Ординатура / Офтальмология / Английские материалы / Ocular Periphery and Disorders_Dartt, Bex, Amore_2011

MGD and posterior blepharitis. Many studies have looked at the association between blepharitis or chronic blepharitis and dry eye. However, the conclusions drawn about the association depend on the definitions being used in the study. Although MGD occurs in nearly three-quarters of patients with chronic blepharitis, it also occurs in approximately 20% of people with normal tear function. In MGD, the orifices of the meibomian glands are blocked, reducing the secretion of meibomian lipids onto the ocular surface. Regular warm compresses help to open the orifices and allow normal lipid secretion. Some patients with chronic blepharitis have similar symptoms to those of dry eye and are prescribed tear lubricants for palliative purposes, that is, they do not resolve the blepharitis.

Chalazion

The blockage of the meibomian glands can lead to formation of a chalazion (Figure 7). This is a cyst on the eyelid that is normally sterile and composed of a lipid granuloma. It looks similar to a stye, which is caused by an infected sebaceous gland of the eyelash, but can be easily distinguished clinically because a chalazion is painless and develops gradually, whereas a stye is always painful and forms over a few days. For both conditions, warm compresses are recommended. In extreme cases of chalazion, it is surgically incised and the granulomatous material is removed by curettage. Antibiotics are often prescribed to treat a stye.

Surgical Damage

Treatment of trachoma is a common cause for surgical damage to the meibomian gland. Trachoma, a leading cause of blindness, is an infectious disease of the palpebral conjunctiva that leads to the eyelids folding inward (entropion), causing the lashes to rub against the cornea. The lids, and hence the meibomian glands and ducts, are cut to relieve this condition. It is yet to be determined whether this compromises the functionality of the outer lipid layer of the tear film. Other surgical procedures such

as correction of lid malpositioning, particularly ptosis (drooping of the upper eyelid) and genetic entropion, can also sometimes require the cutting of the meibomian glands.

Contact Lenses and the Lipid Layer

Anomalies of the lipid layer, in themselves, are not a deterrent for contact lens wear. Lipids or proteins or both are deposited on contact lenses during wear. These deposits can block the small pores of the contact lenses, which are essential for the passage of air to the cornea for its metabolism. It is impossible to ascertain beforehand how long a contact lens needs to be worn for it to be unsuitable for an individual as the amount and pattern of lipid deposition depends on the composition of the ocular lipids, which can vary between people, and the specific material the lens is made from. Contact lens cleaning agents are designed for the specific type of contact lens and normally contain a surface active agent that removes lipid deposits.

See also: Dry Eye: An Immune-Based Inflammation; Ocular Mucins; Tear Film.

Further Reading

Bron, A. J., Benjamin, L., and Snibson, G. R. (1991). Meibomian gland disease, classification and grading of lid changes. Eye 5: 395–411.

Bron, A. J., Tiffany, J. M., Gouveia, S. M., Yokoi, N., and Voon, L. W. (2004). Functional aspects of the tear film lipid layer. Experimental Eye Research 78: 347–360.

Butovich, I. A., Millar, T. J., and Ham, B. M. (2008). Understanding and analysing Meibomian lipids – a review. Current Eye Research 33: 405–420.

Glasgow, B. J., Marshall, G., Gasymov, O. K., et al. (1999). Tear lipocalins: Potential scavengers for the corneal surface. Investigative Ophthalmology and Visual Science 40: 3100–3107.

Goto, E. and Tseng, S. C. G. (2005). Kinetic analysis of tear interference images in aqueous tear deficiency dry eye before and after punctual occlusion. Investigative Ophthalmology and Visual Science 44:

1897–1905.

Gouveia, S. M. and Tiffany, J. M. (2005). Human tear viscosity: An interactive role for proteins and lipids. Biochimica et Biophysica Acta 1753: 155–163.

Holly, F. J. (1973). Formation and rupture of the tear film. Experimental Eye Research 15: 515–525.

Hykin, P. G. and Bron, A. J. (1992). Age related morphological changes in lid margin and Meibomian gland anatomy. Cornea 11: 334–342.

Jester, J. V., Nicolaides, N., and Smith, R. E. (1981). Meibomian gland studies: Histologic and ultrastructural investigations. Investigative Ophthalmology and Visual Science 20: 537–547.

Mathers, W. (2004). Evaporation from the ocular surface. Experimental Eye Research 78: 389–394.

McCulley, J. P. and Shine, W. (1997). A compositional based model for the tear film lipid layer. Transactions of the American Ophthalmological Society 55: 79–93.

Millar, T. J., Tragoulias, S. T., Anderton, P. J., et al. (2006). The surface activity of purified ocular mucin at the air–liquid interface and interactions with meibomian lipids. Cornea 25: 91–100.

Nagyova, B. and Tiffany, J. M. (1999). Components responsible for the surface tension of human tears. Current Eye Research 19: 4–11.

