Ординатура / Офтальмология / Английские материалы / Ocular Therapeutics Eye on New Discoveries_Yorio, Clark, Wax_2007

standard diagnostic equipment, such as the slit lamp biomicroscope. Because of this, and since uveal inflammation spills into the anterior chamber and/or vitreal cavity, from a clinical standpoint, uveitis has a looser definition and refers to active inflammation in the intraocular compartments of the eye (Read, 2006).

As reviewed below in the “Basic mechanisms” section, from an etiologic standpoint, uveitis can be classified as either infectious or non-infectious. However, since the underlying etiology is often not apparent initially, from a clinical perspective uveitis is classified based on the location of the ocular involvement. Accordingly, clinical uveitis is classified as being “anterior”, “intermediate”, “posterior”, or if it involves all ocular compartments as “panuveitis”. Anterior uveitis, which is by far the most common form of uveitis, is also referred to as “iritis”, terms that are often used interchangeably. In iritis, active inflammation is seen in the anterior chamber. Along with this inflammation, there is breakdown in the blood–aqueous barrier and concomitant leakage of proteins in the aqueous humor that fills the anterior chamber – leading to the clinical description of “flare” when the slitlamp beam is focused through the anterior chamber. The clinical grade of the inflammation is therefore based on the degree (1–4 ) of cells and flare in the anterior chamber, parameters that are used both initially at the time of diagnosis, as well as in follow-up to gauge response to therapy. Other clinical signs of anterior uveitis may include “ciliary flush”, which refers to vascular engorgement in the perilimbal vasculature; “keratic precipitates”, which refer to collections of mononuclear cells (primarily T-cells and macrophages) adherent to the corneal endothelium; and spillover inflammation into the anterior vitreal space.

There are many dozens of infectious and non-infectious etiologies and syndromes associated with anterior uveitis, and hence their review is well beyond the scope of this brief overview. However, several issues are worthy of mention. First, it is important

to emphasize that given current diagnostic tests, close to one-half of anterior uveitis cases are still classified as idiopathic, since no one underlying etiology can be identified in spite of extensive laboratory testing. Most of these cases are unilateral and limited to one or several acute attacks; chronic disease is not the norm, though it may occur in a substantial minority of cases(Curi et al., 2005). Among adults, in particular younger males, HLA B27-associated disease comprises a large subset of those afflicted with anterior uveitis. The HLA B27 gene locus is associated with several syndromes including ankylosing spondylitis, Reiter syndrome, psoriatic arthritis, and inflammatory bowel disease. For reasons that are not quite clear, the coincidence of spondyloarthropathy (presented as lower back pain), uveitis, and other autoimmune disorders affecting the joints and/or gut has been appreciated for a long time. Patients with HLA B27-associated disease typically have anterior disease affecting one eye at a time, though recurrent disease may “jump” from one eye to the other from attack to attack. While in the majority of cases these attacks are acute and limited to the anterior compartments of the eye, some patients with HLA B27-associated uveitis develop more chronic disease, or involvement of the posterior segment, requiring more aggressive management.

Patients with intermediate uveitis have inflammation that is most pronounced in the intermediate compartment of the eye involving the ciliary body (“cyclitis”, “pars planitis”) and anterior vitreous cavity (“vitritis”). As mentioned above, even in anterior uveitis, cells can “spill over” into the vitreous cavity, but the degree of inflammation in the anterior chamber is always more marked than that seen in the anterior and mid vitreous. In contrast, in intermediate uveitis the degree of inflammation in the pars plana and anterior vitreous cavity is more than that in the anterior chamber. Most cases of intermediate uveitis are of unknown cause, though a number of systemic autoimmune conditions (e.g. multiple sclerosis,

syphilis, sarcoidosis, etc.) can present as intermediate uveitis. The most common form of intermediate uveitis is termed “pars planitis”, which is by definition of unknown cause (idiopathic). Often bilateral, pars planitis is characterized by smoldering low grade inflammation. However, a significant number of patients affected by this condition have associated vision loss due to development of cystoid macular edema (CME) in the posterior pole, presumably due to diffusion of proinflammatory factors through the vitreous to the retina, where these factors can promote vascular leakage from the retinal vessels, causing accumulation of fluid into the interstitial spaces of the retina, leading to CME.

