Interferon gamma regulates platelet endothelial cell adhesion molecule 1 expression and neutrophil infiltration into herpes simplex virus-infected mouse corneas.

In a mouse model of herpes simplex virus (HSV) 1 corneal infection, tissue destruction results from a CD4+ T cell-mediated chronic inflammation, in which interleukin 2 and interferon (IFN) gamma are requisite inflammatory mediators and polymorphonuclear leukocytes (PMN) are the predominant infiltrating cells. In vivo neutralization of IFN-gamma relieved inflammation at least in part through a specific block of PMN extravasation into HSV-1-infected corneas. Intercellular adhesion molecule (ICAM) 1 and platelet endothelial cell adhesion molecule (PECAM) 1 were upregulated on the vascular endothelium of inflamed corneas. Reduced PMN extravasation in anti-IFN-gamma-treated mice was associated with a dramatic reduction of PECAM-1 but not ICAM-1 expression on vascular endothelium. PMN accumulated in the lumen of corneal vessels after in vivo IFN-gamma neutralization. PECAM-1 was readily detectable on PMN inside the vessels but was not detectable on PMN that extravasated into the infected cornea. Moreover, flow cytometric analysis revealed reduced PECAM-1 expression but elevated major histocompatibility complex class I expression on PMN that recently extravasated into the peritoneal cavity when compared with PMN in the peripheral blood. We conclude that IFN-gamma contributes to HSV-1-induced corneal inflammation by facilitating PMN infiltration; this appears to be accomplished through upregulation of PECAM-1 expression on the vascular endothelium; and PMN downregulate PECAM-1 expression during the process of extravasation.

Infected cell protein (ICP)47 enhances herpes simplex virus neurovirulence by blocking the CD8+ T cell response.

The herpes simplex virus (HSV) infected cell protein (ICP)47 blocks CD8+ T cell recognition of infected cells by inhibiting the transporter associated with antigen presentation (TAP). In vivo, HSV-1 replicates in two distinct tissues: in epithelial mucosa or epidermis, where the virus enters sensory neurons; and in the peripheral and central nervous system, where acute and subsequently latent infections occur. Here, we show that an HSV-1 ICP47- mutant is less neurovirulent than wild-type HSV-1 in mice, but replicates normally in epithelial tissues. The reduced neurovirulence of the ICP47- mutant was due to a protective CD8+ T cell response. When compared with wild-type virus, the ICP47- mutant expressed reduced neurovirulence in immunologically normal mice, and T cell-deficient nude mice after reconstitution with CD8+ T cells. However, the ICP47- mutant exhibited normal neurovirulence in mice that were acutely depleted of CD8+ T cells, and in nude mice that were not reconstituted, or were reconstituted with CD4+ T cells. In contrast, CD8+ T cell depletion did not increase the neurovirulence of an unrelated, attenuated HSV-1 glycoprotein (g)E- mutant. ICP47 is the first viral protein shown to influence neurovirulence by inhibiting CD8+ T cell protection.

T cell receptor-gamma/delta cells protect mice from herpes simplex virus type 1-induced lethal encephalitis.

Increased numbers of T cell receptor (TCR)-gamma/delta cells have been observed in animal models of influenza and sendai virus infections, as well as in patients infected with human immunodeficiency virus and herpes simplex virus type 1 (HSV-1). However, a direct role for TCR-gamma/delta cells in protective immunity for pathogenic viral infection has not been demonstrated. To define the role of TCR-gamma/delta cells in anti-HSV-1 immunity, TCR-alpha-/- mice treated with anti- TCR-gamma/delta monoclonal antibodies or TCR-gamma/delta x TCR-alpha/beta double-deficient mice were infected with HSV-1 by footpad or ocular routes of infection. In both models of HSV-1 infection, TCR-gamma/delta cells limited severe HSV-1-induced epithelial lesions and greatly reduced mortality by preventing the development of lethal viral encephalitis. The observed protection resulted from TCR-gamma/delta cell-mediated arrest of both viral replication and neurovirulence. The demonstration that TCR-gamma/delta cells play an important protective role in murine HSV-1 infections supports their potential contribution to the immune responses in human HSV-1 infection. Thus, this study demonstrates that TCR-gamma/delta cells may play an important regulatory role in human HSV-1 infections.

