AAO

Ocular Immunology
Overview of the Ocular Surface Immune Response
For an in-depth discussion of the various features of the innate and adaptive immune system,
including types of hypersensitivity reactions with relevant ocular examples, see BCSC
Section 9, Intraocular Inflammation and Uveitis. This chapter is an overview of the ocular
surface immune response, which involves components of the immune system, the tear film,
and the lacrimal functional unit, a complex apparatus consisting of the lacrimal glands, ocular
surface (cornea, conjunctiva, and meibomian glands), and eyelids, as well as the sensory and
motor nerves that connect these structures (see Chapter l).
Tear Film
The normal tear film is a complex structure that contains a variety of elements, including
components of the complement cascade, proteins, growth factors, and an array of cyto- kines.
Cytokines such as interleukin-l (IL-l) and tumor necrosis factor a (TNF-a) are significantly
upregulated in a variety of corneal inflammatory disorders, such as corneal graft rejection and
dry eye disease. Similarly, increased expression of growth factors, pros- taglandins,
neuropeptides, and pro teases (Table 6-1) has been observed in a wide array of immune
disorders of the cornea and ocular surface. Effective immune responses to foreign antigens
require cells to "traffic" through tis- sues. Chemokines (chemotactic cytokines) are critical
mediators that provide the traffick- ing signals to immune cells. These are low-molecularweight proteins, of which more than 50 have been identified to date; they have been classified
into subgroups based on their amino acid sequence. Although there is some overlap in the
function of these cytokine species, they can also be classified functionally into those that
promote neutrophil recruit- ment (eg, IL-8); T helper-1 (Thl) lymphocyte recruitment and
activation (macrophage inflammatory protein -1 ~ [MIP-1 ~] ); monocyte-macrophage
recruitment (monocyte che- motactic protein-1 [MCP-1]); and eosinophil recruitment
(eotaxin). Many chemokines have been identified as playing important roles in corneal
inflam- mation. A brief tabulation of some important soluble mediators involved in immune
and inflammatory responses of the cornea and ocular surface appears in Table 6-1.
lmmunoregulation of the Ocular Surface
Immunoregulation of the ocular surface occurs through tolerance and regulation of the innate
and adaptive arms of the ocular immune response.

The normal, uninflamed conjunctiva contains polymorphonuclear leukocytes (neutrophils);

lymphocytes (includ- ing regulatory T cells [Treg cells], which dampen the immune
response); macrophages; plasma cells; and mast cells. In addition, the conjunctival stroma has
an endowment of dendritic antigen-presenting cells (APCs). The epithelium contains a
special subpopu- lation of dendritic APCs known as Langerhans cells, which are capable of
both antigen uptake and priming (sensitizing) of naive (antigen-inexperienced) T
lymphocytes. Hence, these dendritic cells serve as the sentinel cells of the immune system of
the ocular surface. In addition to containing immune cells, the conjunctiva has a plentiful
supply of blood vessels and lymphatic vessels, which facilitate the trafficking of immune
cells and antigens to the draining lymph nodes, where the adaptive immune response is
generated. This oc- curs through the recruitment ofTreg cells, which return to the ocular
surface to modulate and suppress the local immune response (Fig 6-1). The normal,
uninflamed cornea, like the conjunctiva, is endowed with dendritic cells. Like those in the
conjunctiva, the dendritic cells in the corneal epithelium are called Lang- erhans cells. They
are located primarily in the corneal periphery and limbus. These APCs are in an activated,
mature state (expressing class II major histocompatibility complex [MHC] antigens and
costimulatory molecules) and hence are capable of efficiently stimu- lating T cells. In
addition to these dendritic cells (Fig 6-2), small numbers oflymphocytes are present in the
peripheral epithelium and anterior stroma of the cornea. A highly regulated process, mediated
by vascular endothelial adhesion molecules and cytokines, con- trols the recruitment of the
various leukocyte subsets from the intravascular compartment into the limbal matrix. Immune