The most heterogeneous group of uveitis conditions are those comprising posterior uveitis. A wide array of infectious (e.g. toxoplasmic, syphilitic, mycobacterial, and herpetic) conditions and non-infectious syndromes (e.g. vasculitides, systemic lupus erythematosus, etc.) can present as posterior uveitis. Some of these present as primarily choroidal diseases with variable degrees of inflammation seen in the vitreous, such as tubercular infections. In other cases, the presentation is mostly limited to the inner retina and vitreous, such as in toxoplasmosis which can lead to a form of retinal necrosis. Other cases, such as in lupus and in many vasculitic syndromes (e.g. Wegener granulomatosis, etc.), are primarily characterized by a pathological picture focused on the blood vessels. Perivascular “cuffing” or “sheathing” (representing localized areas of vasculitis), vascular occlusions, hemorrhaging and neovascularization are all possible presentations in these forms of posterior uveitis. In general, the vision is more severely affected in posterior uveitis than in anterior uveitis, particularly if there is involvement of the macula and/or the condition is more chronic.

2. Basic mechanisms

Uveitis is classified as either infectious or non-infectious. Infectious uveitis can be

caused by a wide variety of microorganisms including viruses, fungi, bacteria, protozoa, helminths, and rickettsiae. Specific microorganisms implicated in infectious uveitis include Toxoplasma gondii, Histoplasma capsulatum, Toxocara canis, and Mycobacterium tuberculosis. The predominant form of uveitis is non-infectious and believed to be elicited by autoimmune responses to tissue-specific antigens in the retina and uveal tract. Uveitis of the anterior segment accounts for approximately 75% of the cases of uveitis, and as the name implies, inflammation occurs in the iris and ciliary body, but can spread to the vitreous. However, retinal involvement is not a feature of anterior uveitis.

Animal models have shed light on the pathophysiology of uveitis. Murine experimental autoimmune uveoretinitis (EAU) is the most widely utilized animal model of this disease and has provided important insights into the mechanisms of non-infec- tious uveitis. Immunization with retinal antigens such as retinal S antigen (S-Ag), interphotoreceptor binding peptide (IRBP), rhodopsin, or phosducin in the presence of complete Freund’s adjuvant (CFA) elicits EAU in rodents, which displays many of the histopathological and clinical features of human uveitis (Agarwal and Caspi, 2004). A wealth of data indicate that EAU is a CD4 T-cell-mediated disease involving T-helper cells. It is widely believed that EAU is produced by CD4 T-helper type 1 (Th1) cells that elicit inflammation through their production of interferon-γ (IFN-γ). However, an aberrant form of EAU can be elicited in IFN-γ knockout (KO) mice and appears to be mediated by a Th2 type of autoimmunity (Jones et al., 1997).

One of the enigmas of non-infectious uveitis is the mechanism by which retinaspecific CD4 T-cells circulating in the bloodstream find their target antigen in the eye and breach the blood–retina barrier. Compelling data suggest that the first wave of uveitogenic CD4 T-cells enter the uveal tract randomly (Prendergast et al., 1998). Activation of the T-cells is crucial, as T-cells that have not been recently activated are

