Ординатура / Офтальмология / Английские материалы / Dry Eye and Ocular Surface Disorders_Pflugfelder, Beuerman, Elliot Stern_2004

respond to inflammation. Finally, we will provide a synopsis of how these cellular and molecular mechanisms are involved in the disease pathogenesis of perhaps the most common form of chronic ocular surface disease, dry eye syndrome.

A wide array of cells and molecules participate in the natural defense of the ocular surface. From a functional and mechanistic standpoint, it may be helpful to divide these mediators into two groups: (1) those involving the resident mesenchymal cells of the cornea and ocular surface, and (2) those involving the bone marrow or hematopoietic elements that normally reside in these tissues.

II.RESIDENT AND MESENCHYMAL CELLS OF THE OCULAR SURFACE

The immune-mediated responses of the ocular surface are influenced by its unique anatomy and physiology. The ocular surface consists of three distinct anatomical regions, the cornea, the limbus, and the conjunctiva, which function both independently and in concert as specific barriers against microbial, immunogenic, and traumatic insults. Although the conjunctiva and cornea are anatomically proximate and are bathed in the same tear film, their immune responses are distinctly different from each other.

A.The Cornea

The normal cornea (in contrast to the conjunctiva) is considered an “immuneprivileged” site due to the high success rate of corneal transplantation [1], the lack of corneal blood or lymphatic vessels, the expression of Fas ligand on its surface that is thought to be relevant to deleting Fas-expressing effector T cells [2], and its general resistance to expressing cell-mediated immunity such as delayed hypersensitivity responses [3]. Since the incident light that will ultimately reach the retina is first refracted at the preocular tear–air interface, and must travel through a clear corneal medium for minimal distortion on its way to the retina, it is axiomatic that the cornea can ill afford any immune or inflammatory response that is not absolutely necessary. This rationale has been applied to explain the central cornea’s high threshold for manifesting immune responsiveness [4]. Interestingly, peripheral areas of the cornea have distinct morphological and immunological characteristics that render them significantly more prone to inflammatory reactions. For example, unlike the avascular central regions of the cornea, the limbus and the peripheral cornea derive much of their nutrient supply from the capillary arcades, which extend only approximately 0.5 mm into the clear cornea [5]. However, this greater reliance of the peripheral cornea on vascular supply also renders it more responsive to locally or systemically incited

inflammation due to recruitment of cellular and molecular mediators of inflammation from the intravascular compartment.

B.The Conjunctiva

The conjunctiva-associated lymphoid tissue (CALT) is a relatively unique component of the mucosa-associated lymphoid tissue (MALT) system, with the limbus serving as a transition zone between the conjunctiva and cornea. The conjunctiva also has other unique cell populations, including goblet cells, mast cells, and accessory lacrimal glands, all of which are absent from the normal cornea [5]. In addition, the conjunctiva has a luxurious vascular supply as well as a rich lymphatic network, which means that it is much more intimately connected anatomically to the immune system than the cornea. The lymphatics afford easy access of antigen(s) and antigen-presenting cells to lymphoid reservoirs, and the vascular supply allows ready access of cellular and molecular immunological effectors to the conjunctiva.

The conjunctival epithelium, unlike the corneal epithelium, lacks an organized basement membrane and rests loosely on the fibrovascular tissue of the substantia propria (stroma). The substantia propria of the conjunctiva consists of two layers, a superficial lymphoid layer and a deeper fibrous layer. The lymphoid layer is made up of a connective tissue matrix containing a homogeneous-appear- ing population of (mostly T) lymphocytes. They interact with mucosal epithelial cells through reciprocal regulatory signals mediated by growth factors, cytokines, and neuropeptides. However, under normal conditions, no germinal follicles are present in the conjunctiva [6]. In addition to these microanatomical features that facilitate immunogenic inflammation in the conjunctiva, the conjunctival epithelium also expresses a unique cohort of cellular adhesion molecules and surface antigens that regulate the interactions among conjunctival epithelial cells, as well as interactions between these cells and bone marrow-derived immune cells. For example, β-integrin complexes, adhesion molecules known as very late antigens (VLAs), function as receptors for collagen, fibronectin, and/or laminin, which form the extracellular matrix. Flow cytometry has shown that β-integrin VLA-1, -2, and -3 adhesion factors are constitutively expressed by the normal human conjunctival epithelium [7].