responses are also mediated by Treg cells in the regional lymph nodes and perhaps at the
local level as well.
Unlike the conjunctiva, the normal cornea is considered an immunologically priv- ileged site,
so called because the generation of immune response to foreign (including transplant)
antigens is relatively suppressed. The normal cornea's immune privilege is due to a multitude
of factors, including the following:
absence of blood vessels, which impedes delivery of immune effector cells
absence oflymphatics, which minimizes flow of antigens and APCs to the draining lymph
nodes
expression of immunosuppressive factors, including transforming growth fac- tor ~ (TGF~), and neuropeptides, such as a-melanocyte-stimulating hormone (a-MSH)
expression ofFas ligand (CD95) by corneal cells, which is believed to play a criti- cal role in
inducing Pas-mediated apoptosis (programmed cell death) of activated lymphocytes
exposure to the anterior chamber, which contains immunosuppressive factors, in- cluding
corticosteroid
The downregulatory immune response to antigens in the cornea and anterior cham- ber may
lead to immune unresponsiveness or even immunologic tolerance.
Niederkorn )Y. Cornea: window to ocular immunology. Curr Immunol Rev. 2011;7(3): 328335. Stern ME, Schaumburg CS, Dana R, Calonge M, Niederkorn )Y, Pflugfelder SC. Autoimmunity at the ocular surface: pathogenesis and regulation. Mucosal Immuno/. 20
10;3(5):425-442.
Angiogenesis and Lymphangiogenesis in the Cornea
Though not normally present in the cornea, blood and lymphatic vessels may extend into the
cornea- as sprouts of the vascular endothelium from the limbal tissue-after inflam- matory,
infectious, traumatic, chemical, or toxic insults. The cellular and molecular mechanisms of
corneal angiogenesis and lymphangiogenesis are not completely understood. Inflammatory
cells infiltrate tissue at local sites of vascular remodeling, where they secrete proangiogenic
factors and metalloproteinases. Vascular endothelial growth factor (VEGF) is upregulated in
inflamed and vascularized corneas in humans and in animal models. VEGF-A and fibroblast
growth factor 2 (FGF-2) induce lymphangiogenesis in corneas of C57BL/6 mice. VEGF-C
also induces lymphangiogenesis in various animal models and, under certain conditions,
angiogenesis in both humans and animal models. Lymphangio- genesis is thought to be
secondary to angiogenesis, suggesting common molecular and cellular origins for the 2
processes. Pooyan 66961526-7 Vascularization of the cornea increases the risk of immune
rejection after corneal transplantation, leading to a rate of graft rejection greater than 50%.
This may occur even when a strict regimen of topical and systemic immunosuppressive
agents is used. In fact, stratification of risk factors for immunologic rejection in penetrating

keratoplasty has identified recipient vascularization as a critical proximal cause of earlier and
more fulmi- nant rejection episodes. Lymphatic neovessels may grow in parallel with the
blood vessels; this facilitates access of donor and host APCs and antigenic material to
regional lymph nodes, accelerating sensitization to graft antigens.