unable to enter the eye and do not produce uveitis (Agarwal and Caspi, 2004). It has also been suggested that uveitogenic T-cells cannot cross the blood–retinal barrier unless there has been a systemic signal, which renders the retinal vascular endothelial cells amenable to lymphocyte diapedesis (Xu et al., 2004). The CD4 T-cells that enter the retina elaborate a variety of cytokines and chemokines that recruit additional inflammatory cells including activated T-cells, granulocytes, macrophages, and antigen presenting dendritic cells (Jiang et al., 1999). Although EAU is mediated by CD4 T- cells, macrophages also contribute to tissue damage and disease (Dick et al., 2004). The amplifying capacity of CD4 T-cells is remarkable. It has been estimated that as few as 15 retinal antigen-specific CD4 Th1 effector cells are capable of initiating the inflammatory cascade and producing EAU in rodents (Caspi, 2006). Macrophages are recruited to the inflammation site and are activated by cytokines – principally inter- feron-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) – which are elaborated by retinaspecific CD4 Th1 cells. Within the inflammatory site, the activated macrophages release large amounts of reactive oxygen species including nitric oxide, which produce extensive tissue damage (Robertson et al., 2002). Thus, the initial phase of EAU is retinal antigen-specific and mediated by CD4 T-cells that recognize tissue-specific epitopes on the cells in the retina and uveal tract, but the subsequent damage to the eye is produced by an inflammatory process that indiscriminately inflicts injury to cells that may not even express the retina-specific antigens initially recognized by the CD4 T- cells.

3. Current and future therapy

a. Current therapy – Due to the complexity and variations in the presentation of uveitis, and the myriad complicating variables that can affect the treatment, readers are referred to more focused and exhaustive

reviews for these for guidelines (Kim and Foster, 2006). Herein, we will provide some broad issues relevant to therapy rather than provision of prescriptive steps in management of clinical uveitis.

The management of the uveitis patient should begin with taking appropriate diagnostic steps to determine, if possible, the underlying etiology. Laboratory (serum, urine), and radiographic studies are typically done to rule out underlying auto-immune or infectious disease. A high suspicion for infection will require a thorough review of systems (and incorporation of a non-ophthalmologist such as an infectious disease expert, if necessary), and at times a diagnostic tap of the eye for culturing or PCR testing.

In the vast majority of cases, instituting a reasonable and effective anti-inflammatory therapy is key to management of the uveitis patient. The vast majority of patients with acute anterior disease can be treated with topical corticosteroid therapy alone. Cycloplegia is also frequently provided to relax the ciliary muscles and prevent spasm and development of posterior synechiae. Patients prone to chronic or severe inflammation, and hence at risk of developing CME, can also benefit from topical non-steroidal anti-inflammatory(NSAID)therapy.NSAIDs, either topical or oral, are effective in prophylaxis and treatment of CME, and their use in high risk patients, if not contraindicated, can be very useful.

A minority of patients with anterior uveitis, and a majority of those with posterior uveitis, require more than just topical therapy for management of their condition. Topical therapies do not penetrate the deep ocular structures at levels required to control intense inflammation. For this reason, regional therapy and/or systemic treatment is required for cases of uveitis not limited to the anterior segment. Regional therapy is focused primarily on injection of a depot of corticosteroid in the peribulbar area, either in the sub-Tenon space or trans-septally through the eyelid. Depot injections of

corticosteroid can be quite effective in reducing inflammation over weeks to months. Their benefit is in not exposing the patient to systemic immune modulation. However, they also have significant limitations. First, spikes in intraocular pressure are common and can be severe and last for weeks to months as a result of these injections. Second, uveitis due to active systemic disease, such as vasculitis, cannot be adequately controlled by local injections, and needs systemic immune modulation. Third, these local injections are best for monocular cases and in cases where both eyes are involved, often due to active systemic disease, as systemic immunomodulation would be the preferred approach. Finally, corticosteroids are potent suppressors of innate immunity, including macrophage function, and hence can profoundly suppress the host response in many infections. Certain patients with infectious uveitis, such as toxoplasmosis, are not good conditions for regional depot injections of corticosteroids (Koo and Young, 2006).