The exocrine and immune elements of the conjunctiva are intricately related to one another. Two types of accessory lacrimal glands are present in the conjunctiva: the glands of Krause and the glands of Wolfring. Their structures are similar to that of the main lacrimal gland, all belonging to the mucosa-associated lymphoid tissue system. B cells home to sites adjacent to the lacrimal and accessory lacrimal gland epithelia, producing immunoglobulins (Ig), especially IgA. The IgA in tears is unique in that it is dimerized with a polypeptide known as secretory component, acquired from the adjacent epithelia [8]. Secretory IgA is

relatively resistant to proteolysis and is a poor activator of complement, potentially providing defense without significant cytolytic damage to the epithelial cells.

C.The Limbus

The convergence of the cornea and conjunctiva at the limbus separates two vastly different tissues. The phenotypic and biochemical characteristics of the limbal epithelium are intermediate between corneal and conjunctival epithelium [9]. The substantia propria of the limbus has capillaries, lymphatics, sensory nerve endings, and mast cells that are similar to the conjunctiva. Limbal and conjunctival lymphatics drain to the preauricular and submandibular lymph nodes and facilitate the delivery of antigen-presenting cells (APCs) to these lymph nodes, which can contribute to immunogenic inflammation such as rejection of corneal grafts [10,11]. The eye–lymphatic axis is critically relevant for induction of corneal immunity, which is mediated in large part by the limbus [12]. While the functional significance of lymphatic access by ocular surface cells and antigens is increasingly appreciated, the molecular mechanisms that regulate leuko- cyte–lymphatic interactions, potentially involving vascular endothelial growth factor (VEGF)-associated signaling [13], are just being revealed.

III.BONE MARROW-DERIVED CELLS OF THE OCULAR SURFACE: RESIDENT T CELLS

T cells form the predominant lymphocyte subpopulation of the normal ocular surface, outnumbering B cells by up to 20-fold [14]. Table 1 describes the distribution of T cells and other immune cells on the normal ocular surface. T cells are subdivided into CD4+ and CD8+ cells. CD8+ cells are major histocompatibility complex (MHC) class I-restricted and generally function as classic regulatory T cells in addition to being cytotoxic T cells. CD4+ T cells are MHC class II-restricted, and those with helper function [named T-helper (TH) cells] can be further subdivided into either TH1- or TH2-helper types, depending on the profile of cytokines they secrete. The majority of T cells in the uninflamed conjunctiva are CD8+ cells with a CD4/CD8 ratio of 0.75 [15]. T lymphocytes recognize antigens through two structurally distinct T-cell receptors [16]. The α/β T-cell receptor (composed of α- and β-glycoprotein subunits) is expressed on a majority of peripheral blood T cells and is the predominant T-cell receptor subtype on the normal ocular surface, whereas the γ/δ T-cell receptor is expressed on a small proportion of blood lymphocytes and also on a small fraction of lymphocytes in the conjunctival epithelium [15].

*The majority of corneal lymphocytes are found in the limbus and the perivascular areas of the surrounding superficial limbic conjunctiva.

IV. BONE MARROW-DERIVED CELLS OF THE OCULAR SURFACE: RESIDENT B CELLS

Ocular surface B cells are found primarily in the conjunctival substantia propria, where they may form aggregates in association with the overlying specialized lympho-epithelium [17]. This organized tissue, termed conjunctiva-associated lymphoid tissue, resembles the mucosa-associated lymphoid tissue of the gut and upper respiratory tract, and is found variably in humans. Its presence in the conjunctiva correlates with increasing age and antigenic stimulation [17,18]. The specific role played by conjunctiva-associated lymphoid tissue in ocular surface immune responses remains speculative. While it may be involved in homeostatic processes such as secretory IgA production [19], it may also play a role in disease processes such as allergic conjunctivitis [20] and conjunctival B-cell lymphomas [21]. B cells are seen in substantial numbers in a heterogeneous group of ocular diseases. For example, B cells are found in conjunctival biopsies of Sjögren’s syndrome and Epstein-Barr virus (EBV)-associated infections [22]. Together with mast cells and eosinophils, B-lymphocytes play a critical role in IgE-mediated type 1 hypersensitivity reactions, which underlie a number of ocular surface allergic diseases including seasonal allergic conjunctivitis, vernal