Figure 6-1 A, lmmunoregulation of the ocular surface: The ocular surface tissues contain a
variety of soluble and cellular factors to reduce inflammation-induced pathology in the
lacrimal functional unit Those implicated in immunoregulation within the ocular surface
tissues include the fo llowing: (1) Natural regulatoryT cells (nTreg cells) (eg, CD4, CDS,
and natural killerT cells), which include many of the conjunctival intraepithelial lymphocytes,
are thought to dampen or inhibit the inflammatory/autoimmune response on the ocular
surface. (2) The anti-inflammatory cytokine transforming growth factor ~ (TGF-~) is present
on the ocular surface and has profound suppressive effects on resident dendritic cell (DC)
maturation in the cornea; proliferation, differentiation, and surviva l of auto reactive T cells;
and regulatory T cell (Treg cell) differentiation and maintenance. The activity of the potent
acute-response, proinflammatory cytokine interleukin-1 (IL-1) is modulated by the IL-1
receptor antagonist (IL-1 RA), which is expressed and secreted by corneal and conjunctival
epithelial cells. Vasoactive intestinal peptide (VIP) also seems to be protective; VIP secreted
by sensory nerve endings in the cornea increases production ofTGF- ~ and IL-10 and inhibits
expression of the proinflammatory cytokines and chemokines, I L-1 ~.tumor necrosis factor
a, interferon-y, and chemokine (C-X-C motif) ligand 2. Hormones are also implicated in
curbing inflammation and maintaining homeostasis. In addition, the cornea l epithelium
expresses vascular endothelial growth factor (VEGF)

receptor-1 to sequester VEGF and reduce neovascularization. (3) Antigen-presenting cells

(APCs) bearing self-antigen derived at the ocular surface may migrate to the regional lymph
nodes to induce antigenspecific Treg cells (iTreg cells). B, lmmunoregulation in the lymphoid
organs: nTreg cells may exert their immunosuppressive function through (1) re lease of
soluble factors (eg, TGF-~, IL-10); (2) ce ll-cell contact which disables pathogenic effectorT
cel ls (Teff cel ls) and/or APCs; and/or (3) competition for soluble factors (eg, IL-2). (4) iTreg
cells may use simi lar mechanisms to inhibit cells bearing or responding to autoantigens. It is
possible that these Treg-dependent mechanisms may also function within the ocular surface
tissues. C, Other peripheral immunoregulatory mechanisms: additional mechanisms also limit
access and effector function of autoreactive T cells within the ocular surface tissues: (1) TGFp and (2) nTreg and iTreg cells are thought to suppress infiltrating autoreactive lymphocytes,
and (3) lowlevel expression of integrins in endothelial cel ls of the healthy ocular surface,
coupled with expression of the programmed death ligand-1 (PD-L 1), negatively regulates
activated T cells w ithin the ocular surface tissues. (Modified with permission from Stern ME,
Schaumburg CS, Dana R, Calonge M, Niederkorn JY, Pflugfelder
SC. Autoimmunity at the ocular surface: pathogenesis and regulation. Mucosal lmmunol.
2010;3(5):425-442.)
Therefore, targeting angiogenesis in order to modulate immune responses after corneal
transplantation has been the primary area of interest for many researchers. Treatment of
corneal neovascularization after corneal transplantation may limit both the afferent
(sensitization) and efferent (rejection) arms of alloimmunity and thus re- duce the tendency
toward inflammatory reactions, which can jeopardize graft survival. VEGF inhibitors,
including pegaptanib sodium, ranibizumab, and bevacizumab, are used to treat neovascular
age-related macular degeneration. Recently, there has been increasing interest in using topical
and subconjunctival anti-VEGF to treat corneal neovascularization.
Bourghardt Peebo B, Fagerholm P, Traneus-Rockert C. Lagali N. Time-lapse in vivo imaging
of corneal angiogenesis: the role of inflammatory cells in capillary sprouting. Invest
Ophthalmol Vis Sci. 2011;52(6):3060-3068. Dastjerdi MH, Saban DR, Okanobo A, eta!.
Effects of topical and subconjunctival beva- cizumab in high-risk corneal transplant survival.
Invest Ophthalmol Vis Sci. 2010; 51(5):2411-2417. Ecoiffier T, Yuen D, Chen L. Differential
distribution of blood and lymphatic vessels in the murine cornea. Invest Ophthalmol Vis Sci.
2010;51(5):2436-2440.