There is a large body of literature on the use of systemic immune modulators in the treatment of uveitis. However, there is a dearth of well-controlled prospective randomized studies; rather, the literature is replete with small uncontrolled case series of patients with highly heterogeneous disease who receive one or the other treatment. There are, therefore, few firm guidelines that have reached wide consensus. However, several broad guidelines are worthy of emphasis. First, a “step-ladder” approach is advocated, where treatment with a less toxic strategy is initiated, and based on the patient’s response (or lack of response), the therapy is altered to potentially include a more effective, but toxic and risky, medication. An example would be a patient with retinal vasculitis who would first be treated with methotrexate and prednisone, but who after several months continues to progress gradually. A decision to institute cytotoxic therapy would then be possibly warranted. Second, in cases when the underlying disease is known, it is probably

best to start therapy with medications that have a “tried and true” history of success. Examples would be use of methotrexate in rheumatoid arthritis, or cyclophosphamide in Wegener granulomatosis. Third, the last decade has witnessed a surge in interest of “biologic” agents that target specific molecular pathways (e.g. CD25, TNF-α, IL-1, etc.), and there is a growing literature that some of these agents are safer than standard cytotoxic agents, and are indeed quite effective in management of ocular inflammatory disease (Lim et al., 2006). Fourth, it is critical that ophthalmologists involved in the care of these patients appreciate the limits of their expertise, and the ability of their practices, to monitor patients on systemic therapy and manage the side effects. Few are able to assume this level of care, and hence it is highly recommended that the ophthalmologist work with other experts (e.g. rheumatologists or oncologists) in managing patients who require systemic immune suppression. Finally, it is important to appreciate, and to educate the patient with severe uveitis, that in spite of the most aggressive management strategy, vision loss is quite common. In particular, cases involving vascular disease and ischemia, and retinal or optic nerve disease can be associated with significant levels of vision loss.

b. Future therapy – The advantage of animal models such as EAU is that they permit prospective analysis of potential therapeutic modalities. In the case of an immunemediated disease, therapies can either inhibit the induction phase of an immune response to retinal autoantigens or disable the effector stage once it has been evoked. Antigens introduced via mucosal surfaces or into immune privileged sites, such as the anterior chamber (AC) of the eye, elicit immune tolerance in which destructive CD4 Th1 immune effector cells are silenced by T- regulatory cells (Tregs) (Weiner, 1997; Egan et al., 2000; Streilein, 2003; Mowat et al., 2004; Ashour and Niederkorn, 2006; Caspi, 2006). Injecting soluble antigens into the AC

of the eye induces a systemic down regulation of Th1 immune responses that has been termed anterior chamber-associated immune deviation (ACAID) (Streilein, 2003; Ashour and Niederkorn, 2006). Interestingly, ACAID can be imposed on hosts that have been previously immunized with antigens such as IRBP, resulting in the mitigation of EAU, even after intraocular inflammation has begun (Hara et al., 1992). Oral administration of antigen elicits a form of immune tolerance that bears a striking resemblance to ACAID and produces a similar mitigation of EAU (Rizzo et al., 1999). However, attempts to utilize oral tolerance with retinal S-antigen to treat uveitis patients have been disappointing (Nussenblatt et al., 1997).

Most therapeutic strategies for managing EAU have been directed at the efferent arm of the immune response, as this is the stage at which uveitis patients will be identified. The pathophysiology of EAU is a sequential process that begins with the retinal antigen-specific, CD4 Th1 cells traversing the blood–retinal barrier and entering the retina and uveal tract. Therefore, this stage can be targeted with monoclonal antibodies against the cell adhesion molecules, such as intercellular adhesion molecule-1 (ICAM-1), which are expressed on the vascular endothelium and serve as adhesins for facilitating the entry of the pathogenic T- cells into the retina and uveal tract. Blocking the adhesion of circulating retinal antigenspecific CD4 Th1 cells to the retinal vascular endothelium, in theory, is a potential strategy for preventing their entry into the retina and uveal tract. Once CD4 Th1 cells have entered the retina and have begun producing pro-inflammatory cytokines such as IFN-γ and TNF-α, systemic administration of neutralizing antibodies to these cytokines is a potential strategy for controlling immune-mediated injury to the eye. The successful use of anti-TNF-α antibody (Remicade®) and soluble TNF-α receptor (Enbrel®) in the treatment of rheumatic diseases suggests that blocking this