Sullivan, D. A., Sullivan, B. D., Evans, J. E., et al. (2002). Androgen deficiency, Meibomian gland dysfunction, and evaporative dry eye.

Annals of the New York Academy of Science 966: 211–222. Tiffany, J. M. (1995). Physiological functions of the Meibomian glands.

Progress in Retinal and Eye Research 14: 47–74.

Los Angeles, CA, USA

ã 2010 Elsevier Ltd. All rights reserved.

Glossary

Acinus – Originating from the Latin word grape, it refers to the sac-like ending of a secretory exocrine gland. Endocytosis – The process of internalization of plasma membrane as well as membrane-bound constituents and extracellular fluid by invagination of the plasma membrane, budding of the membrane vesicle, and its movement to the interior. Different types of endocytosis are known, including clathrin-mediated and caveolar endocytosis. Exocytosis – The process by which a cell releases the contents of secretory vesicles to the extracellular environment by fusion of secretory vesicles with a plasma membrane domain.

Motor proteins – Mechanochemical proteins that utilize the energy of ATP hydrolysis to generate motive force along a polar surface, typically an actin filament or a microtubule.

Rab proteins – Small GTP-binding proteins that utilize the GTP binding and hydrolysis cycle to trigger protein on and off states, and which serve as molecular zip codes to specify the accurate sorting and targeting of membranes.

SNARE proteins – Proteins associated with donor and acceptor membranes which associate to form a fusion pore, allowing the contents of opposing membrane vesicles to intermingle, or allowing the extrusion of membrane-encapsulated contents to the cell exterior.

Transcytosis – The process by which macromolecules are transported through a polarized cell.

trans-Golgi network – A post-Golgi processing compartment responsible for the accurate segregation of contents into membrane vesicles destined for regulated exocytosis, constitutive exocytosis, or for targeting to intracellular membrane compartments.

Anatomy of the Main Lacrimal Gland

The human main lacrimal gland, located laterally above the eye, measures approximately 20 12 5 mm and has an almond-like shape. The major part of the gland, designated as the orbital portion, or the intraorbital gland,

is located in the shallow lacrimal fossa of the frontal bone, while the smaller palpebral, or extraorbital portion, which is separated from the orbital portion by the lateral horn of the levator palpebrae muscle, is located above the temporal segment of the superior conjunctival fornix. In contrast, the mouse and rat have two pairs of lacrimal glands including a small orbital gland which is located laterally beneath the upper lid and a larger extraorbital gland which is located ventral and anterior to the eye. The rabbit lacrimal gland is unusually large and is comprised of a larger portion (4 cm) located below the eye and a smaller portion (0.5 cm) located above the eye. The lacrimal gland is constituted largely (80%) of acinar epithelial cells organized within the tubuloacinar units that are arranged into multiple globuli surrounded by fibrovascular septa. The remaining 20% of the mass of the lacrimal gland is composed of ducts, nerves, myoepithelial cells, leukocytes, and connective tissue. A schematic diagram showing the positioning of the gland relative to the ocular surface and the organization of several of the cell types within the gland is shown in Figure 1.

Cell Types within the Lacrimal Gland

Acinar Cells

The acinar epithelial cells within the lacrimal gland are triangular-shaped cuboidal cells organized in single cell layers in clusters with a narrow microvillus-covered apical domain oriented toward a central lumen and a more extensive basolateral domain which faces the tissue interstitium. Tight junctions near the apices segregate these two domains and result in polarization of the cells, which also are cytoplasmically coupled through gap junctions, thus making the acinus a single functional unit. The apical side of the cell is enriched in numerous large (1–2 mm) secretory granules or vesicles, containing proteins released upon cell stimulation, while the Golgi apparatus and endoplasmatic reticulum compartments are located more toward the basolateral side adjacent to the basolateral nucleus. Figure 2 shows the characteristic distribution of secretory vesicles and other organelles within an acinar cell from mouse lacrimal gland.

Ductal Cells

The lumena of several acini come together to form a duct; each duct merges with others into gradually larger ducts

Figure 1 Schematic diagram showing the positioning of the human main lacrimal gland relative to the ocular surface and the organization of several of the cell types within the gland. AM, apical membrane; BM, basolateral membrane; CNS, central nervous system; D, duct; L, lumen; LG, lacrimal gland; ME, myoepithelial cell; N, nucleus; NE, nerve ending; OS, ocular surface; SV, secretory vesicle.

N

SV

L

2000 nm

Figure 2 Transmission electron micrograph of mouse lacrimal gland. N, nucleus; L, lumen; SV, secretory vesicle.

that finally, in humans, drain into 6–12 major ducts with openings in the upper lateral fornix. In rat and mice, the ducts from the extraorbital gland join to a single major duct that then joins the duct from the intraorbital gland before it empties onto the conjunctiva in the lateral canthus of the eye. In rabbit, a single duct each forms from the upper and lower portions of the gland, which empty onto the conjunctiva of the upper and lower lids, respectively, near the temporal angle. The ducts are formed by one to two layers of cuboidal epithelial cells. Similar to the acinar cell, the ductal epithelial cells of small ducts are polarized by tight junctions in the apical area. However, in these cells, the Golgi and endoplasmic reticulum are located more apically, and secretory vesicle content is lower. Interlobular ducts are embedded with and supported by perivasculoductal connective tissue containing associated structures such as nerve fibers, capillaries, and mast cells and a dense population of fibroblast-associated collagen fibrils.

