5 Corneal Epithelial Physiology

T or F, the basement membrane of the corneal epithelium is an ECM

T

ECM can be divided into two categories:

pericellular matrix and interstitial matrix

ECM is a dynamic, 3D network of molecules that…

provide structural support for cells and tissues

The basement membrane of epithelia and glycocalyx are what category of ECM?

pericellular matrix

Stroma of cornea (and cartilage/bone) is what category of ECM?

interstitial matrix

What cell/tissue processes are regulated by ECM?

proliferation, migration, differentiation, angiogenesis, immune function, autophagy, and tissue separation/shaping

Major components of ECM (just to see them)

proteoglycans, collagens, MMPs, elastic fibers, lysyl oxidase, laminin, tenascin, fibronectin

Significance of proteoglycans having negative charge in ECM

binds to water

MMPs

matrix metalloproteinases, break down components of ECM

Two layers of any basement membrane

lamina lucida, lamina densa

What is the principal component of lamina lucida of any basement membrane?

Laminin (protein)

Principal component of lamina densa

Collagen IV

What are the anchoring fibrils between lamina densa to sublamina densa?

Collagen type VII

What adheres BM (ECM) to cells?

hemidesmosomes

Hemidesomosomes are in the __ membrane of corneal epithelial cells

basal membrane

What are the components of hemidesmosomes?

intermediate filaments, plectin, integrins

What are intermediate filaments?

A structural cytoskeletal component that strengthens hemidesmosomes

What is plectin?

forms intracellular plaque in hemidesmosomes, connects intermediate filament with integrins

What are integrins?

transmembrane proteins in PM, bind to laminin extracellularly (in lamina lucida), and Plectin (plaque) intracellularly

Describe molecular structure of Collagen IV

Molecular subunit of 4 proteins; chicken wire-like network → strong BM

What might cornea erosions indicate is wrong?

failure of adherence of corneal epithelial cells to BM/Bowman’s via hemidesmosomes (EMBD)

T or F, there is no pain associated with cornea erosions

F, exposed nerve endings induces pain

What is the most common cause of cornea erosions?

corneal injury (trauma)

What diseases come with genetic predisposition to cornea erosions?

Alport’s syndrome, Epidermolysis bullosa

Alport’s syndrome

mutations in Collagen IV → epithelial cell death → cataracts

Epidermolysis bullosa

disruptions in other hemidesmosomal/BM genes

EBMD names

Epithelial BM dystrophy aka Cogan’s microcystic epithelial dystrophy aka map-dot-fingerprint dystrophy

EMBD presentation

map-like lines and subepithelial microcysts where BM has intruded into more superficial layers of cornea

In EBMD, does BM have abnormal thickened or thinned region?

thickened, extra deposition of ECM

2 characteristics of stem cells

1) unspecialized/undifferentiated capable of renewing themselves through cell division

2) have potential to differentiate into a specific cell type

Stem cells can be grouped into two kinds:

1) Embryonic stem cells (pluripotent or totipotent)

2) Somatic “adult” stem cells (multipotent)

pluripotent

can give rise to all tissue cell types

multipotent

can give rise to multiple cell types

unipotent

gives rise to a single cell time

All ocular tissues are derived from what pluripotent stem cell line?

ectoderm (ocular structures) and mesoderm (vasculature, muscles)

Transit Amplifying Cell

highly proliferative cell derived from stem cell, after it committed to developing towards a specific line

Which has a higher proliferation rate, stem cells or transit amplifying cells?

TACs, don’t want stem cells to proliferate uncontrollably

Describe X+Y=Z

X: basal cell proliferation anteriorly (mitosis)

Y: centripetal movement of the epithelial cells from limbus towards center

Z: cells being shed from surface

Describe the conflicting hypotheses of LESC vs CESC

LESC: SC found exclusively in limbus, and TACs move/divide centripetally to refresh corneal epithelium during homeostasis and injury

CESC: SC in limbus AND basal cornea epithelium, TACs move/divide centrifugally. CESCs contribute during homeostasis and injury, while LESCs contribute only during injury

What causes exfoliation/desquamation of corneal epithelial cells?

• constant cell loss (sloughing off)

• Blinking

• Minor abrasions

• Eye rubbing

Centripetal movement of cells (~10-15mm/day) is likely due to __ and causes entire corneal epithelium to be renewed in __ months

growth pressure; 9-12 months

Where are limbal epithelial thickenings (source of epithelial cells)?

between palisades of Vogt

What is the source of epithelial cells?

limbal epithelial thickenings between palisades of Vogt

There is more or less proliferation in the posterior/peripheral cornea

More proliferation

LSC location

Limbal stem cells are in the limbal basal cells

Do LSCs (limbal basal cells) have high or low mitotic activity? Short or long life span?

Low; long

LSCs are __potent

multipotent

When needed, cell division of LSC produces what 2 cells?

• 1 remains a multipotent LSC

• 1 becomes a TAC and differentiates

What happens if the LSC becomes a TAC?

TAC has increased mitosis rate, migrates centripetally toward center and anteriorly (become wing and superficial cells)

Do TACs become more or less multipotent with each division?

less

When do TACs become post-mitotic (terminally differentiated)?

stop dividing once anterior?

Stem cell “niche”

region that protects stem cell population and helps maintain their multipotency

What is the stem cell niche of limbal stem cells?

epithelial thickenings in limbus

What factors aid multipotency in niche?