CD8(+) T cells can block herpes simplex virus type 1 (HSV-1) reactivation from latency in sensory neurons.

Recurrent herpes simplex virus type 1 (HSV-1) disease usually results from reactivation of latent virus in sensory neurons and transmission to peripheral sites. Therefore, defining the mechanisms that maintain HSV-1 in a latent state in sensory neurons may provide new approaches to reducing susceptibility to recurrent herpetic disease. After primary HSV-1 corneal infection, CD8(+) T cells infiltrate the trigeminal ganglia (TGs) of mice, and are retained in latently infected ganglia. Here we demonstrate that CD8(+) T cells that are present in the TGs at the time of excision can maintain HSV-1 in a latent state in sensory neurons in ex vivo TG cultures. Latently infected neurons expressed viral genome and some expressed HSV-1 immediate early and early proteins, but did not produce HSV-1 late proteins or infectious virions. Addition of anti-CD8alpha monoclonal antibody 5 d after culture initiation induced HSV-1 reactivation, as demonstrated by production of viral late proteins and infectious virions. Thus, CD8(+) T cells can prevent HSV-1 reactivation without destroying the infected neurons. We propose that when the intrinsic capacity of neurons to inhibit HSV-1 reactivation from latency is compromised, production of HSV-1 immediate early and early proteins might activate CD8(+) T cells aborting virion production.

Ex vivo model of leukocyte migration into herpes simplex virus-infected mouse corneas.

We are investigating neutrophilic infiltration into herpes simplex virus type 1 (HSV-1)-infected mouse corneas. Using a novel ex vivo corneal migration assay, we demonstrated two waves of neutrophil chemotaxis in HSV-1-infected corneas. An early chemotactic gradient developed within 48 h in concert with the appearance of HSV-1 epithelial lesions, peaked by 4 days post-infection (p.i.), and degraded by 6-8 days p.i.. A second chemotactic gradient appeared 10 days after infection, just before the initiation of chronic inflammation. The gradient was maintained in corneas that developed inflammation but rapidly degraded in corneas that failed to develop inflammation. The early chemotactic gradient was established in the absence of T lymphocytes, but the second chemotactic gradient was virtually abrogated by T cell depletion. We conclude that HSV-1 infection induces two temporally separated neutrophil chemotactic gradients in the mouse cornea: an early, transient, T cell-independent gradient; and a later, chronic, T cell-dependent gradient.

Contribution of virus and immune factors to herpes simplex virus type I-induced corneal pathology.

In vivo T-lymphocyte subpopulation depletion techniques were used to identify the roles of L3T4+ (CD4) and Lyt-2+ (CD8) T-lymphocytes in the pathogenesis of corneal stromal disease induced by two different strains of Herpes simplex virus type 1 (HSV-1). Histologic examination of infected corneas revealed significant differences in the composition of the inflammatory corneal infiltrates induced by the RE and KOS strains of HSV-1. The RE strain induced a predominantly polymorphonuclear leukocyte (PMN) infiltrate, which began approximately 1 week after infection and progressed through day 21. Depletion of CD4 cells before corneal infection with RE HSV-1 greatly reduced the incidence and severity of corneal disease; depletion of CD8 cells had no effect. The strain KOS HSV-1 induced an early PMN infiltrate that became predominantly mononuclear by day 21. Depletion of CD4 cells did not change the incidence or severity of KOS HSV-1-induced corneal stromal disease. The corneal lesions of these mice contained numerous CD8 cells. Depletion of CD8 cells before KOS HSV-1 infection of the cornea moderately reduced the incidence of stromal disease. However, in CD8-depleted mice with the disease, PMNs were the most prevalent infiltrating cells, and the disease appeared identical to that seen in RE HSV-1 infected corneas. Simultaneous depletion of CD4 and CD8 cells before KOS HSV-1 infection eliminated stromal disease. However, when T-cell depletion was discontinued in these mice, stromal disease developed in concert with the appearance of T-cells in the lymphoid organs and corneas. Thus, T-lymphocytes are a necessary component of HSV-1 corneal stromal disease. These results further suggest that RE HSV-1 preferentially activates CD4 cells in the cornea, and KOS HSV-1 preferentially activates CD8 cells in the cornea.