Myoepithelial Cells

The acini are surrounded by stellate-shaped myoepithelial cells with long slender processes. The myoepithelial cells not only exhibit characteristics of other epithelial cells, such as expression of cytokeratin, but also exhibit properties of smooth muscle cells such as expression of a-smooth muscle actin. The exact role of the myoepithelial cells in the regulation and maintenance of the lacrimal gland remains unclear, but it has been shown that they express receptors for neurotransmitters, suggesting that they play a role in facilitating the secretion from the lacrimal gland. It is also likely that an important role is to support and maintain the structure of the lacrimal gland.

Bone-marrow-derived Cell Population

The lacrimal gland is a part of the mucosal-associated lymphoid tissue (MALT). The bone-marrow-derived cells in the lacrimal gland are mainly immunoglobulin A (IgA)-producing plasma cells and T and B lymphocytes, but macrophages and mast cells are also present. The bone-marrow-derived cells are clustered into lymphoid follicles scattered in the stroma surrounding the acini.

Innervation of the Lacrimal Gland

The lacrimal gland is innervated by parasympathetic, sympathetic, and sensory nerves. Parasympathetic nerves originate in the lacrimal nucleus of the pons and travel along the nervus intermedius, the deep and superficial petrosal nerves, and the vividian nerve before they synapse in the pterygopalatine ganglion. The postganglionic parasympathetic fibers can take different routes to the lacrimal gland. They leave the ganglion through the pterygopalatine nerves but can also reach the lacrimal gland via the maxillary portion of the trigeminal, the zygomatic, or the lacrimal nerves. Parasympathetic fibers can also travel along a branch of the middle meningeal artery to join the ophthalmic or lacrimal artery en route to the lacrimal gland. Sympathetic nerves originate from the superior cervical ganglion and travel along with the parasympathetic nerves through the pterygopalatine ganglion, reaching the lacrimal gland through the lacrimal branch of the zygomatic branch of the maxillary trigeminal nerve that joins the ophthalmic branch of the trigeminal nerve. The sensory nerves innervating the lacrimal gland carry sensory information from the gland through the ophthalmic branch of the trigeminal nerve to the trigeminal ganglion.

The parasympathetic nerves, being the most abundant, regulate the lacrimal gland mainly through the release of neurotransmitters, acetylcholine and vasoactive intestinal

peptide (VIP), with a possible co-secretion of nitric oxide (NO). Acetylcholine activates M3 muscarinic receptors located in the basolateral membrane of the lacrimal cells, while VIP binds to VIP receptors that are similarly located. The sympathetic nerves exert their effects on the lacrimal gland through release of norepinephrine that binds to a- and b-adrenergic receptors, and possibly through neuropeptide Y receptors also located at the acinar cell basolateral membranes. The sensory nerves release substance P and calcitonin gene-related peptide. Not every individual acinar cell is independently innervated; rather cells that are not directly innervated can respond to stimulation of neighboring cells due to the intercellular gap junctions that connect the cells. The density of synapses within each acinus varies according to the species: in the rat, orbital glands fewer than 15% of acinar cells have an adjoining nerve fiber in contrast to the mouse orbital glands where close to 100% of the cells have an adjoining nerve fiber.

Blood Supply

The main blood supply to the lacrimal gland is not only through the lacrimal artery, a branch of the ophthalmic artery, but it also receives minor contributions from the infraorbital and the meningeal arteries. The veins mainly follow the same pathways as the arteries inside the orbit and drain into the superior ophthalmic vein.

Contents of Lacrimal Fluid

The tear film consists of three layers: a mucus layer located directly above the ocular surface epithelium, an aqueous layer, and a thin external lipid layer. The lacrimal fluid produced by the main lacrimal gland constitutes the major part of the aqueous layer of the tear film, to which the accessory glands of Krause and Wolfring and the ocular surface epithelium also contribute, in a minor fashion. Although the major part of the aqueous layer is water, it also contains electrolytes and a high concentration of proteins. Human tear fluid, for instance, has a protein concentration of about 8 mg ml 1. Although the three layers of the tear film largely originate from different sources, these sources can contribute in part to each layer; therefore, it is hard to determine the origin of a specific protein. Recently, researchers identified 419 different proteins in human tear fluid; however, many of these may not have an active function in the tear fluid but are simply debris shed from epithelial cells. Three proteins constitute 80% of the total protein within the aqueous tear film, that is, lipocalin, lysozyme C, and lactoferrin. The functions of the different proteins in the tear fluid are varied. For instance, many proteins contribute

to the antimicrobial properties of the tear fluid. Secretory IgA and cytokines are involved in immune responses, while others such as lysozyme C and lactoferrin provide a more direct defense against bacteria. The novel protein, lacritin, acts as a mitogen in corneal regeneration. Other proteins in the lacrimal fluid are involved in diverse activities in wound healing, blood coagulation, and oxidative stress reduction – all functions essential to maintain a healthy ocular surface. The protein pattern of both active and inactive proteins in the tear fluid can reflect disease states, including diabetes, dry eye, and cystic fibrosis.

