damage and stromal exposure lead to activation of underlying keratocytes. These
release chemical messengers to encourage neighboring epithelium to detach from
the basement membrane and migrate as sheets to close the defect. Proliferation
and migration according to the X, Y, Z pattern aid regeneration of epithelial
layers Fig.1.7 (Maltseva et al.,
· Corneal epithelium wound healing:
Microvilli on the apical surface have filaments that interact with
mucins that expand into the tear film, supporting it and forming a glycocalyx
gel Fig. 1.5. Increased surface area
of the microvilli provides a strong anchor that stabilizes the tear film and
protects the cornea. The mucin matrix decreases surface tension and facilitates uniform
re-wetting of the epithelium and close interaction between the hydrophilic
aqueous component and the hydrophobic epithelial cell membranes. Cellular tight
junctions on the corneal epithelium form a barrier that provides protection
from inflammatory and microbial insults. Corneal epithelial cells live
approximately 7 to 10 days and undergo an organized apoptosis and desquamation
that is highly regulated by matrix metalloproteinases and other signaling
molecules. Complete turnover occurs weekly as deeper basal epithelium moves
toward the apex of the cornea (DelMonte and Kim, 2011).
Size of the layer
Number of Layers
Flat, microvilli, microplicae
Tight junctions, desmosomes
Suprabasal wing-like cells
Gap junctions, desmosomes
Gap junctions, desmosomes, hemidesmosomes
1.2: Characterstics of Superficial, Suprabasal and Basal Cells of the
Corneal Epithelium (Cruzat et al., 2011).
Basal epithelial cells attach to the epithelial
basement membrane, which is adjacent to the Bowman’s layer. The characteristics
of corneal epithelial cells and their junctional complexes are shown in Table 1.2. Tight junctions (zonula occludens) play an effective barrier
role and are present between the superficial cells. Desmosomes, on the other hand,
are present in all layers. Further, actin filaments, intermediate filaments, and
microtubules, which form the intracellular cytoskeleton, are present in corneal
epithelial cells (Cruzat et al., 2011).
Fig.1.6: (a) Layers of the cornea (b) Corneal epithelium
The corneal surface is covered by a non-keratinized
stratified squamous epithelium and has a thickness of approximately 50 ?m. The
corneal epithelium is comprised of 5-7 layers, consisting of superficial
squamous epithelial cells, suprabasal epithelial cells with wing-like
extensions, and a monolayer of columnar basal epithelial cells.
· Structure and function: Fig 1.6
The cornea is a transparent structure at the front of the eye. It
is a powerful refractive surface and a robust barrier that protects the ocular
contents. The cornea is oval shaped, with a 12.6 mm
horizontal and a 11.7 mm vertical diameter. The central cornea is spherical;
the peripheral cornea is flatter and thicker than the central portion
(Read et al., 2006) (Khoramnia et al., 2007).
Water and electrolytes are secreted across all conjunctival
cells using basolateral Na+ / K+ ATPase pump activity, with
water being transported transcellularly by aquaporins (Verkman
et al., 2008). This can be stimulated by noradrenergic or purinergic mechanisms (Nakamura
et al., 2010).
Mucin is stored in large secretory granules at apical
surface of goblet cells. Neuronal control of secretion allows mucin release in
response to surface irritation or microtrauma. Goblet cells are not directly
innervated, but cholinergic (acetylcholine (ACh) and vasoactive intestinal
peptide (VIP)) and adrenergic (noradrenaline) neurotransmitters diffuse from
the surrounding vascular and subepithelial conjunctival autonomic plexuses. Cholinergic
neurotransmitters provide the predominant goblet cell stimulation (Diebold et al., 2001).
· Storage and
The glycocalyx provide viscosity, and a low surface tension that aids
uniform re-wetting of the hydrophobic ocular surface. Therefore it
renders the ocular surface polar and thus wettable (Govindarajan and
b. The glycocalyx:
lower surface tension, which produces the smooth, uniform, optically advantageous
properties of the tear film.
surface wettability by overcoming corneal epithelial hydrophobicity.
reservoir for immunoglobulins.
epithelial surface; it spreads rapidly to heal defects and cover foreign
bodies. It play a role in preventing adherence and
interaction of microbes, debris, and inflammatory cells with the epithelium.
lubrication, allowing the palpebral and bulbar conjunctiva to slide over each
other with minimal trauma during blinking or eye movements (Cher, 2008).
Fig. 1.5: Transmission electron
micrographs of the surface cell layer of the cornea showing corneal epithelial
microvilli with transmembrane mucins that extend into the mucin/aqueous glycocalyx
(Blalock et al., 2008).
