molecules to enter the interfibrillar spaces. This causes swelling and distortion of collagen fibres. The peptide bonds of

the exposed and distorted fibres are hydrolysed by the alkali. Also, the proteoglycans are cleaved at their linkage with GAGs, all these result in obliteration of the regular interfibrilar distance and hence the loss of transparency.

Immunochemical damage

When the alkali burns is severe (>0.1 M alkali exposure) the initial chemical damage is followed by destructive inflammatory response as the reparative process cannot commence due to cell destruction. The response is by infiltration of polymorphonuclear lymphocytes within the first 2 days of the burns, ulceration of tissues, release of matrix metallo proteinase, which will further lyse and damage the corneal collagen and proteoglycan proteins. This continual degradation of corneal collagen can lead onto perforation of cornea.

Cell turnover and wound healing in Cornea

Epithelium is constantly being regenerated by mitotic activity in the basal layer of cells. The initial response following epithelial debridement is migration of cells as a flattened sheet of single layer across the stroma to close the defect. Migration is achieved by the redistribution of cytoskeletal actin – myosin fibrils. Adhesion of epithelium to basement membrane and Bowman’s layer is through hemidesmosomes that form within 18 hours. The lamina densa and the anchoring fibrils of type VII collagen take many days to form. Thus, single cell layer is restored to its six layered architecture.

Stroma: Immediate effect is to cause wound gaping and imbibing of water from tears by GAGs causing local opacity. Then, keratocytes are activated to synthesize GAGs and collagen. In early stages of healing, there is loss of specialization in keratocytes and loss of regularity in the arrangement and size of fibrils leading to further opacity in the cornea. Only after (many months) a long time, cornea restores clarity by producing normal corneal matrix component in small, well defined wound and not in extensive wounds.

Endothelium: does not have mitotic activity, but undergoes cell slide i.e. migration and loss of cells. If considerable number of endothelial cells are lost, the pumping cannot be performed; cornea will imbibe water and become opaque.

Vascularisation occurs during healing, if the defect does not close promptly.

Metabolism of Cornea

The primary metabolic substrate for energy in cornea is glucose. Glucose is provided to the stroma, primarily from aqueous humor by carrier mediated transport through the endothelium to the epithelium by passive diffusion through stroma. Oxygen is supplied largely by the atmosphere. The percentage of glucose utilization through various pathways is given below.

pressure of O2 falls below 54 mmHg. The recent use of rigid gas permeable lenses has largely eliminated this problem as these lenses allow the passage of O2 more efficiently than even soft contact lenses. Actual O2 consumption rate is only 2 µl/mg of corneal tissue / hour.

Diabetic cornea

Diabetes mellitus produces 3 pathological effects on the cornea:

1.Reduced epithelial adhesion to the corneal stroma.

2.Loss of corneal neural sensitivity.

3.Possible increase in corneal thickness principally in the stroma.

1.Reduced epithelial adhesion is due to the glycation of the protein laminin that forms the extra cellular matrix needed for spreading and attachment of epithelial cells. This impairs the ability of corneal epithelium to repair itself especially after corneal surgery, the collagen anchoring fibrils are defective leading on to tissue erosions.

2.Loss of corneal neural sensitivity is due to protein AGEs (Advanced Glycation End products) in the basal lamina of Schwann cells at the anterior cornea. It impedes the ability to sense corneal contact such that patient is unaware of bacterial infections and so undetected corneal ulceration can occur.

3.Increase in corneal thickness results in corneal swelling from the osmotic effect due to stimulated polyol pathway.

UVEAL TRACT

Uveal tract is the vascular, middle compartment of the eye consisting of three parts viz. Iris, ciliary body, anteriorly and choroid in the posterior uvea.

The major functions of uvea are:-

1.To regulate the pupil size for optimal vision through the muscles of the iris viz. sphincter iridis and dilator pupillae muscles.

2.To regulate the production and composition of aqueous humor.

3.To influence the ionic environment and metabolism of lens, cornea and

trabecular meshwork.

Blood vessels of iris have tight junction and lack fenestrations rendering them relatively impermeable to large molecules, thus forming a second component of blood – aqueous barrier.

Stroma of the iris is composed of pigmented melanocytes and non-pigmented epithelial cells and matrix of collagen Type III fibrils with hyaluronic acid as GAG. Anterior border is avasucular and allows the passage of aqueous humor through the loose stroma.

Ciliary body bridges the anterior and posterior segments of the eye and forms the blood aqueous barrier. It forms the aqueous humor, maintains the intraocular pressure, and uveoscleral outflow of aqueous humor apart from its role in accommodation of lens through ciliary muscle.

Ciliary muscle forms the major part of ciliary body with 3 bundles of outer longitudinal, middle radial and inner circular fibres. The cells contain multiple myofibrils, with mitochondria and nucleus.A basal lamina surrounds the smooth muscle cells.

Contraction of ciliary muscle relaxes the zonules allowing the lens to adapt a more spherical shape owing to the elasticity of the lens capsule. This accommodation is mediated by parasympathetic stimulation of longitudinal fibres followed by the action of circular fibres. It produces greater ability to focus on near objects owing to the increased refractive (diopteric) power of the lens.

Biomedical importance

Myopia may be induced by intense near work for prolonged periods and in children sleeping in dimly lit rooms due to persistence of poorly focused images through thin eyelid skin.

Ciliary processes are inward projections of ciliary body; zonular fibres of lens are attached in between these ciliary processes. These processes consist of a central core of highly vascularised connective tissue stroma and specialized double layer of epithelium covering the stromal core. The connective tissue consists of fibroblasts and Type III collagen fibrils. The vessels are highly fenestrated leaking most of the plasma components into the stroma.

Double layer epithelium

The inner layer is non-pigmented epithelium (NPE) and is in direct contact with the aqueous humor.

The outer layer is pigmented (PE) and lies in between NPE and stroma. The basement membrane of PE is the Bruch’s membrane. The PE cells are cuboidal and contain numerous melanosomes, but poor in other intracellular organelles. The internal limiting membrane of the ciliary body is the basement membrane of NPE (Fig. 13.9).

Fig 13.9: Posterior Chamber