Mechanisms of Protein Secretion in the

Lacrimal Gland

The acinar cells of the lacrimal gland are professional excretory cells that engage in several types of secretion that collectively contribute to the lacrimal fluid. Protein secretion at the apical membrane into lacrimal fluid can be subdivided into several types: regulated exocytosis, constitutive exocytosis, and transcytosis. Regulated exocytosis is a process in which proteins destined for a particular plasma membrane domain are sorted into secretory vesicles after their biosynthesis within a post-Golgi sorting compartment called the trans-Golgi network. These secretory vesicles mature and migrate toward the site of release, where they are stored until the appropriate signal triggers their movement and fusion with the acceptor membrane domain. Examples of content proteins thought to be released from the lacrimal acini in animal model systems through regulated exocytosis at the apical plasma membrane include peroxidase and b-hexosaminidase. Constitutive exocytosis occurs, similarly, as components for release to the exterior of the cell are sequestered into vesicles at the trans-Golgi network that are immediately targeted to the acceptor membrane. Unlike regulated exocytosis, constitutive exocytosis is not reliant on extracellular activation of receptors by a ligand (such as a hormone or a neurotransmitter) to elicit the event. Although both forms of exocytosis have been observed in lacrimal acini, most studies have focused on the role of regulated exocytosis in the release of proteins at the apical plasma membrane of the acinar cell into lacrimal fluid. The secretory vesicles in the lacrimal gland acinar cells are generally larger (1–2 mm) and considerably more heterogeneous in both size and content compared to vesicles in other exocrine glands such as exocrine pancreas and the salivary gland. The spectrum of proteins secreted from the lacrimal acinar secretory vesicles appear to span a greater functional range than the spectrum release from other exocrine tissues as well, including nutrient and protective factors as well as factors that protect the mucosal surface from pathogens. This is an area of very active research since there are an unusually large number of

proteins of unknown function in the lacrimal gland secretory proteome.

Transcytosis is a process in which material internalized at the basolateral membrane is recruited into vesicles by endocytosis, followed by the movement of these vesicles to apically located compartments, and ultimately their targeting to the apical plasma membrane for release. Major cargo known to be carried through this pathway includes dimeric IgA, through association with the polymeric IgA receptor. Although not specifically characterized in lacrimal acini, other abundant tear proteins, including albumin and transferrin, are known to be transported through transcytotic pathways in other epithelial cells, suggesting that these proteins may be comparably transcytosed into lacrimal fluid by lacrimal acini.

Regulated exocytosis and transcytosis utilize a number of different processes, globally referred to as membrane trafficking, to achieve the unidirectional transport of cargo-laden vesicles over short and long distances followed by their targeted release. Several of these processes have been studied in acinar cells, including cytoskeleton and motor proteins, rab proteins, and soluble N-ethylmaleimide-sensitive factor attachment protein receptors or SNARE proteins. All mammalian cells contain filamentous structures collectively referred to as the ‘cytoskeleton’, which includes actin filaments, intermediate filaments, and microtubules. Each of these structures is formed from individual subunit proteins that exist in equilibrium with polymeric assemblies. Cytoskeletal filaments are critical in maintaining the integrity of cell shape as well as conferring cellular polarity or asymmetry, a function critical in aiding the movement of materials to different membrane domains in a polarized cell. Filamentous actin and microtubules, in particular, participate in several capacities in the movement of membrane vesicles to the apical plasma membrane, where the release of proteins into lacrimal fluid takes place. Microtubuleand actindependent membrane transport events can be facilitated either by the use of compressive force associated with cytoskeletal assembly to physically compress or direct membrane traffic, and/or by the use of these polymers as tracks which support the movement of motor proteins which carry membrane vesicles to specific destinations.

In lacrimal acini, actin filaments are localized in a dense network below the apical membrane called the subapical actin network, and this network is also present to a lesser extent below the basolateral membrane. Beneath the subapical actin, the ends of microtubules are anchored. Microtubules extend from the subapical region to the basolateral membrane. Both actin filaments and microtubules sustain aspects of protein secretion in lacrimal acinar cells. When microtubules are disrupted in acinar cells using the agent, nocodazole, stimulated protein secretion is reduced because the microtubule scaffolding required for vesicle motility has been disrupted. Other studies have

suggested that a particular motor protein, cytoplasmic dynein, is required for the movement of membrane vesicles involved in secretory vesicle maturation and, possibly, transcytotic vesicle transport, to the subapical cytoplasm.