pro-inflammatory cytokine may be useful in the management of uveitis. Ultimately, induction of retinal antigen-specific Tregs is the most desirable strategy for managing autoimmune diseases such as uveitis. However, a formidable obstacle in employing

Tregs is the difficulty in identifying the relevant antigens that elicit the generation of

autoimmune CD4 Th1 cells. To date, this has eluded researchers, but may ultimately be achieved with continued research and the emergence of more sophisticated molecular technology.

H. Corneal Allograft Rejection

1. Clinical disease

Corneal grafting, or penetrating keratoplasty (PK), is the most common form of tissue transplantation. More corneal transplants are performed each year than all other forms of transplantation combined. In the United States alone, nearly 40,000 cases are performed annually, but this far underestimates the medical “need” for this procedure, since corneal blindness is the second most common cause of vision loss worldwide after cataracts. This means that there are millions of people with scarred and/or vascularized corneas that may benefit from corneal surgery, but due to a host of factors, such as economic poverty, lack of access to eye banking and donor tissues, and lack of access to appropriate health care, many people who could benefit from this procedure are not receiving it.

Most patients who receive a corneal graft for the first time for a corneal scar or opacity in an uninflamed eye, say for bullous keratopathy or keratoconus, have a very good prognosis with PK. In these cases, the 2-year survival rate under cover of local immune suppression, afforded by corticosteroid therapy, is over 85% (Rocha et al., 1998; Qian and Dana, 2001) While topical steroid therapy may be fraught with many side effects, including elevation of intraocular pressure and glaucoma, infection, and

stromal thinning, it is still remarkable that topical therapy can lead to such extraordinary rates of success that can be achieved in other solid grafts only with profound systemic immune suppression. This high rate of success has been related to various features of the cornea and ocular microenvironment that together account for its so-called “immune-privileged” status (Streilein, 1999a,b). However, many corneal grafts are still rejected, and immune rejection is the leading cause of corneal graft failure (Rocha et al., 1998; Niederkorn, 1999a). Inflammation in the corneal graft bed with attendant neovascularization (NV) are the leading tissue characteristics that herald a high risk of rejection to a transplant (Mader and Stulting, 1991; Dana and Streilein, 1996). Unfortunately, NV is a ubiquitous element of corneal pathology that accompanies a vast array of traumatic, inflammatory, infectious, and toxic insults (Epstein et al., 1987). Grafts placed into “high risk” beds with NV exhibit rejection rates which increase to well over 50–90% even with maximal local and systemic immune suppression (Mader and Stulting, 1991).

The clinical presentation of the rejecting corneal graft can range considerably (Qian and Dana, 2001). The classic presentation is the graft whose endothelium is being visibly destroyed by an endothelial “rejection line”, comprised of mononuclear cells including T-cells and macrophages, that advances usually from the interior and leaves in its wake a swollen graft without adequate endothelial function. Concomitant with this would be inflammation in the anterior chamber with or without keratic precipitates. However, only about one-third of rejecting eyes present with this picture. The remainder present with varying degrees of inflammation in the anterior chamber with a graft that becomes progressively swollen, typically in a matter of days to weeks. These presentations all comprise “endothelial rejection”, the principal form of corneal transplant rejection and the one that leads to eventual graft decompensation. The other

principal form of rejection occurs in the epithelium, and is seen as a faint line, most visible with the application of fluorescein, running across the graft epithelium. While not in itself a major threat to graft viability, it could be the harbinger of an alloimmune process, and hence should be seen as a “red flag” by the clinician that unless steroids are increased an endothelial rejection process may commence.

