Geeta Pararajasegaram

Hematopoietically derived retinal perivascular microglia initiate uveoretinitis in experimental autoimmune uveitis.

Abstract Background: Hematopoietically derived cells in the retina were studied for the expression of molecules associated with antigen presentation. Methods: Bone marrow cells of (Lewis × Brown Norway) F1 rats (LBNF1) were transplanted to sublethally irradiated Brown Norway (BN) rats to construct chimeric rats (LBNF1→BN). Each of 21 established chimeras received an adoptive transfer of uveitogenic Lewis T lymphocytes. Three rats were killed on each of 7 consecutive days. The right eye of each rat was processed for flat-mount preparation of the retina; the left eye of each was frozen for cryostat sectioning. All tissues were then stained with one of the following antibodies: OX-3 (Lewis-specific MHC class II marker), anti-ICAM, anti-B7-1, anti-TNF-α or anti-IL-1β. Results: Initial clinical signs of EAU appeared first on day 4; by day 6, full-blown EAU was noted. The flat-mount preparations revealed the presence of OX-3+ cells in the retina, perivascularly exhibiting dendritic morphology on day 2. These cells were observed in the retinal nerve fiber layer (NFL). No B7-1+, ICAM-1+, TNF-α+ or IL-1β+ cells were detected. Cryostat sections revealed positive cell staining of perivascular microglia and astrocytes in the retinal NFL with anti-IL-1β and anti-TNF-α antibodies. Conclusions: Since only perivascular bone marrow-derived cells are seen to express MHC class II molecules prior to onset of EAU, and since these cells also generate the cytokines IL-1β and TNF-α, it appears that initial presentation of antigen in the retina could be by these cells.

Small Heat Shock Protein αA-Crystallin Prevents Photoreceptor Degeneration in Experimental Autoimmune Uveitis

The small heat shock protein, αA-crystallin null (αA−/−) mice are known to be more prone to retinal degeneration than the wild type mice in Experimental Autoimmune Uveoretinitis (EAU). In this report we demonstrate that intravenous administration of αA preserves retinal architecture and prevents photoreceptor damage in EAU. Interestingly, only αA and not αB-crystallin (αB), a closely related small heat shock protein works, pointing to molecular specificity in the observed retinal protection. The possible involvement of αA in retinal protection through immune modulation is corroborated by adaptive transfer experiments, (employing αA−/− and wild type mice with EAU as donors and Rag2−/− as the recipient mice), which indicate that αA protects against the autoimmune challenge by modulating the systemic B and T cell immunity. We show that αA administration causes marked reduction in Th1 cytokines (TNF-α, IL-12 and IFN-γ), both in the retina and in the spleen; notably, IL-17 was only reduced in the retina suggesting local intervention. Importantly, expression of Toll-like receptors and their associated adaptors is also inhibited suggesting that αA protection, against photoreceptor loss in EAU, is associated with systemic suppression of both the adaptive and innate immune responses.

Anti-inflammatory effects of vitamin E on experimental lens-induced uveitis.

The anti-phlogistic effect of dietary vitamin E supplementation on the acute inflammation observed in experimental lens-induced uveitis in Brown Norway rats was studied. The effects of vitamin E were examined using histopathologic parameters as well as by measuring the levels of arachidonic acid metabolites. Histologic examination of the eyes revealed that the vitamin E-deficient animals had the most severe destruction of the retina, while those animals receiving the vitamin E-supplemented diet exhibited the best preservation of the retinal architecture. Levels of arachidonic acid metabolites, as determined by radioimmunoassay, were significantly higher in vitamin E deficient rats as compared with rats on a normal diet.

