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G Peng, J Yang, G Tezel, RV Patil, MB Wax; Apoptosis of Activated T Lymphocytes Induced by Retinal Microglial Cells . Invest. Ophthalmol. Vis. Sci. 2002;43(13):3383.
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Purpose: We investigated the effects of rat retinal microglia on T cell viability using co-cultures of the rat retinal microglia and heat shock protein (hsp) antigen-activated T cells. Methods: Primary cultures of rat retinal and brain microglial cells were characterized by staining CD11b and GFAP antibodies using flow cytometry to assure homologous populations. Cell lines were stimulated by granulocyte macrophage-clonal stimulated factor (GM-CSF) prior to identification. The T cell line was obtained from rats immunized with hsp60, hsp27 or bovine serum albumin (BSA) one month prior to sacrifice, and were stimulated by hsp60 or hsp27 for several cycles of stimulation and proliferation. T cells and microglial cells were then co-cultured in the presence of the antigens for 12, 24 or 48 hours in culture medium. Apoptosis of co-cultured lymphocytes was assessed in these co-cultures by DNA ladder formation using agarose gel electrophoresis, DNA in situ end-labelling and Flow cytometry. Results: Using flow-cytometry, 90.4% and 93.2% of the retinal and brain microglia, respectively, were positive for CD11b, a marker of microglial cells. However, only 6.4% and 5.9% of retinal and brain microglial cells, respectively, were positive for GFAP, a marker of glial cells. While, a typical DNA ladder formation was observed in T cells co-cultured with retinal or brain microglial cells, no evidence of DNA fragmentation was present in retinal or brain microglial cells cultured alone. The numder of apoptosic cells increased as time of co-culture went on. Quantitative analysis of apoptosic cells showed that apoptosic cells accounted for 4∼5% after 12 hours, 10∼12% after 24 hours, 17∼18% after 48 hours. Conclusion: These results suggest that the retinal microglial cells play a role in inducing apoptosis of activated T cells. Further studies are in progress to identify the specific mechanisms involved in the apoptosis.
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