2. Basic Mechanisms of Corneal Allograft

Rejection

Although the immunologic basis for corneal allograft rejection was established over 50 years ago (Maumenee, 1951), the precise immune effector mechanism remains poorly understood. Studies employing rodent models of penetrating keratoplasty have firmly established that T-cells, namely CD4 T-cells, are required for corneal allograft rejection (Niederkorn, 1999a,b, 2001, 2002). Although corneal allografts stimulate the production of serum antibodies directed at the histocompatibility antigens expressed on the donor cornea, efforts to demonstrate a role for antibody-mediated rejection have failed (Hegde et al., 2002; Hargrave et al., 2003). The expression of complement regulatory proteins on the corneal cells and in the aqueous humor neutralize the potential damaging effects of complement activation by donor-specific antibodies (Lass et al., 1990; Bora et al., 1993; Goslings et al., 1998; Hegde et al., 2002; Hargrave et al., 2003). Moreover, corneal allograft rejection occurs in B-cell deficient mice that are incapable of mounting antibody responses to the corneal donor’s histocompatibility antigens and in complement-deficient mice that cannot mount complement-mediated cytolysis of target cells (Goslings et al., 1999; Hegde et al., 2002).

Histopathological and immunohistochemical analysis of rejected corneal allografts in humans and rodents have revealed the presence of a mixed inflammatory infiltrate that includes CD4 T-cells,

CD8 T-cells, macrophages, and PMNs. The “rejection line” that has been observed in rejecting corneal allografts in patients and animals led some to believe that corneal allograft rejection was the result of piecemeal necrosis of corneal cells by cytotoxic T-lym- phocytes (CTL). Indeed, histopathological examination of rejected corneal allografts in rodents reveals the presence of CD8 T-cells (the phenotype of CTL) that are in direct contact with necrotic corneal cells. However, orthotopic corneal grafts are notoriously poor at inducing CTL responses, and corneal graft rejection occurs unabatedly in mice that have been depleted of CD8 T- cells using monoclonal antibodies and in CD8 knockout (KO) mice that cannot develop CTL (Hegde and Niederkorn, 2000; Niederkorn, 1999b, 2001, 2002). By contrast, corneal graft rejection is sharply reduced in CD4 KO mice and in normal mice depleted of CD4 T-cells by the systemic administration of anti-CD4 monoclonal antibody (He et al., 1991; Ayliffe et al., 1992; Yamada et al., 1999; Hegde et al., 2005). However, closer scrutiny of the data reveals that CD4 T-cells alone do not account for all episodes of corneal allograft rejection. In fact, corneal allografts undergo rejection in 33% of the mice and 64% of the rats treated with anti-CD4 antibody and in 45% of the CD4 KO mice (He et al., 1991; Ayliffe et al., 1992; Yamada et al., 1999; Hegde et al., 2005). Recent findings in a mouse model of penetrating keratoplasty indicate that CD4 T-cell-independent rejection can occur and can be mediated by two populations of CD4 T-cells; one population is comprised of CD4 , CD8 T-cells and the other is made up of CD4 , CD8 T-cells (Niederkorn et al., 2006b). Thus, there is considerable plasticity and redundancy in the cell populations that are capable of mediating corneal allograft rejection.

The precise mechanisms that lead to corneal allograft rejection remain poorly understood. Delayed type hypersensitivity (DTH) to the cornea donor’s histocompatibility antigens remains the most reliable