Lysis of herpes simplex virus (HSV) infected targets. IV. HSV-induced change in the effector population.

Herpes simplex virus type 1 (HSV-1) stimulation of peripheral blood lymphocytes from patients (PBL-P) with both recurrent corneal and oral-facial HSV-1 lesions results in altered natural killer (NK) activity. Freshly isolated PBL-P exhibit high lysis of HSV-1 infected allogeneic fibroblasts but low lysis of uninfected fibroblasts. After stimulation with HSV-1, PBL-P exhibit markedly increased lysis of uninfected fibroblasts such that the HSV-1 infected and uninfected fibroblasts are lysed with equal efficiency. The NK activity in freshly isolated PBL-P is mediated by effector cells that are phenotypically distinct from those responsible for the NK activity in HSV-1 stimulated PBL-P. In freshly isolated PBL-P most of the lysis of HSV-1 infected fibroblasts is mediated by lymphocytes expressing the phenotype OKM1+, OKT3+, and Leu-7+. After incubation with HSV-1, most of the lysis of HSV-1 infected and uninfected fibroblasts is mediated by lymphocytes that express the OKM1 antigen but lack detectable OKT3 or Leu-7 antigens. The change in function and phenotype appears to reflect in part the activation of Leu-7-, OKM1+, and OKT3- precursors of cytotoxic lymphocytes that are inactive in freshly isolated PBL-P.

Cellular immune function of patients with retinitis pigmentosa.

Although initial investigations of peripheral blood lymphocytes (PBL) from patients with retinitis pigmentosa (RP) demonstrated a reduced percentage of Leu-4 positive (pan-T) lymphocytes, the authors recently determined that the absolute number of Leu-4 positive cells per milliliter of blood is normal. This investigation studied the production of the lymphokines gamma-interferon (gamma-IFN) and interleukin-2 (IL-2) in cultures of concanavalin-A stimulated PBL. The difference in the mean production of gamma-IFN or IL-2 by PBL from 33 RP patients versus PBL from 16 controls did not achieve statistical significance at P less than 0.05. Overall, the data did not demonstrate significant cellular immune abnormalities in patients with RP.

Cell-mediated immune tolerance to HSV-1 antigens associated with reduced susceptibility to HSV-1 corneal lesions.

We have investigated the involvement of cell-mediated immune responses to Herpes simplex virus type 1 (HSV-1) in the pathogenesis of HSV-1 induced corneal stromal lesions in mice. Topical corneal (TC) HSV-1 infection induced a vigorous delayed hypersensitivity response, as well as lymphoproliferative and cytotoxic responses in the regional lymph nodes. The cytotoxic response involved HSV-1 specific and genetically restricted cytotoxic T lymphocytes, and activated natural killer cells. Half of the TC HSV-1 infected mice developed corneal stromal inflammation and scarring, the cause of visual morbidity in human herpetic disease. However, injection of HSV-1 into the ocular anterior chamber (AC) prior to, or simultaneously with, TC HSV-1 infection resulted in a profound state of cell-mediated immune tolerance of HSV-1 antigens. The tolerance was characterized by a substantial reduction in delayed hypersensitivity, lymphoproliferative, and cytotoxic responses to HSV-1 and was associated with virtually complete protection from corneal stromal lesions induced by HSV-1. These findings suggest a pathogenetic role for cell-mediated immunity and indicate the feasibility of preventing stromal disease through proper manipulation of the immune response.