The subapical actin cytoskeleton plays complex roles in lacrimal acinar secretion. With its location immediately beneath the apical plasma membrane in a dense network, it poses an intracellular barrier for vesicle fusion to the apical membrane. For fusion to occur, this actin barrier must be disassembled to allow access of large secretory vesicle to the apical plasma membrane. Recent work has shown in fact that regions of the actin cytoskeleton located immediately beneath the regions of apical plasma membrane do disassemble, but that actin filaments also reassemble and contract around the base of multiple fusing secretory vesicles. The force generated through compression and retraction of actin filaments toward the apical membrane aids in compound fusion and content extrusion from the fusing vesicles. Regulated exocytosis can therefore be further characterized in lacrimal acini into a type known as multivesicular exocytosis. Two specific actin-dependent motors have been implicated so far in this actin remodeling and compound fusion of activated secretory vesicles, a conventional myosin motor known as nonmuscle myosin 2 and an unconventional myosin motor known as myosin 5c, with the possibility that other members of the myosin motor superfamily may also participate in this complex process.

Other major membrane trafficking effectors that have been implicated in acinar cell protein secretion include rab proteins. Rabs are major effectors of all intracellular steps of membrane trafficking and fusion in the eukaryotic cell, serving as the molecular address code on donor membrane vesicles which specify the acceptor compartment destination. GTP binding and hydrolysis serves as the on/off switch that activates these proteins. Specific rabs are localized to distinct compartments, conferring identity to these compartments. For instance, rab3D is enriched in secretory vesicles in lacrimal acinar cells and appears to regulate compound fusion of these vesicles. Other data suggest that rab27 isoforms also participate in the maturation and fusion of secretory vesicles during regulated exocytosis in lacrimal acini. By analogy with other systems, rab4 and rab5 isoforms are likely to participate in early events in acinar transcytosis, specifically basolateral endocytosis and sorting, while rab 11 isoforms are enriched in apical endosomes and may facilitate terminal transcytotic traffic of materials destined for the lacrimal fluid.

Specific types of SNARE proteins are located on donor and acceptor membranes and interact to form fusion pores which allow membrane contents to mingle or secretory vesicle contents to be extruded to the cell exterior. Several types of SNARE proteins have been demonstrated in lacrimal acini. Regulated exocytosis of secretory vesicles in acini is thought to use both vesicle-associated

Figure 3 Protein secretion in lacrimal acinar cells at the apical membrane. (a) Depicts vesicles participating in the transcytotic pathway from the basolateral membrane (BM) to the apical membrane (AM) as shown in blue vesicles. Depicted as well is the maturation and movement to the AM of secretory vesicles after budding from the trans-Golgi network (TGN) as shown in red vesicles. Initially both transcytosis from the BM and movement from the TGN are reliant on microtubule-based motor proteins. As the vesicles move to the actin-rich subapical region, unconventional myosin motors become important in actin-based movement through the subapical actin network.

(b) Depicts multivesicular exocytosis after stimulation with secretogogs such as carbachol. Unconventional myosins such as myosin 5c have been shown to have an important role in the reorganization of actin filaments around clusters of secretory vesicles primed for fusion. Actin and conventional myosins then work together to compress the fusing secretory vesicles and to promote content extrusion. Rab and SNARE

membrane protein 2 (VAMP 2) and VAMP 8 on donor membranes, and syntaxin 2 and SNAP23 on acceptor membranes. Figure 3 shows the organization of the two major protein secretory pathways that contribute proteins to lacrimal fluid: regulated exocytosis and transcytosis, as well as some of the effectors within each pathway.

Morphological analysis of ductal epithelial cells has revealed the presence of large secretory vesicles, presumably containing additional constituents destined for release into the lacrimal fluid. However, due to limitations in the ability to isolate these individual cells and conduct cellular investigations into the membrane trafficking mechanisms, little is known about the precise mechanisms involved in ductal cell exocytosis and transcytosis.

Mechanisms and Regulation of Electrolyte and Water Secretion by the Lacrimal Gland