The glycocalyx is a membrane-bound network of mucins attached to
the apical microvilli of corneal and conjunctival epithelial cells. The Corneal and conjunctival epithelium express transmembrane
mucins (MUC 1, 2, 4), which anchor the aqueous/mucin glycocalyx to the cell
surface Fig.1.5 (Blalock et al., 2008).
b. Water and electrolytes: it is
secreted by conjunctival goblet and non – goblet epithelial cells (Watanabe,
a. Mucins (glycoproteins): it
is secreted by conjunctival goblet cells. Mucins are high molecular weight
proteins with many carbohydrate side groups. It maintain a high water content
and confer a viscous texture to mucous.
The mucus layer is the closest to the cornea with thickness about
0.02-0.04 µm. It consists of:
· Composition and
Layer and Glycocalyx:
§ The glands of Wolfring are in the tarsal conjunctiva
situated near the upper border of superior tarsal plate in upper lid and
occasionally lower lid (Doughty and Bergmanson, 2003).
§ The glands of Krause are mostly forniceal; 40-42 in the
superior and 6–8 in the inferior fornix.
non-reflex tear production is from the accessory lacrimal glands of Krause and
Wolfring that open onto the conjunctival surface (Takahashi
et al., 2013).
(iii) Lacrimal gland secretion: The acinar secretory cells produce a primary
secretion that is similar to plasma; this is modified by the epithelial cells
lining the ductules which secrete additional K+ and Cl?. At
low flow rates, this is hypertonic to plasma; at high flow rates, it is
isotonic. Acinar cell secretion is maintained by basolateral Na+
/ K+ ATPase pump activity (Dartt, 2009).
Microscopic picture: The lacrimal gland is a lobulated tubuloacinar gland.
Multiple acini drain into progressively larger tubules which drain into the superolateral
fornix (Dartt, 2009). The acini consist of columnar secretory cells.
Myoepithelial cells basal to secretory cells have contractile properties to
help express secretions. The acini are surrounded by an interstitium with a
dense network of capillaries and immunological cells (macrophages, eosinophils,
lymphocytes, and plasma cells). Plasma cells produce IgA that is secreted in
tears (Knop and Knop, 2005).
Fig 1.4: Structure of the main lacrimal gland (Lorber, 2007).
anatomy: The lacrimal gland is found superiorly in the
anterolateral superior orbit. It is divided into a superior orbital part and an
inferior palpebral part. These are continuous with each other around the
lateral horn of the levator aponeurosis (Obata, 2006) (Lorber, 2007).
a) Main Lacrimal gland: Fig 1.4
95 % of aqueous
volume is replenished from main lacrimal gland secretion while 5 % from the
accessory glands of Krause and Wolfring.
The aqueous contains nutrients and waste products important in
corneal and conjunctival metabolism. Regulation of tear pH is essential for
optimal epithelial cell function and survival. Tear
pH is lowest on awakening due to overnight buildup of acid by-products while on
eye opening, it rapidly corrects due to loss of CO2 and remains stable through the day due to
buffering systems (Ohashi et al., 2006).
Electrolyte concentration of this layer is similar to that of
serum, resulting in an average osmolarity of 300 mOsm/L. Normal osmolarity is essential to maintain cellular volume, enzymatic
activity, and cellular homeostasis. Matrix Metalloproteinases, particularly
MMP-9, serve an important role in wound healing and inflammation, and are
substantially up-regulated in dry eye syndrome. Tear osmolarity is lower
during closure overnight due to reduced evaporative loss while during the day, it
stabilizes. Tear osmolarity correlates highly with
dry eye syndrome and will likely be increasingly utilized as a metric for
diagnosis and classification of the disorder (Lemp et al., 2011).
Table 1.1: Tear
film aqueous layer proteins (Klenkler et al., 2007).
The aqueous component comprises the major portion (90%) of the tear
film, with a total thickness about 6 µm. The aqueous portion of the
mucin/aqueous gel contains proteins, electrolytes, oxygen, and glucose. Some of
the major protein constituents of the aqueous are outlined in Table 1.1 (Klenkler et al., 2007) (Dartt, 2011).
· Composition and
B. Aqueous Layer:
secretion occurs on blinking due to contraction of the muscle of Riolan.
Increased blink rate and force might increase the volume of secreted meibum (McMonnies, 2007).
glands have androgen and estrogen receptors. Meibomian gland secretion is
influenced by lipid synthesis, which is regulated by circulating androgen and
estrogen levels. Androgens appear to stimulate lipid synthesis and secretion by
meibomian gland (Sullivan et al., 2002).
b. Hormonal regulation:
glands are innervated richly by sensory, sympathetic, and parasympathetic
nerves. However, how these nerves regulate meibum secretion is unknown (Seifert and Spitznas, 1996).
a. Neural regulation:
Regulation of meibum secretion:
spilling over the eyelid. This occurs because the skin’s sebum has mostly non-polar
lipids and tends to repel meibum which has a greater proportion of polar lipids
(Pappas, 2009) (Butovich, 2011).