Lysis of herpes simplex virus (HSV) infected targets. III. Effect of HSV on natural killer activity.

Freshly isolated peripheral blood lymphocytes from patients (PBL-P) with recurrent herpetic corneal and skin lesions, and controls (PBL-C) with no recollection of herpetic disease, effectively lysed herpes simplex virus type 1 (HSV-1) infected but not uninfected allogeneic fibroblasts. However, after 48 hr of stimulation with HSV-1, the lytic activity of PBL-C declined, while that of PBL-P increased. PBL-P and PBL-C produced similar amounts of interferon when stimulated with HSV-1. Thus, the decline in lytic activity by HSV-1 stimulated PBL-C was not due to a lack of interferon production. Interestingly, the lytic activity of HSV-1 stimulated PBL appeared to be directed against a component commonly expressed on both uninfected and HSV-1 infected fibroblasts. This was indicated by the fact that after incubation with HSV-1, PBL-P lysed the uninfected fibroblasts as effectively as the HSV-1 infected fibroblasts. The augmented lysis of uninfected fibroblasts was not due to infection of the "uninfected fibroblasts" by HSV-1 carried over or produced by the HSV-1 stimulated PBL. Furthermore, the capacity of HSV-1 to augment lytic activity was not diminished markedly by ultraviolet irradiation, suggesting that augmentation is due to the stimulatory capacity rather than the infectivity of the virus. Our data show that interaction of PBL-P with HSV-1 results in enhancement of nonspecific lytic activity and loss of the capacity to discriminate HSV-1 infected from uninfected allogenic fibroblasts.

The effect of cellular immune tolerance to HSV-1 antigens on the immunopathology of HSV-1 keratitis.

Previous studies have revealed that herpes simplex virus type 1 (HSV-1) corneal stromal lesions do not develop in the absence of a cell-mediated immune (CMI) response to HSV-1 antigens. HSV-1 glycoprotein C (gC) has been shown to play an important role in the induction of the cytotoxic T lymphocyte (CTL) response to HSV-1 infections at anatomical sites other than the eye. Here we report that a deletion mutant lacking gC (gC-39) when used to infect the corneas of A/J mice was a poor inducer of both CTL and delayed type hypersensitivity (DTH) responses. We have also followed histologically and immunohistochemically the course of HSV-1 stromal disease in A/J mice following topical corneal (TC) infection with wild type (WT) HSV-1, TC infection with gC-39 HSV-1, and simultaneous TC and anterior chamber (TC + AC) infection with WT HSV-1. The latter type of infection has been shown to induce a profound state of DTH and CTL tolerance of HSV-1 antigens. Following TC infection with WT HSV-1, stromal disease progressed to severe ulcerative keratitis with neovascularization by day 21. Histologic and immunohistochemical analysis revealed a predominantly mononuclear infiltrate consisting of numerous plasma cells as well as L3T4+ (T helper/inducer) and Lyt-2+ (T suppressor/cytotoxic) T lymphocytes. Following TC infection with gC-39, or simultaneous TC + AC infection with WT HSV-1, the severity of stromal disease did not progress beyond day 7. On day 21, there was at most a mild stromal cellular infiltrate consisting predominantly of polymorphonuclear neutrophils. These findings indicate that early stromal disease consists of a nonspecific inflammatory response, but severe stromal disease involves a CMI response to HSV-1. AC injection of HSV-1 inhibits the CMI response, thereby halting the progression of stromal disease. Similarly, gC-39, a poor inducer of CMI responses, is also a poor inducer of stromal disease.

Augmentation of intraocular inflammation by melanin.