Acinar Cells

The lacrimal fluid is hypertonic due to a high Cl and Kþ content, whereas the levels of Naþ, HCO3 , and Ca2þ are similar to the plasma levels. The electrolyte concentration of the lacrimal fluid is however not static, but varies with the flow rate to become more isotonic with an accelerated flow rate. Fluid secretion is an osmotic process driven by ion movement through the membrane of the acinar cells. Parasympathetic stimulation of the acinar cells triggers an acute increase of cytosolic Ca2þ and adenosine 30,50- cyclic monophosphate (cAMP) which opens Cl channels in the apical membrane, resulting in a movement of Cl into the lumen. The increase in cytosolic Ca2þ also activates Kþ channels in the apical as well as the basolateral membrane, causing Kþ to move out of the cell. Naþ follows the flux of Cl and Kþ, moving from the basolateral side toward the lumen traveling through paracellular channels between the cells. To maintain an isotonic secretion, water exits the cell through water-channel proteins called aquaporins. The movement of Cl and Kþ out of the cell is dependent on their electrochemical gradient, that is, the intracellular concentration of these ions must be higher than in the extracellular fluid. This is made possible by ion pumps and co-transporters located in the basolateral membrane. Naþ/Kþ-ATPase transports Kþ into the cell and Naþ out of the cell; the Naþ/Kþ/2Cl co-transporter (NKCC1) moves all three ion types into the cell; and Cl /HCO3 and Naþ/Hþ anti-porters transport Cl and Naþ into the cell and Hþ and HCO3 out of the cell. Figure 4 illustrates the ion channels and transporters present in the lacrimal acinar cell.

proteins participate in the targeting and fusion events in each pathway. It should be noted that the rabs and the SNAREs participating in transcytosis and exocytosis are different for each pathway.

Lumen

Figure 4 Schematic diagram showing the major ion transporters active in the electrolyte and water release from the lacrimal gland acinar cell. Increases in cytosolic Ca2þ and cAMP following neural stimulation opens Kþ and C channels, resulting in an outward flux of these ions. Naþ/Kþ-ATPase transports

Kþ into the cell and Naþ out of the cell, the Naþ/Kþ/2Cl co-transporter moves all three ion types into the cell, and Cl /HCO3 and Naþ/Hþ anti-porters transport Cl and Naþ into the cell and Hþ and HCO3 out of the cell.

Ductal Cells

Due to difficulties in specifically isolating the ductal cells, the ion transport mechanisms have not been extensively studied. However, it has been hypothesized that the water and ion transport events continue as the lacrimal fluid travels through the ductal system in a pattern comparable to that in the acinar cells. Recently, several approaches to study the ductal cells have been developed, including microdissection and culturing of individual ducts and laser capture microdissection of individual ductal cells. In these studies, some of the most common acid/base transporters were characterized in the ductal cells. The same work also showed that the ion transport in the ductal cells can be regulated by parasympathetic neurotransmitters. Furthermore, studies showed that the lacrimal fluid released from the acinar cells is isotonic, leading to the hypothesis that the ductal cells are responsible for the high Kþ levels in tears. The finding that NaþKþ-ATPase and NKCC1 are expressed at higher levels in the ductal cells than in the acinar cells supports this hypothesis.

Conclusion

Previous and ongoing studies have established many of the functions of the principal cells of the lacrimal gland. The major cell type, the acinar cell, is responsible for the regulated release of proteins, fluid, and electrolytes into

the lacrimal fluid, while ductal cells appear to further modify the electrolyte composition and likely also contribute additional proteins to the aqueous tear film. Some of the signaling pathways have been elucidated that stimulate the production of lacrimal fluid, while some of the molecular mechanisms involved in the fundamental exocytotic and transcytotic events have likewise been elucidated. However, the complexity of the signaling and membrane trafficking events even in lacrimal acinar cells, the best-studied cell type in this complex organ, means that considerable work remains to be done. Although some insights regarding changes in signaling and membrane trafficking pathways that result in altered production of lacrimal fluid have been obtained in dry eye disorders, considerable additional work is required in order to truly understand the etiology of these disorders. In some studies, changes in tear protein composition have been associated with dry eye disorders, so future challenges also include the identification of tear biomarkers that can be used to diagnose different types of dry eye disorders to aid in determination of the appropriate course of treatment.

See also: Dry Eye: An Immune-Based Inflammation; Innate Immune System and the Eye; Lacrimal Gland Hormone Regulation; Lacrimal Gland Signaling: Neural; Meibomian Glands and Lipid Layer.

Further Reading

Cohen, A. J., Mercandetti, M., and Brazzo, B. G. (eds.) (2006). The Lacrimal System, Diagnosis, Management and Surgery. New York: Springer.

Hodges, R. R. and Dartt, D. A. (2003). Regulatory pathways in lacrimal gland epithelium. International Review of Cytology 231: 129–196.

Jerdeva, G., Wu, K., Yarber, F. A., et al. (2005). Actin and non-muscle myosin II facilitate apical exocytosis of tear proteins in rabbit lacrimal acinar epithelial cells. Journal of Cell Science 118: 4797–4812.

Marchelletta, R. R., Jacobs, D., Schechter, J. E., Cheney, R., and Hamm-Alvarez, S. F. (2008). Myosin Vc facilitates actin filament remodeling and compound fusion of mature secretory vesicles during exocytosis in lacrimal acini. American Journal of Physiology (Cell Physiology) 295: C13–C28.

Pflugfelder, S. C., Beuerman, R. W., and Stern, M. E. (eds.) (2004). Dry Eye and Ocular Surface Disorders. New York: Marcel Dekker, Inc.