• Thickness of epithelium

• Amount of melanin pigments

• Array of BM proteins different from central cornea to promote stemness and proliferation

• Presence of nearby blood vessels for O2, GFs, antioxidants

What do blood vessels provide for stem cells in a niche?

O2, growth factors, antioxidants

Where are stromal stem cells relative to the limbal basal cells?

subjacent to limbal basal cells

What SC niche likely nurtures stromal stem cells?

limbal stem cell niche

What is the definition of stem cells that stromal SCs fit?

• can self renew

• Are multipotent

What do stromal stem cells differentiate into?

keratocytes

Conjunctival stem cells are concentrated in __ conjunctiva or spread throughout conj, not sure

fornicial conj

What do conjunctival stem cells differentiate into (they are multipotent)?

epithelial and goblet cells

How are conjunctival stem cells different from limbal stem cells?

Conj SCs do not help repopulate cornea (don’t express corneal specific proteins), and limbal don’t supply the conj(?)

What happens if limbal stem cells can’t repopulate corneal epithelium?

conjunctival stem cells spread into cornea

What would patient complain of with a limbal stem cell deficiency?

blurry vision, foreign body sensation, photophobia, tearing, pain

What can be observed with slit-lamp with limbal stem cell deficiency?

Loss of palisades of Vogt

Would corneal epithelium thin or thicken with limbal stem cell deficiency?

thin, limbal SCs not repopulating cornea

What can limbal SC deficiency be caused by?

autoimmune disorders, chemical/thermal injury, contact lens wear, surgical damage, infections, congenital malformations…

OSSN general definition and presentation

Ocular Surface Squamous Neoplasia: stem cells divide inappropriately → range from mild dysplasia to invasive squamous cell carcinoma

What areas are more susceptible to OSSN?

conj of limbus (from basal limbal stem cells); also affects cornea

Risk factors for OSSN

UV exposure, HIV, HPV

Pterygium

encroachment of bulbar conj onto the cornea

What resembles an aberrant wound healing response?

pterygium

2 potential etiologies of pterygium

• Mutation of limbal stem/epithelial cells → proliferate/differentiate into conj cells

• Destruction of limbal stem/epithelial cells (usually a barrier to conj proliferation into cornea) = local limbal stem cell deficiency

Which involves zigzag vessel patterns, OSSN or pterygium?

OSSN

Which involves straight vessel patterns of neovascularization, OSSN or pterygium?

pterygium

Which involves surface keratinization, OSSN or pterygium?

OSSN

Which can become invasive/malignant, OSSN or pterygium?

OSSN

Which involves growth of subepithelial conjunctival fibroblastic tissue over cornea, OSSN or pterygium?

pterygium

Which variably involves growth of conj, limbal/cornea epithelia, OSSN or pterygium?

OSSN

Which involves underlying stroma of activated fibroblasts and ECM remodeling, OSSN or pterygium?

pterygium

Which involves goblet cell hyperplasia, OSSN or pterygium?

pterygium

What is a leading edge of altered limbal epithelial cells characteristic of?

pterygium

Pinguecula

UV damage of fibroblasts beneath epithelium → inappropriate (excess) production of EM proteins (elastin) and possible increase in fibroblast cell number

Where do pinguecula occur?

bulbar conj and limbus

What is a creamy-colored, chalky growth on conjunctival surface characteristic of?

pinguecula

Pinguecula vs pterygium

Pinguecula is more extracellular mass from some damage of cells (no proliferation), pterygium is mutation in p53 (tumor supressant) → fibroblasts proliferate

How can pingueculas affect limbal stem/epithelial cells?

extracellular mass beneath epithelium pushes epi superficially, disrupting protective mechanisms (more exposed to UV, pushed away from vasculature of niche) → can cause mutation of DNA in limbal stem/epithelial cells → pterygium and pinguecula can occur together

Stages of epithelial wound repair

Stage 1. Latent phase, 4-6 hrs after wound - recruit immune cells, change molecular pathways, secreting EGF

Stage 2: Cell migration - cover cells

Stage 3: Epithelial mitosis - make up for loss of cells, activating stem cell pathway

Stage 4: Reassembly of adhesive contacts (hemidesmosomes)

What morphology changes occur in the epithelial cells in the latent phase of epithelial wound repair?

• Loss of surface microplicae/glycocalyx

• Retraction of epithelial cells at wound edge

• Basal cells flatten

In what stage of epithelial wound repair is there signaling/chemotaxis? What is released?

Stage 1, latent phase; cellular stress causes release of Ca2+, H2O2, and nerve damage releases cytokines and substance P

What happens to the basement membrane components (hemidesmosomes, ECM…) in latent phase of epithelial wound repair?

They degrade

What facilitates degradation of hemidesmosomes and ECM in latent phase?

PMNs phagocytize damaged cells and signal for more PMNs

T or F, mitosis is stimulated in latent phase

F, mitosis is inhibited

PMNs infiltrate tears from…

lymphoid follicles and diffuse lymphoid tissue beneath conj

PMNs infiltrate stroma from…

limbal blood vessels upon corneal abrasion

How many cells thick is the leading edge of migrating cells during stage II of epithelial wound repair?

one cell thick

Focal adhesions in migration in wound repair are attachments of what molecules?

actomyosin and transmembrane integrins

What do hemidesmosomes attach to in ECM?

intermediate filaments

What cellular processes help epithelial cells sense the environment in cell migration, and extend from margin of epithelial wound?

lamellipodia (broad) and filopodia (narrow)

What are lamellipodia and filopodia (or cell migration?) driven by?

cytoskeleton polymerization - actin, myosin