The inflammatory response in endogenous uveitis or after anterior segment surgery was noted to be substantially greater in heavily pigmented eyes. Because varying amounts of melanin are released into the anterior chamber after intraocular inflammation, it was hypothesized that a proinflammatory effect of melanin might account for the enhanced inflammatory response in these eyes. To test this hypothesis, albino (BALB/c) or pigmented (C57BL/6) mice were challenged in the anterior chamber 2 weeks after a subcutaneous foot pad injection of horse serum or conalbumin dissolved in Freund's complete adjuvant. The degree of inflammation in the challenged eyes was determined by histologic examination 72 hr after the challenge. In all cases, the inflammatory infiltrate consisted mainly of polymorphonuclear leukocytes suggestive of an Arthus reaction. An anterior chamber challenge of horse serum-sensitized BALB/c or C57BL/6 mice with horse serum alone resulted in mild inflammation, which was augmented markedly by challenge with a combination of horse serum and melanin. The presence of melanin in the anterior chamber similarly increased the inflammatory response of conalbumin-sensitized mice to anterior chamber challenge with conalbumin. Melanin in the anterior chamber also significantly (P less than 0.05) augmented the inflammatory response of conalbumin-sensitized mice to a horse serum challenge, but it did not significantly augment the inflammatory response of horse serum-sensitized mice to a conalbumin challenge. The heterologous antigens induced minimal inflammation in the absence of melanin. Injection of melanin alone did not evoke an inflammatory response. Ocular challenge with melanin alone or in combination with antigen induced minimal inflammation in nonsensitized mice. However, preincubation of melanin with sera from horse serum-sensitized mice significantly increased its proinflammatory capacity when injected with horse serum into the anterior chamber of nonsensitized mice. In vitro binding studies using fluorescein isothiocyanate-conjugated mouse immunoglobulin G showed a high binding capacity of melanin for immunoglobulin G. It was concluded that the presence of free melanin in the anterior chamber can increase intraocular inflammation. Although the mechanism(s) by which melanin augments inflammation has not been defined, these data suggest that the binding of serum components (such as antibodies) to melanin may contribute to its proinflammatory effect.

The effect of flurbiprofen on herpes simplex virus type 1 stromal keratitis in mice.

The use of steroidal compounds to reduce the inflammation and scarring associated with herpes simplex virus type 1 (HSV-1) stromal keratitis can result in severe exacerbation of the corneal disease. We compared the nonsteroidal anti-inflammatory drug (NSAID) flurbiprofen sodium with dexamethasone for the treatment of HSV-1 induced corneal stromal disease in an inbred mouse model. Stromal disease was induced by the direct intrastromal injection of HSV-1. A stromal opacity and corneal neovascularization (CNV) developed in 100% of the injected eyes, with no epithelial involvement until late in the course, when the stromal disease was quite severe, such that it was possible to test the effectiveness of drugs in animals with an intact epithelium. Dexamethasone treatment had a variable effect on the severity of disease, ranging from a significant reduction in severity to significant exacerbation of disease, compared with placebo-treated controls. The most frequent effect of dexamethasone treatment was a worsening of corneal stromal opacities and CNV. In contrast, treatment with the NSAID did not exacerbate HSV-1 stromal disease. Flurbiprofen treatment resulted in a significant reduction of the maximum intensity of stromal opacity in some experiments, whereas in other experiments the effect was not statistically significant. In vitro studies of the effect of the anti-inflammatory drugs on HSV-1 replication in Vero cells revealed that both dexamethasone and flurbiprofen inhibited HSV-1 replication in a dose-dependent manner. Flurbiprofen had a greater inhibitory effect, which appears to be due, at least in part, to a direct virucidal effect. Dexamethasone did not exhibit virucidal activity.(ABSTRACT TRUNCATED AT 250 WORDS)

A reproducible method for injecting the mouse corneal stroma.