Ubels, J. L., Hoffman, H. M., Srikanth, S., Resau, J. S., and Webb, C. P. (2006). Gene expression in rat lacrimal gland duct cells collected using laser capture microdissection: Evidence for Kþ secretion by duct cells. Investigative Ophthalmology and Visiual Science 47: 1876–1885.

Walcott, B., Moore, L., Birzgalis, A., Claros, N., and Brink, P. R. (2002). A model of fluid secretion by the acinar cells of the mouse lacrimal gland. Advances in Experimental Medicine and Biology 506(Pt. A): 191–197.

Wu, K., da Costa, S. R., Jerdeva, G., et al. (2006). Mechanisms of exocytosis in lacrimal gland. Experimental Eye Research 83: 84–96.

Zierhut, M., Stern, M. E., and Sullivan, D. A. (eds.) (2005). Immunology of the Lacrimal Gland, Tear Film and Ocular Surface. New York: Taylor and Francis.

Glossary

CD86 – A co-receptor expressed on the surfaces of antigen-presenting cells. When engaging either of its cognate receptors – CD28 and CTLA-4 – on the surface of T cells, it generates signals essential for T-cell activation and activates signaling cascades within the antigen-presenting cells.

Chemokines – The proteins that promote recruitment of lymphocytes and leukocytes to inflamed tissues and to lymphoid tissues. Hypophysectomy – The surgical removal of the pituitary gland.

Interferon gamma (IFN-g) – A cytokine released primarily by Tcells and natural killer cells that induces T cells to express the TH1 phenotype, activates macrophage to express microbicidal functions, and induces B cells to switch from immunoglobulin

M (IgM) to complement-fixing IgG isotypes.

Interleukin 1 alpha and 1 beta (IL-1a, IL-1b) – The related cytokines released primarily from macrophages, endothelial cells, and epithelial cells that induce inflammatory responses.

Interleukin 6 (IL-6) – A cytokine that promotes inflammatory responses and supports survival of T and B cells.

Interleukin 12a (IL-12a) – A cytokine released in innate immune responses that induces expression of IFN-g and, thereby, promotes the evolution of adaptive responses mediated by TH1 cells. Lactation – The production and secretion of milk. Lactogenesis – The secretory differentiation of the mammary epithelium.

Orchiectomy – The surgical removal of the testes.

Sex hormone-binding globulin (SHBG) – A protein which binds estrogens and androgens. It is produced by the liver and secreted into the blood. Estrogens stimulate its production and androgens suppress its production.

Sodium-potassium-dependent ATPase (Na,K-ATPase) – The sodium–potassium pump enzyme; it generates the chemiosmotic energy essential for lacrimal fluid production by pumping Na+ out of, and K+ into, the cytosol.

Transforming growth factor-beta (TGF-b) –

A cytokine released by T cells and macrophages, as well as by some epithelial cells and mesenchymal cells. Its actions on immune cells include: inhibiting T-cell proliferation and expression of effector

functions; inhibiting B cells from proliferating and inducing them to undergo IgM-to-IgA isotype classswitch recombination; and suppressing macrophage activation. It often exerts antiproliferative or proapoptotic influences on epithelial cells.

Gender-Related Dimorphisms

The lacrimal glands produce most of the aqueous fluid that comprises the milieu exte´rieur sustaining the live, metabolically active cells of the superficial layers of the cornea and conjunctiva, and insufficient production of this fluid and alterations of its composition are associated with dry eye disease. Because dry eye disease is considerably more prevalent among women, it has seemed intriguing that morphological differences can be readily discerned between the acini – that is, the primary secretory structures of the lacrimal glands – of male and female rats. The structural dimorphisms were, some years ago, found to be accompanied by equally striking biochemical and functional dimorphisms. Classic work by Sullivan and colleagues showed that many of the evident dimorphisms are supported by the higher levels of androgens characteristic of males. However, it is taking much longer to learn how, in mechanistic terms, the gender-related dimorphisms might relate to females’ greater predilection for lacrimal dysfunction. Indeed, one of the first functional dimorphisms to be documented appeared paradoxical: basal precorneal tear volume is smaller in intact male rats than in females, and it increases in males after they are castrated – a surgical maneuver that causes the size of the acini to decrease, that is, to become more female like.

One of the products that the lacrimal glands contribute to the ocular surface fluid is secretory immunoglobulin A (sIgA), which is the effector of the adaptive mucosal immune defense against microbial infection. The lacrimal glands of rats exhibit several readily quantified dimorphisms relating to the production and secretion of dimeric IgA (dIgA). The stromal spaces of the lacrimal glands of male rats are populated by larger numbers of dimeric IgA-secreting plasmacytes. Whole gland extracts contain greater masses of dIgA. Glandular epithelial cells express higher levels of the polymeric Ig receptor (pIgR), which mediates uptake of dimeric dIgA at the stromal-facing surface of the lacrimal epithelium, chaperones it through

the cells’ transcytotic apparatus, and provides the secretory component (SC) portion of secretory IgA (sIgA). Lacrimal gland fluid from sexually mature male rats contains both more sIgA and SC.

