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P. B. Thomas, D. M. Samant, Y. Wang, S. Selvam, D. Stevenson, A. K. Mircheff, J. E. Schechter, F. Apparailly, M. D. Trousdale; Adeno-Associated Virus Vector Mediated vIL-10 Gene Therapy for Autoimmune Dacryoadenitis in a Rabbit Model. Invest. Ophthalmol. Vis. Sci. 2009;50(13):3654.
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To evaluate the effect of adeno-associated virus (AAV) vector mediated vIL-10 gene expression on lacrimal gland immunopathology and ocular surface disease in a rabbit model of induced autoimmune dacryoadenitis (ID).
An inferior LG was excised from each rabbit, epithelial cells were purified and cultured for 2 days, gamma irradiated, and then co-cultured for 5 days with autologous peripheral blood lymphocytes. Autoimmune dacryoadenitis was induced in rabbits by autoadoptive transfer of activated PBL from the autologous mixed cell reaction. Schirmer test, tear break up time (BUT) and rose Bengal staining were done to assess clinical disease. After confirming disease at 4 weeks an AAV vector carrying the vIL-10 gene was injected into the remaining inferior LG of the treatment group (ID/Rx). All LG were removed for analysis at 16 weeks post disease induction. Cryosections were immunostained to quantify the CD4, CD8, RTLA (Rabbit T Lymphocyte Antigen), MHC II and CD18 expression. One portion of the gland was used for cytokine analysis by real time RT-PCR.
The tear production was decreased in the ID group and partially restored within two weeks of Rx. Tear BUT and rose Bengal score also showed overall improvement by the end of the study. Histopathological examination of ID group’s LG revealed some large scattered lymphocytic foci and areas of altered or distorted acini. In the ID/Rx group the lacrimal glands had scattered small lymphocytic foci around ducts and venules. The CD4:CD8 ratio was approximately 1:1 in normal animals, 8:1 in ID animals, and 0.4:1 in the ID/Rx group. CD18 expression was almost 5 fold less in the ID/Rx group. There was no significant difference between TNF-alpha and IFN-gamma mRNA transcripts in the ID and ID/Rx group. However, IL-10, IL-4 and IL-6 transcripts increased in ID/Rx group compared to ID group.
Animals with experimentally induced autoimmune dacryoadenitis appear to benefit from AAV mediated vIL-10 gene therapy. Tear production increased, rose Bengal staining decreased and lacrimal gland histopathology decreased following gene therapy. The cytokine expression profile suggested that the response may have involved induction of CD8+ regulatory cells.
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