and consistent immune effector function associated with corneal graft rejection. Moreover, maneuvers that inhibit the generation or expression of DTH to the donor’s histocompatibility antigens promote corneal allograft survival. We still do not have a clear understanding as to what constitutes the lethal hit that leads to the demise of the corneal allograft and the loss of its function as a refractive organ (i.e. its clarity). The survival of corneal allografts denuded of their epithelium, and the capacity of the corneal epithelium to be re-established from stem cells in the graft bed, support the notion that immune destruction of the corneal epithelium alone is not the cause of corneal allograft failure. The overwhelming consensus is that irreparable damage to the corneal endothelium is the critical event that leads to corneal graft failure. As mentioned earlier, DTH is the immune effector function that is most closely associated with corneal allograft rejection. The elicitation of DTH, which is classically mediated by CD4 T- cells, is exquisitely antigen-specific, yet the expression of the DTH inflammatory response is remarkably antigen-non-spe- cific and culminates in extensive damage to innocent bystander cells due to the generation of toxic molecules, such as reactive oxygen species, nitric oxide, proteases, and the recruitment of secondary inflammatory cells, such as macrophages and PMNs. In addition to the DTH inflammatory response, CD4 , CD8 and CD4 , CD8 T-cells are capable of inducing apoptosis of the donor’s corneal endothelial cells via contact-depend- ent, perforin-independent mechanisms (Niederkorn et al., 2006b,c). However, the effector molecules that produce apoptosis remain to be identified.

Thus, in spite of over half a century of laboratory investigations, much remains to be learned about the immune effector elements and the mechanisms that lead to corneal allograft rejection. Understanding this process will be crucial for developing improved strategies for promoting corneal

allograft survival, especially in the high risk patient.

3. Current and Future Therapy

a. Current therapy – The therapy of the PK patient has two dimensions: prevention of rejection (prophylactic therapy), and treatment of the rejecting graft. There are many nuances to the treatment regimens employed in PK patients that are well beyond this brief review; hence, we will focus only on broad general guidelines. In general, for the first-time PK recipient who is at low risk of rejecting the graft, the patient is maintained on topical steroids for 9–12 months and is finally tapered off treatment if there is no sign of an immune response. This is particularly the case in younger phakic patients, in whom the risks of long-term steroid therapy (cataract and glaucoma) can outweigh the potential benefits. Conversely, in the older pseudophakic patient with no history of glaucoma, the patient is often retained on low dose/frequency steroid therapy since the benefit of this therapy in retarding or preventing rejection outweighs the incremental risks of chronic steroid therapy. In all cases the patients are monitored closely for high intraocular pressure.

The approach toward prophylactic therapy in the patient at high risk of rejection, with an inflamed and neovascularized bed, or with a history of graft rejection, is different. While there is no consensus regarding risk classification or management of high risk patients, most authorities would agree that more immune suppression should be employed than that customarily used for the low risk patients. This can range from employing higher frequency topical steroids for a longer duration, say in a patient with a history of graft rejection, but with an otherwise normal recipient bed, to the use of systemic immunosuppressives (e.g. combination of low dose cyclosporine-A (CsA) and prednisone) in addition to topical therapy for the patient at very high risk

of rejection, say the patient with extensive deep stromal vascularity in the host bed. After adequate education of the patient regarding potential risks to systemic immune suppression, an “aggressive” therapeutic approach may be employed to minimize chances of an immune attack against the graft. Therapy often begins just before the transplant with oral prednisone, or on the day of transplantation with an intravenous dose of methyl-prednisolone. Oral steroids are tapered over a few weeks while the patient is maintained on low dose oral CsA for a prolonged period.

The approach toward treating, rather than preventing, a graft rejection is quite different. Here, hours to days can make a major difference in eventual graft outcome; hence, the most critical facet in the management of the rejecting corneal transplant is early detection and initiation of therapy in what is truly an ocular emergency. Typically, as soon as an endothelial rejection line is appreciated or suspected (due to presence of other signs as reviewed above), the patient is placed on topical prednisolone on an hourly basis. Clinical exam and serial pachymetry can be used to gauge response to therapy; indeed, a responding graft may regain near-normal thickness within a matter of days to weeks. Similar to risk stratification in the prophylactic approaches to these patients, the treatment approaches can vary based on the history and presentation. For example, if a graft is rejecting for the second or third time in an inflamed host bed, institution of topical prednisolone therapy may be inadequate to salvage the transplant. Other approaches could include injection of depot steroids periocularly, or placement of the patient on systemic immune suppression, including steroids. In severe cases, or when attempting to salvage a graft in the only-seeing eye of a patient, use of more potent immunosuppressives such as CsA or Cellcept may be employed. The success of these treatment strategies can vary widely. Close to twothirds of a first rejection episode in a low risk