Suppression of S antigen-induced uveitis by vitamin E supplementation

The anti-inflammatory effects of vitamin E were investigated using the S antigen model of uveoretinitis. Thirty-six 3-week-old Lewis rats were separated into three groups and maintained on a specially formulated diet. One group of animals received a diet deficient in vitamin E; a second group received a normal diet containing vitamin E, and the third group, in addition to receiving the normal diet, received vitamin E supplementation. At 9 weeks of age, all rats were sensitized to S antigen. Six animals in each group were killed on day 14 and the remaining animals on day 21 following immunization. Both histopathologic and biochemical studies were conducted to evaluate the tissue damage observed in animals maintained on different dietary levels of the vitamin. The intraocular inflammation in the vitamin E-supplemented group was considerably smaller than in the other two groups (p less than 0.01). The former group had the highest level of vitamin E in both the eye and plasma (mean value 1.13 micrograms/mg protein and 23.9 micrograms/ml, respectively), while the vitamin E-deficient group had the lowest levels (mean values of 0.16 micrograms/mg protein and 0.48 micrograms/ml in the eye and plasma, respectively). Results of the radioimmunoassay for the determination of the arachidonic acid metabolites revealed significantly lower levels of thromboxane B2 in the vitamin E-supplemented group (2.04 +/- 0.45 pg/mg) than in the normal (4.33 +/- 0.98 pg/mg) or the vitamin E-deficient (5.21 +/- 1.12 pg/mg) groups (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Treatment of S Antigen Uveoretinitis with Lipoxygenase and Cyclo-Oxygenase Inhibitors

The role of metabolites of arachidonic acid (AA) in experimental autoimmune uveoretinitis was studied using inhibitors of AA metabolism. Nordihydroguaiaretic acid (NDGA), which inhibits predominantly the lipoxygenase (LO) pathway, or indomethacin, a specific cyclo-oxygenase (CO) inhibitor, was administered to rats immunized with retinal S antigen. Levels of various AA metabolites were measured in the inflamed uvea, and the severity of intraocular inflammation was quantitated by morphometric analysis. Histopathologically, the uveoretinitis was significantly suppressed following treatment with NDGA, while indomethacin treatment resulted in augmentation of the disease (p less than 0.05). These results tend to indicate that the inhibition of the LO rather than the CO pathway may be more beneficial in the treatment of autoimmune uveitis.

Inhibition of interphotoreceptor retinoid-binding protein-induced uveitis by monoclonal antibodies

Immunization of Lewis rats with interphotoreceptor retinoid-binding protein (IRBP) resulted in development of uveoretinitis in 100% (12/12) of the animals so injected. In the present study, we attempted immunotherapy of this intraocular inflammation using the monoclonal antibodies anti-I-A and anti-I-E. The antibodies were injected 1 day before and on days 1 and 2 after IRBP sensitization, and the animals were killed 16 days later. Anti-I-A treatment inhibited the disease in 83% (10/12) of the animals, while anti-I-E antibodies resulted in inhibition of the disease in 67% (8/12) of the animals. All rats injected with preimmune IgG (control animals) developed uveoretinitis. The anti-I-A and anti-I-E antibodies exerted a specific suppressor effect on the immune response of these animals. Thus, anti-I-A and anti-I-E antibodies administered at the time of antigen priming abolished the immune function and inhibited the development of autoimmune uveitis.

Histochemical Analysis of Experimental Granulomatous Uveitis

We have previously demonstrated the effects of various inhibitors of arachidonic acid metabolism on experimental lens-induced granulomatous uveitis. In the present study, we investigated the effect of these same inhibitors on the expression of lysosomal enzymes at different stages of choroidal inflammation in experimental lens-induced granulomatous uveitis and compared this to the inflammation observed at each stage examined. Lysosomal enzymes such as acid phosphatase, beta-glucuronidase and succinate dehydrogenase are known to be liberated during the maturation of mononuclear phagocytes to epithelioid cell granulomas. Although animals treated with nordihydroguaiaretic acid showed less severe inflammation than did indomethacin-treated or control animals, none of these agents appeared to affect the expression of acid phosphatase and beta-glucuronidase, as determined histochemically. Succinate dehydrogenase could not be detected in any of the eyes examined, even though sections of liver and kidney from these same animals were positive for this enzyme.