conjunctivitis, and perennial atopic keratoconjunctivitis [23]. In allergy, antigenpresenting cells and allergen activate the TH2 subset of helper T cells, which in turn direct IgE-specific B-lymphocyte proliferation and maturation [24]. IgE complexed with allergen on the surface of mast cells stimulates their degranulation, releasing vasoactive amines, neutrophil and eosinophil chemotactic factors, and arachadonic acid metabolites [24]. These result in conjunctival edema, extensive epithelial damage, and scarring seen in atopic and vernal conjunctivitis [25].

V.RESIDENT ANTIGEN-PRESENTING CELL POPULATIONS

Antigen-presenting cells serve as the principal immune sentinels to the foreign world and can be divided into “professional” and “nonprofessional” types. While the latter are found among nonlymphoid tissues (e.g., vascular endothelial or some tissue epithelial cells), professional antigen-presenting cells, such as dendritic cells, macrophages, and B cells, form an integral part of the immune system and are bone marrow (BM)-derived. The cardinal properties of antigen-presenting cells include their ability to take up, process, and present antigen, to migrate selectively through tissues, and to stimulate and direct T-cell-dependant responses. Expression of MHC class II antigens on antigen-presenting cells, whose primary function is to distinguish between self and non-self, plays an integral role in antigen recognition and presentation. Two populations of bone marrow-derived cells,

(1) macrophages/monocytes and (2) dendritic and Langerhans cells, are the main antigen-presenting cells of the ocular surface immune response.

A.Macrophages

Macrophages are bone marrow-derived monocytic cells that reside in virtually every tissue. They are integral to the innate immune response because of their phagocytosis of foreign material, expression of a variety of surface receptors specific for pathogens or antigens, and secretion of proinflammatory cytokines [26–28]. Macrophages synthesize and secrete a variety of powerful biological molecules, including proteases, collagenase, angiotensin-converting enzyme, lysozyme, interferon (IFN)-α, IFN-β, interleukin (IL)-6, tumor necrosis factor (TNF)-α, fibronectin, transforming growth factor (TGF)-β, platelet-derived growth factor, macrophage colony-stimulating factor (M-CSF), granulocytestimulating factor, granulocyte-macrophage colony-stimulating factor (GMCSF), prostaglandins, leukotrienes and oxygen metabolites. They develop from myeloid progenitor cells, enter the bloodstream as monocytes, and migrate into tissues as macrophages. Their expression of low levels of MHC class II and co-stimulatory molecules enables them to act as antigen-presenting cells, although much less efficiently than dendritic cells [29]. However, resident tissue

macrophages are in general poorly responsive to activation signals [30]. Macrophages also play roles in other processes including immune regulation and suppression, tissue reorganization, and angiogenesis [31].

Previous studies have demonstrated resident tissue macrophages in the conjunctiva, limbus, iris, ciliary body, tunica vesiculos lentis, uvea, and retina [32–38]. Until recently, resident macrophages of the ocular surface were thought to reside in the conjunctiva and limbus only [32,38], whereas the cornea was thought to be devoid of any bone marrow-derived antigen-presenting cells. In the conjunctiva, macrophages are the second most commonly found leukocyte cell type after T cells, with a density of 6.5 cells/mm2 in the tarsal epithelium and 32.2 cells/mm2 in the tarsal substantia propria, with similar numbers in the bulbar conjunctiva [32]. Very recently, resident tissue macrophages have been found by confocal microscopy in the normal cornea of mice, located mainly in the posterior third of the stroma [39,40]. Resident stromal macrophages may provide a critical first line of defense against pathogens that breach the epithelial barrier of the cornea by producing antimicrobial substances, as well as other inflammatory cytokines and chemokines to attract and activate additional macrophages, neutrophils, and dendritic cells.

