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The present study determined whether topical latanoprost, a prostaglandin (PG) F(2alpha) analog, influences the induction of anterior chamber-associated immune deviation (ACAID), corneal neovascularization (NV) or survival of corneal allografts in mice. BALB/c mice received topical latanoprost or PGE(2) once or multiple times daily starting 4 weeks prior to or the day of anterior chamber injection of C57BL/6 splenocytes. Induction of allo-specific ACAID was assessed by ear challenge with C57BL/6 splenocytes 1 week after subcutaneous immunization. In a separate experiment, orthotopic corneal transplantation was performed using C57BL/6 mice as donors and BALB/c mice as recipients. Recipients were randomized in a masked fashion to receive topical latanoprost or PGE(2). Graft fate was assessed clinically under surgical microscopy. Presence of MHC class II(+) CD11c(+) or CD11b(+) cells in normal BALB/c mouse eyes following latanoprost or PGE(2) administration was assessed immunohistochemically. Control mice received topical 20% dimethyl sulfoxide or no treatment. Allo-specific ACAID was induced after 2 or 6 weeks of once daily treatment with latanoprost, and was induced even after 6 weeks of multiple treatments with latanoprost. Conversely, mice receiving PGE(2) failed to develop ACAID. Opacity and corneal NV scores for allografts treated with latanoprost were statistically indistinguishable from those for control allografts (p>0.05), whereas all allografts treated with PGE(2) were rejected. Opacity and NV scores were significantly higher in these allografts than in controls (p<0.05). A number of MHC class II(+) CD11c(+) cells were present in the central cornea after PGE(2) treatment. Topical application of latanoprost does not influence induction of ACAID or graft outcomes including opacity and NV, whereas PGE(2) does. Immune privilege of corneal allograft is maintained after topical latanoprost application in mice.

Human amniotic epithelial cells (HAEC) may be a source of soluble anti-inflammatory factors. The purpose of this study is to determine the effect of topically applied HAEC culture supernatant on corneal inflammatory reactions. HAEC were obtained from a placenta and cultured for 48 hr, and the supernatant was collected. The conditioned medium from HAEC contained small amounts of human interleukin-1 receptor antagonist (IL-1ra). Intrastromal sutures were placed in the cornea of BALB/c mice to induce corneal neovascularisation. Superficial cauterisation was applied to induce recruitment or activation of antigen presenting cells (APCs) in the cornea without neovascularisation. HAEC conditioned medium, placebo, or recombinant human IL-1ra was topically applied three times daily for 2 weeks. Suture-induced corneal neovascularisation was evaluated microscopically for 8 weeks. The cauterised corneas were harvested at 2 weeks, and the MHC class II(+) APCs were quantified by immunofluorescent staining and confocal microscopy. Inflammatory cytokine gene expression in the cauterised corneas was analyzed by a multiprobe ribonuclease protection assay. Conditioned medium from HAEC led to a profound suppression of corneal neovascularisation and fewer MHC class II(+) APCs in the epithelium. In contrast, human IL-1ra was only slightly effective in suppressing corneal inflammatory reactions. mRNA expression of murine IL-1ra and IL-1beta in the cauterised corneas was markedly suppressed after application of the conditioned medium. These results suggest that HAEC are a source of soluble anti-inflammatory factors and that conditioned medium from HAEC contains factors other than IL-1ra that suppress corneal inflammation.

The purpose of this study was to investigate the role of CD80 and CD86 costimulatory molecules in corneal allograft rejection. Anti-CD80 and anti-CD86 monoclonal antibodies (mAbs) were administered after orthotopic corneal allograft transplantation. Graft rejection was observed by biomicroscopy. Population and localization of CD80(+)and CD86(+)cells in the cornea, cervical lymph nodes, and spleen were examined by flow cytometry and immunohistochemistry. The combined use of anti-CD80 and anti-CD86 mAbs was effective in prolonging corneal allograft survival. In the untreated mice bearing rejected graft, many CD86(+)and CD80(+)cells were found around the host-graft junctional area in the cornea, and CD86(high)cells were found in the cervical lymph node and spleen. In contrast, few CD86(+)or CD80(+)cells were observed in the cornea, cervical lymph node, and spleen from the mice treated with anti-CD80/CD86 mAbs. These results demonstrated a significant role of CD80 and CD86 costimulatory molecules in corneal allograft rejection., (Copyright 2002 Elsevier Science Ltd.)