Sex Steroids

Androgens

As noted above, it was established early on that the androgens support the gender-related dimorphisms of acinar size and of precorneal tear volume, which is presumably related to basal rates of lacrimal fluid production. It was also found that the androgens also support epithelial cell expression of pIgR and secretion of SC. However, androgen effects on the numbers of IgAþ-plasmacytes populating the glands’ stromal spaces varied considerably among individual animals.

Although different laboratories have reported discrepant findings, there is some evidence for the theoretical paradigm that the androgens exert general trophic influences on the glandular epithelium. Hypophysectomizing rats decreases circulating levels of gonadal and adrenal steroids, as well as pituitary hormones (see Box 1). This maneuver reduces the lacrimal glands’ gross weight and their weight as a fraction of total body weight. In some studies, administration of dihydrotestosterone (DHT) did not change lacrimal gland weight appreciably. In other studies, administration of DHT partially reversed hypophysectomy-induced decreases in the total amounts of protein and Na,K-ATPase catalytic activity measured in lacrimal gland lysates. Ovariectomizing female rabbits decreases serum sex steroid levels. This maneuver decreased the total protein and DNA contents of lacrimal gland lysates. Administration of DHT prevented the ovariectomy-induced decreases, and it increased the Na,K-ATPase catalytic activity measured in lacrimal gland lysates. Surprisingly, in view of the role Na,K-ATPase plays in lacrimal fluid production, DHT did not increase the basal rate of lacrimal fluid production. However, it significantly increased the volume of fluid intact glands produced when they were stimulated with cholinergic agonists. These findings make it clear that several independent layers of regulation determine lacrimal fluid production: long-term regulation of the mass of cells comprising the glandular epithelium and of the levels at which the epithelial cells transcribe the genes specifying Na,K-ATPase and other iontransport proteins, and acute regulation – presumably neurally mediated – of the transport proteins’ functional states. A more complex paradigm, however, is needed to account for why the magnitude of the lacrimal gland regression caused by ovariectomy is small compared to the extent of atrophy that nuclear magnetic resonance (NMR) imaging studies have documented in the lacrimal glands of aging females. One possible paradigm is that in mature, but not aged, female rabbits the lacrimal glands compensate for ovariectomy-induced loss of testosterone

by converting the weak androgen, dihydroepiandrosterone (DHEA) – which is produced by the adrenal cortex (see Box 1) – to testosterone and DHT. A second is that the loss of a small trophic influence must be compounded over time before its consequences become evident. A third is that androgens influence other parameters in addition to the cellular mass of the epithelium, and that it is the consequences of loss of those influences that are compounded over time.

In ex vivo studies, DHT increased proliferation in models of acinar cells from rabbit lacrimal glands. Compared to the action of epidermal growth factor (EGF), however, the influence of DHT was relatively modest. When Azzarolo and colleagues tested the hypothesis that androgens support the number of cells in the epithelium by preventing apoptosis, as well as by promoting cell proliferation, they found that epithelial cell apoptosis is relatively rare, but plasmacytes in the glands’ stromal space began apoptosing within an hour following ovariectomy. Moreover, administration of DHT prevented ovari- ectomy-induced plasmacyte apoptosis.

The finding that androgen withdrawal leads to a wave of apoptosis in the lacrimal gland plasmacyte population is one of several that accord with the paradigm that their higher levels of androgens protect men both from Sjo¨gren’s autoimmune dacryoadenitis and from the histopathophysiological syndrome commonly found in aging women by influencing immunophysiological processes within the gland. Indeed, it has been found that administration of DHT suppresses lacrimal gland disease in certain mouse models for Sjo¨gren’s syndrome, and preliminary clinical experiences suggest that androgen supplementation of hormone replacement therapy may improve symptoms and clinical signs in menopausal women with inflammatory autoimmune lacrimal diseases. In the models in which androgen administration is therapeutic, the responses are more pronounced in the lacrimal glands than in other affected organs. This finding led Sullivan and coworkers to propose that the androgens control the expression of critical immunoregulatory paracrine mediators by lacrimal gland epithelial cells. Recent microarray analyses indicate that androgens influence the expression of large numbers of gene transcripts in the lacrimal glands and corneas of mice. Some of the androgen actions might be expected to diminish inflammatory processes. For example, testosterone decreases expression of certain chemokines; interferon (IFN) response factors 4 and 7; caspase-1, which converts the inactive interleukin (IL)-1b precursor to active IL-1b; and the prolactin receptor, which – as discussed below – mediates mitogenic responses both in B and T cells and induces Tcells to express IFN-g. However, other testosterone actions might be expected to enhance inflammatory processes. For example, testosterone increases expression of IL-6; IL-12a; the chemokines CCL1, CCL8, CCL28, CCL5, and CXCL4; CD86; and interferon response factor 5.