transplant can be successfully reversed if diagnosed and managed in a timely manner. In contrast, a rejection episode in a high risk eye, or one that is diagnosed late, is much less apt to respond to therapy.

b. Future therapy

BOX 10.1

It is remarkable that more than 60 years after the widespread adoption of corneal transplantation in the United States, a period that has witnessed very significant advances in eye banking and microsurgical techniques, little has changed in the medical management of this procedure. The currently available pharmaceutical armamentarium for corneal transplant survival is principally comprised of steroids. The introduction of these agents into the field of ophthalmology remains the single most significant factor in promoting corneal graft success in the past few decades. However, beyond their well-known serious complications, steroids show widely variable efficacy in preventing ultimate immunogenic graft failure, and this is particularly the case in high risk keratoplasty. In response to this need, research into immune modulation in corneal transplantation has progressed considerably in the past two decades, identifying new approaches toward promotion of graft success (Dana et al., 2000).

As discussed in the above section on the mechanisms of rejection, CD4 T-cells are thought to play a significant role in effecting graft damage, and one strategy that has shown utility in experimental rodent models of corneal grafting has been depletion of antibody blockade of CD4 T-cells (He et al., 1991; Pleyer et al., 1995). CD4 T- cells play a critical function in host defense against myriad threats, including infection, and hence this may not prove a feasible and practical approach for clinical applications. A second approach that has shown

experimental benefit has centered on prevention of the mobilization of APCs in the graft. These cells are absolutely critical for host sensitization to transplant antigens (Niederkorn, 1995), and maneuvers that can block their infiltration via antagonism of cytokines or chemokines responsible for their mobilization, or that can deplete them in the graft, may prove successful in preventing clinical rejection (Yamada et al., 1998; Dekaris et al., 1999; Yamagami et al., 2005). Recently, it has been shown that these APCs need to traffic to lymphoid reservoirs, including draining lymph nodes, to perform their function in immunizing the host (Yamagami et al., 2002). The access of these cells to lymphatics, which drain into lymph nodes, is in large part mediated by a specific receptor, vascular endothelial growth factor receptor-3 (VEGFR-3), and it has been shown in a mouse model of corneal transplantation that blockade of VEGFR-3 can promote graft survival (Chen et al., 2004).

As stated above, vascularization of the graft bed or growth of blood vessels into the graft can pose a significant threat to graft outcome by facilitating immune responses (CCTS, 1992; Dana and Streilein, 1996). One promising strategy for promoting graft success would be inhibition of angiogenesis, for example, by blocking factors (including VEGF) that are required for vascular endothelial cell migration and proliferation, as has been shown recently in a mouse model of transplantion (Cursiefen et al., 2004). Cell migration and infiltration, whether of immunoinflammatory or vascular endothelial cells, requires the coordinated activity of a number of cell adhesion factors. And consequently, antagonism of these adhesion factors has been shown to be highly effective in suppressing allograft rejection (Whitcup et al., 1993; He et al., 1994; Philipp, 1994; Yamagami et al., 1995; Zhu et al., 2000). Finally, the

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gene expression of many cell adhesion and growth factors is under the regulation of “master” pro-inflammatory molecules, interleukin-1 (IL-1), and tumor necrosis factor-alpha (TNF-α) (Yamagami et al., 1999, 2000). Local or systemic antagonism of these cytokines has been shown to suppress induction of host immunity to corneal grafts, growth of transplant-associated angiogenesis, and transplant rejection.

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