B.Dendritic Cells and Dendritic Cell Precursors

Langerhans cells were originally described by Paul Langerhans in 1868 in the skin epidermis [41], but their function and origin remained obscure for more than a century. It has now been established that Langerhans cells are a population of dendritic cells that reside in epithelia and mediate antigen presentation [32]. Dendritic cells comprise a heterogeneous group of professional bone marrowderived antigen-presenting cells that include members of different lineages and states of maturation [42,43]. Dendritic cells are now recognized as essential regulators of both the innate and acquired arms of the immune system. They serve a unique role because they are the only antigen-presenting cell able to induce primary immune responses, thereby permitting establishment of immunological memory [44,45]. Dendritic cells were first isolated from lymphoid tissue of mice in 1973 [46], and they have an extraordinary capacity to stimulate naïve T cells via MHC molecules and initiate immune responses, including rejection of organ transplants and formation of T-cell-dependant antibodies [44,47]. Recent studies suggest that dendritic cells also regulate the types of T-cell immune responses [43], play critical roles in the induction of peripheral tolerance [48], and function as effector cells in innate immunity against microbes [49,50].

MHC class II-negative proliferating dendritic cell progenitors from the bone marrow give rise to nonproliferating dendritic cell precursors in the blood. These seed nonlymphoid tissues in a stage referred to as “immature” dendritic cells [44,51]. Immature dendritic cells express negligible amounts of MHC

class II on their surface, lack the requisite accessory (co-stimulatory) signals for T-cell activation, such as CD40, CD80 (B7–1), and CD86 (B7–2), and are characterized by a high capacity for antigen capture and processing, but a low T-cell-stimulatory capability [29,47]. In their immature state, they remain dormant until they are signaled from the extracellular milieu through inflammatory mediators (derived from microbes or distressed bystander cells) to induce a rapid change in function, becoming “activated” or mature. Maturation induces redistribution of MHC molecules from the intracellular endocytic compartments of dendritic cells to the cell surface, which renders these cells ineffective at antigen capture but potent in T-cell stimulation [44,47]. The various functions of dendritic cells in immune regulation depend on the diversity of dendritic cell subsets and lineages and on the functional plasticity of dendritic cells at their immature stage. They are found in a variety of nonlymphoid tissues, including the ocular surface, iris, and ciliary body. Some of the phenotypic characteristics of corneal and conjunctival dendritic cells are unique and are discussed below.

C.Corneal Dendritic Cells

Under nonpathological circumstances, Langerhans cells, a subset of dendritic cells, are the only cells that constitutively express MHC class II molecules in the corneal epithelium [52]. Over the last several decades, the search for corneal anti- gen-presenting cells, largely dependent on their presumed universal MHC class II expression for detection, led to the (now disproven) dogma that Langerhans cells are present in the epithelium of the conjunctiva and peripheral third of the cornea, but are essentially absent in the central corneal epithelium and stroma [52–59]. The consensus was that the adult cornea, unlike many other tissues, is devoid of resident professional antigen-presenting cells, although isolated MHC class II+ or CD45+ (leukocyte common marker) cells had been observed in the normal corneal stroma or central epithelium of several species [54,60–64]. This paradigm was recently shaken by the demonstration that the cornea is endowed with resident dendritic cells that are universally MHC class II-negative, but are capable of expressing class II antigen after surgery and migrating to draining lymph nodes of allograft hosts [39,65–68]. Many of these cells appear to be Langerhans cells that reside in the normal central corneal epithelium. Examinations of normal corneas revealed that both the peripheral and central areas of the epithelium contain large numbers of bone marrow-derived Langerhans cells, with the density of these cells decreasing from the limbus (178 Langerhans cells/mm2) toward the center (100 Langerhans cells/mm2) [65]. While a large number of Langerhans cells are MHC class II+ in the periphery, a large population of MHC class II-negative immature/precursor Langerhans cells are present both in the periphery and the center of the epithelium, with the center being exclusively MHC class II-negative and B7 (CD80 or CD86) co-stimulatory marker negative [65].