Figure 6-2 Langerhans ce lls are a subpopulation of dendritic antigen-presenting ce lls of the
ocular surface epithelium. As the sentinel cells of the immune system, they pick up, process,
and present antigens toT cells. This micrograph shows the predominance of major
histocompatibility complex class II+ Langerhans cells in the limbus of the uninflamed eye. !
Courtesy of the laboratory of M. Reza Dana, MD.)
Tissue-Specific Patterns of Immune-Mediated Ocular Disease
Conjunctiva
The conjunctiva is part of the mucosa-associated lymphoid tissue (MALT), which involves a
variety of mucosal tissues in the body, including the lacrimal gland. Humoral immunity in the
conjunctiva largely involves IgA produced by the lacrimal gland, and cellular im- munity is
dominated by CD4+ T cells. Serosal mast cells, which contain neutral proteases, are normally
present in the conjunctiva, and the number of mucosal mast cells with gran- ules containing
only tryptase is increased in the conjunctiva of atopic patients. Mast-cell degranulation
produces conjunctival redness, chemosis, mucus discharge, and itching.
Cornea
The normal cornea can have neither an acute allergic reaction (as it contains no mast cells)
nor a typical Arthus reaction (as there are no blood vessels). However, the cornea does participate in immune reactions by way of humoral and cellular immune elements that enter the
periphery from the limbal blood vessels. These anatomical features may explain why so many
immune-mediated disorders of the cornea occur primarily in the corneal periphery and
limbus. Alternatively, ingress of leukocytes through the ciliary body and iris root and ingress
of plasma proteins through breakdown of the blood- ocular barrier (as occurs in uveitis
syndromes) are other means by which immune effectors gain access to the cornea. The cornea
can act as an immunologic blotter, soaking up antigens from the ocular surface. This
phenomenon was first described by Wessely in 1911, when foreign antigen was injected into
the cornea of a previously sensitized animal and a ring-shaped infil- trate formed in the
corneal stroma concentric to the injection site, much like an antigen- antibody complex in an
immunodiffusion test. Still called a Wessely immune ring, this infiltrate contains complement
factors and/or neutrophils. Circulating antibodies are not required if sufficient local antibody
production is stimulated by antigens deposited in the cornea. The antigen may be a drug, as in

the peripheral corneal infiltrates associated with a neomycin reaction; a foreign body; or an
unknown substance, as in the corneal infil- trates that can occur in contact lens wearers.
Wessely rings may persist for some time in corneas traumatized by a foreign body, even after
the foreign body is removed.
Sclera
Nearly 50% of patients with necrotizing scleritis have an associated systemic immunologic or
connective tissue disease. Immune-complex deposition, granulomatous inflammation, and
occlusive vasculitis have been implicated in the pathogenesis of scleral inflammation.
Diagnostic Approach to Immune-Mediated Ocular Disorders
Many, but not all, immune-mediated ocular disorders are secondary to a systemic disease. As
with most medical problems, diagnostic investigations need to begin with a complete history,
including a review of systems, and a general physical examination, as indicated.Some of the
more common laboratory diagnostic tests that are selected to further narrow the differential
diagnosis are listed in Table 6-2. In general, except in the case of rheuma- toid arthritis, which
has a strong predilection for scleral and corneal involvement, the workup for patients with
immune-mediated corneal disease in whom an underlying dis- ease is suspected is quite
similar to that for the uveitis patient. In a patient presenting with ocular inflammation,
diagnosing systemic vasculitis with tests, including antineutrophil cytoplasmic autoantibody
tests (see Table 6-2), may be instrumental in instituting early life-saving therapy.
See BCSC Section 9, Intraocular Inflammation and Uveitis, for more information on the
diagnostic workup of patients with uveitis. Table 6-3 provides the clinical interpre- tation of
ocular surface cytology for immune-mediated keratoconjunctivitis. Finally, it should be noted
that corneal and ocular surface morbidities may result from underlying autoimmune disease.
Generally, when a systemic disease is suspected, it is advisable to coordinate care with an
internist or rheumatologist, especially if systemic immune sup- pression is being considered.
Pooyan 66961526-7