An improved delivery system for injecting the mouse corneal stroma was developed. This system incorporates the following features: a repeating dispenser that eliminates inaccuracies in depressing a syringe plunger, foot activation which frees both hands for manipulating the needle and permits constant observation of the injection site, and a flexible 30-cm, 32-gauge stainless steel needle with a 30 degrees bevel and a locking hub that resists pulsation due to back pressure while permitting freedom of motion by the operator. These injections were done while observing the cornea with a vertically mounted slit lamp, ideally suited for examining and photographing the eyes of laboratory animals. The reproducibility of the new delivery system, expressed in terms of the signal-to-noise ratio, was estimated and compared with that of a hand-held microsyringe by injecting a solution of radioactive chromium into the corneal stroma of A/J mice. The eyes were removed within 1 hr of injection, and the amount of chromium in each eye was determined in a gamma counter. The new delivery system had significantly (P less than 0.05) greater reproducibility than the hand-held syringe and could be calibrated to deliver up to 0.65 microliter to the mouse cornea.

Lymphocyte subpopulations and S-antigen reactivity in retinitis pigmentosa.

The lymphocyte subpopulations in the peripheral blood of 37 patients with retinitis pigmentosa (RP) and 24 controls were analyzed with the Leu series of monoclonal antibodies in conjunction with fluorescence-activated cell-sorter analysis. The peripheral blood lymphocytes (PBLs) from all RP genetic types had a significantly reduced frequency of Leu-4-positive T lymphocytes than controls, and a small but significant reduction in the frequency of Leu-3a-positive T lymphocytes was seen in patients with RP with the dominant trait. The reduced T-cell population seemed to be associated with an increased frequency of Leu-11a-positive cells. The PBLs from patients with RP did not react to retinal S-antigen, as assessed by the lymphocyte transformation or interleukin-2 assays. We conclude that patients with RP, although not clinically immunologically compromised, have a significantly reduced frequency of T lymphocytes in their PBLs. Furthermore, our study did not demonstrate reactivity to retinal tissue in PBLs from patients with RP.

Suppression of corneal neovascularization with cyclosporine.

We sought to determine if cyclosporine, which has been shown to suppress corneal allograft rejection, could also suppress corneal neovascularization induced by interleukin 2. Thirty A/J mice were treated with daily intramuscular injections of cyclosporine (25 mg/kg in olive oil) for 3 days before and 2 weeks following the intrastromal injection of 0.5 microL (5 IU) of recombinant mouse interleukin 2. Controls received intramuscular injections of olive oil. The mean area of corneal neovascularization 4, 8, and 12 weeks after injection was 9.2, 9.1, and 9.2 mm2, respectively, in controls, and 5.0, 5.2, and 5.2 mm2 in cyclosporine-treated mice (P less than .02; Student's t test). Cyclosporine causes a significant reduction in interleukin 2-induced corneal neovascularization that may, in part, account for its ability to prolong corneal allograft survival in high-risk cases.

An immunologist's view of herpes simplex keratitis: Thygeson Lecture 1996, presented at the Ocular Microbiology and Immunology Group meeting, October 26, 1996.

PURPOSE: To review recent progress in understanding the mechanisms of herpes simplex virus 1 (HSV-1)-induced immunopathology in the cornea, as revealed by studies in a mouse model. METHODS: The corneas of A/J mice were infected with 5 x 10(4) plaque-forming units (PFU) of the RE strain of HSV-1, and the development of inflammation was assessed by slitlamp, histologic, or immunohistochemical examination. The immunopathologic mechanisms were then defined by observing the effect of in vivo depletion of corneal Langerhans' cells, or T-lymphocyte subpopulations, or in vivo neutralization of cytokines on adhesion molecule expression or leukocytic infiltration of the infected cornea. RESULTS: After corneal infection, 60-70% of mice develop corneal opacity due to leukocytic infiltration, neovascularization, and edema. Polymorphonuclear neutrophils (PMN) represent 90% of the infiltrating cells, with numerous CD4+, but few CD8+, T cells present. Depleting CD4+ T cells of Langerhans' cells prevents inflammation from developing. Neutralizing interleukin-2 (IL-2) or interferon gamma (IFN-gamma) can prevent inflammation or cause a remission of existing disease. IFN-gamma neutralization causes a rapid block of PMN extravasation from the blood in association with reduced platelet endothelial cell adhesion molecule 1 (PECAM-1) expression on the corneal vascular endothelium. IL-2 neutralization results in decreased IFN-gamma production, reduced chemotaxis, and loss of PMN viability in the infected cornea. CONCLUSION: Herpes stromal keratitis is a CD4+ T cell-dependent inflammatory process in which PMN infiltration and destruction of the cornea are regulated, at least in part, by the T-helper type 1 (Th1) cytokines IL-2 and IFN-gamma.