To determine whether preservation of the donor cornea prevents allograft rejection, orthotopic corneal transplantation was performed using corneas preserved in storage medium (Optisol-GS((R))). Donor corneas harvested from C3H/He (H-2(k)) mice and B10.D2 (H-2(d)) mice were preserved in storage medium for 0, 3, 7 and 14 days, and then transplanted into the corneal beds of the recipient BALB/c (H-2(d)) mice. Graft survival was determined clinically and histologically. The expression of major histocompatibility complex (MHC) molecules in the preserved corneas was analysed by immunohistochemistry and Western blotting. Donor-specific cytotoxic T lymphocyte (CTL) and delayed-type hypersensitivity (DTH) responses were assessed 3 weeks after grafting. Active suppression of DTH in the recipient mice was also examined 3 weeks after grafting. The survival of 14 day preserved allografts was significantly higher than that of the non-preserved allografts in both MHC and minor histocompatibility (H) antigens, and minor H only disparate combination. The recipients of the preserved allografts failed to induce both CTL and DTH. The active suppression of DTH was not acquired in these recipients. The expression of donor-derived MHC class I antigens was markedly reduced in the corneas after preservation. Preservation of the donor cornea had a remarkable effect on the prevention of corneal allograft rejection. Since the preserved allografts failed to induce donor-specific CTL and DTH, and active suppression of DTH was not acquired in the recipients, the prevention of allo-rejection is due to a failure of allo-sensitization. These results indicate that the reduction of MHC class I antigens and minor H antigens expression in the preserved grafts induces a failure of allo-sensitization and leads to the high rate of acceptance in corneal allografts., (Copyright 2000 Academic Press.)

It has been reported that allograft rejection is mediated by a variety of adhesion molecules. Using a corneal allograft model in mice, we studied the role of very late antigen (VLA)-4 and leukocyte function-associated antigen (LFA)-1 adhesion molecules in corneal allograft rejection and the effects of monoclonal antibodies (mAbs) to them in suppressing corneal rejection. C3H/He donor corneas were transplanted into BALB/c corneal beds. The allografted mice were treated with a control mAb (M18/2), mAbs to VLA-4, or LFA-1 or their combination by i.p. injection until day 7. The expression of VLA-4, LFA-1, major histocompatibility complex (MHC) class II antigens, interleukin (IL)-2, IL-2 receptor and interferon gamma (IFNgamma) in the grafted cornea were studied immunohistochemically. Cytotoxic T lymphocyte (CTL) responses to donor alloantigens were assessed. The skins from a syngeneic donor or a third-part strain were transplanted 8 weeks after the initial keratoplasty onto the mice treated with anti-LFA-1 plus anti-VLA-4 mAbs. Fourteen of 16 allografts in non-treated mice and control mAb-treated mice became opaque by 2 weeks after transplantation. At 2 weeks, non-treated allografts showed expression of MHC class II antigens on keratocytes and mononuclear cells at the host-graft junction. Also, mononuclear cells expressing VLA-4, LFA-1, IL-2, IL-2 receptor and IFNgammawere present in the stroma at the host-graft junction. The allografts treated with either anti-VLA-4 or anti-LFA-1 alone, or anti-VLA-4 plus anti-LFA-1 remained transparent for more than 2 weeks, and the survival rates at 14 weeks was 0%, 16.7%, and 75.0%, respectively. The combined use of anti-VLA-4 and anti-LFA-1 mAbs prolonged graft survival significantly (P<0.05) at 14 weeks as compared with anti-LFA-1 mAb alone. At 3 weeks, CTL responses to donor alloantigens were depressed in mice treated with either anti-LFA-1 alone or anti-LFA-1 plus anti-VLA-4. Specific prolongation of donor-syngeneic skin was observed after treatment with the combination of these two mAbs. These results indicate that VLA-4 and LFA-1 have important roles in rejection process of corneal allografts, and that the combined use of mAbs to these molecules has remarkable effects on inducing alloantigen-specific immunosuppression in corneal transplantation.