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P. Jha, B. Matta, D. Y. Nounamo, P. S. Bora, N. S. Bora; Both Extrinsic and Intrinsic Pathways of Apoptosis Are Involved in the Resolution of Experimental Autoimmune Anterior Uveitis (EAAU). Invest. Ophthalmol. Vis. Sci. 2007;48(13):2645.
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Experimental autoimmune anterior uveitis (EAAU) is an organ-specific autoimmune disease of the eye and serves as an animal model of human idiopathic anterior uveitis. This study was undertaken to investigate the role of apoptosis in the resolution of EAAU.
EAAU was induced in Lewis rats by bovine melanin associated antigen (MAA). Animals were sacrificed at different time points during EAAU and apoptosis within the eye was monitored by TUNEL staining and flow cytometric analysis for annexin V positive cells. The expression of Caspase-3, -8, -9 was analyzed using RT-PCR and their activation was assessed using Western blot analysis. Levels of Bcl-2, Bax and PARP were also investigated using Western blot analysis. Cytochrome C release was monitored using immunoflourescent staining.
Very few TUNEL positive cells were detected during the early phase of EAAU, however, their number increased dramatically during the resolution of the disease. Flow cytometric analysis revealed that most of the apoptotic cells during the early phase of EAAU were at early stages of apoptosis while during the resolution of the disease most of the cells were late apoptotic. Interestingly, the up-regulation of caspase-8 and -9 preceded caspase-3 up-regulation indicating that both extrinsic and intrinsic pathways are activated. The levels of caspase-3 peaked during the resolution of EAAU. The expression of pro-apoptotic protein Bax was high whereas the expression of anti-apoptotic protein Bcl-2 was low during the resolution of EAAU. Furthermore, PARP cleavage products increased during the peak and resolution of EAAU. Finally, immunoflourescent staining demonstrated the release of cytochrome C in cell cytoplasm during the resolution of EAAU.
Our results suggest that both extrinsic and intrinsic pathways of apoptosis play a critical role in the resolution of uveitis by removing the inflammatory cells from eyes with EAAU. These results may have therapeutic potential for idiopathic autoimmune anterior uveitis and the administration of pro-apoptotic proteins may provide a novel therapeutic alternative to current treatments.
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