Interleukin-2 induces corneal neovascularization in A/J mice.

Mitogen-stimulated lymphocytes induce a highly reproducible form of corneal neovascularization (CNV) in inbred mice. To determine if supernatants derived from stimulated lymphocytes and their constituent mediators were also angiogenic, we injected conditioned medium (CM) from mitogen-stimulated lymphocytes, control non-conditioned medium, recombinant interleukin-2 (rIL-2), or control bovine serum albumin (BSA), a component of rIL-2, into the corneas of syngeneic A/J mice. Control, nonconditioned medium was not angiogenic. While the other injections all induced some CNV, the CM and rIL-2 both induced a significantly greater area of CNV than BSA (p less than 0.05). The area of CNV induced by the CM was greater than that induced by IL-2 (p = 0.03). These data show that IL-2, which stimulates vascular endothelial cells in vitro and is elaborated during corneal immune reactions in vivo, is one of the potential mediators of immunologically mediated CNV.

Photodynamic therapy for corneal neovascularization.

We have previously described corneal neovascularization (CNV) induced by the intrastromal injection of interleukin-2 (IL-2) in inbred mice. Photodynamic therapy (PDT), administered by a deeply penetrating 630 nm fiberoptic laser, can destroy neoplasms and their associated neovascularization with some selectivity, but can damage neighboring tissues when used for CNV. We performed PDT with a 514 nm ophthalmic argon laser in an attempt to induce regression of CNV and reduce the associated toxicity. Eight weeks following IL-2 injection, mice with CNV were injected i.v. with dihematoporphyrin ether (DHE). Seventy-two hours later, 11 eyes (group I) were irradiated with eight 800 mW, 1000 mu, 2 s spots. Controls included 11 vascularized corneas from mice that received DHE but no laser (group II), 11 that received laser but no DHE (group III), and 35 untreated vascularized corneas (group IV). Comparison of the mean areas of CNV in groups I through IV pretreatment (6.0, 6.5, 6.7, and 7.6 mm2) and 12 weeks posttreatment (4.3, 6.3, 5.6, and 7.5 mm2) revealed that a significant decrease was seen in group I only (p less than 0.04, ANOVA). Complications in group I included blepharitis (9%) and iris damage (18%). Histologic studies revealed no evidence of posterior segment damage. PDT with the 514 nm laser is safe and efficacious for the treatment of IL-2-induced CNV in this model.

Corneal neovascularization. Pathogenesis and inhibition.

Corneal neovascularization (CNV) can cause significant visual loss because of the scarring and lipid deposition that frequently accompany it. In addition, penetrating keratoplasty in a vascularized recipient carries a significant risk of failure from allograft rejection. Frequently CNV is induced by nonspecific inflammatory stimuli, mediated primarily by polymorphonuclear neutrophils. Neovascularization can also be associated with specific corneal immune reactions, such as herpes simplex keratitis. Immunologically mediated CNV may be more amenable to treatment than CNV that results from nonspecific inflammation. Photodynamic therapy (PDT) following the intravenous injection of hematoporphyrin derivative or purified dihematoporphyrin ether (DHE) has been shown to suppress tumor growth and blood vessel growth in the eye. We have developed a murine model of CNV induced by the intrastromal injection of stimulated lymphocytes or interleukin-2 (IL-2). We have noted corneal DHE retention following its intravenous injection in mice with IL-2 induced CNV. Preliminary studies indicate that PDT can induce regression of CNV in these mice. Other recent studies that have enhanced our understanding of the pathogenesis and treatment of CNV are reviewed, and directions for future research are discussed.