(iii) Prevents contamination with skin lipids (which can destabilize the
evaporation of underlying aqueous.
It is primarily secreted from meibomian glands with additional
contributions from the glands of Moll and Zeiss (McCulley
and Shine, 2003) (Bron et al., 2004).
The lipid layer forms the outermost surface of the tear film; thus,
it is the portion of the tear film that is in direct contact with the
atmosphere. It forms a surface film that is about 0.1 µm thick. The lipid layer
consists of a heterogeneous mixture of lipids like
hydrocarbons, sterol esters, waxy esters, triglycerides, free cholesterol, free
fatty acids, polar lipids and proteins. Polar lipids including
ceramides, cerebrosides and phospholipids form the inner surface of the
lipid layer, with their charged side facing aqueous while
the non-polar lipids form the anterior lipid–air interface (Shine and
McCulley, 2003) (Butovich, 2011).
Fig.1.3: Schematic representation of the structure of the tear film. Left:
Classic: Discrete three layered structure. Contemporary: An aqueous–mucin
glycocalyx gel with a mucin gradient has been proposed (Hosaka et al., 2011).
The ocular surface requires a dynamic yet stable tear
film to meet the environmental, immunologic and optical challenges presented
to it. For decades, a discrete three-layer model was accepted, consisting of an
anterior lipid layer to provide protection from evaporation; an aqueous component
that provided the largest part of tear film volume; and a mucin layer that
provided protection and lubrication of the corneal and conjunctival epithelium.
A more recently proposed model consists of a mucin/aqueous glycocalyx gel that
comprises most of the tear film volume with an external protective lipid layer
to resist evaporative forcesFig.1.1(b) Fig. 1.3 (Hosaka et al.,
of the Tear Film:
Fig 1.2: Slit lamp photographs with fluorescein staining of (a) a normal
subject (b) a dry eye patient (Murube, 2009).
Tear production is approximately 1.2 microliters per
minute with a total volume of 7–10 ?L and a turnover rate of 16% per minute.
70–90 % reside in the upper and lower tear menisci. These are curvilinear
collections of tears that line the ocular surface immediately adjacent to the
lid margins. Tears are continually replenished from the inferior tear meniscus
by blinking (Palakuru et al., 2007).This counters the forces of
gravity and evaporation on the volume of the precorneal tear film and protects
corneal and conjunctival epithelial cells from the shear forces exerted by the
eyelids during blinking. The tear film drains via the menisci through the
lacrimal puncta which are opposed to the globe near the inner canthus (Wang
et al., 2006). Tears are also stored in the upper and lower
conjunctival cul-de-sacs (fornices). Normal basal tear production rate is 1–2
?l/min; in contrast the reflex tear rate is >100 ?l/min (Murube, 2009).
Tear film thickness, as measured by interferometry, is 6.0 ?m ± 2.4 ?m in
normal subjects and is significantly thinner in dry eye patients with measured
values as low as 2.0 ?m ± 1.5 ?m Fig. 1.2 (Hosaka et al., 2011).
and Flow of Tears
Fig. 1.1: The Tear Film (a) Distribution (b) Structure (Garreis et al., 2010).
§ It contains antibacterial constituents (e.g., secretory
immunoglobulin A (sIgA), lysozyme, lactoferrin) and has a low pH to maintain an
antibacterial environment (McKown et al.,
2009) (Garreis et al., 2010).
§ The tear film is the first line of defense against
§ Nutrients (e.g., glucose) pass from the conjunctival
vessels to the cornea via the tear film (Chhabra
et al., 2009).
§ Oxygen dissolves in the tear film from air, supplying
the avascular cornea.
§ Blinking flushes debris and exfoliated cells from the
ocular surface out through the tear duct (Gipson, 2007).
§ The tear film adheres to the bulbar and palpebral
conjunctiva ensuring well- lubricated surface.
§ The air-tear film interface is the most powerful
refractive surface of the eye.
§ The tear film provides a smooth, regular optical
surface for refraction by filling corneal irregularities (Montés-Micó
et al., 2010).
The tear film is a highly ordered fluid layer lining the
cornea and bulbar and palpebral conjunctiva Fig. 1.1(a). Abnormal
constitution or volume impairs the ocular surface and may reduce corneal
transparency (Tiffany, 2008). The tear film has four main
functions: optical, mechanical, nutritional, and defensive