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C.A. Broderick, A.J. Smith, T. Georgiadis, J. Bainbridge, F. Schlichtenbrede, A. Thrasher, A.D. Dick, R.R. Ali; Control of Retinal Inflammation via Adeno-Associated Viral Gene Delivery . Invest. Ophthalmol. Vis. Sci. 2003;44(13):748.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: The immune privileged environment of the eye results, in part, from active control by immunoregulatory factors such as IL-10, TGFß, FasL and CD200. In spite of these (and other) specialised mechanisms, autoimmune uveoretinitis is a major cause of blindness. Current treatments are non-specific and cause systemic effects. In mice, experimental autoimmune uveitis (EAU) progression coincides with leucocyte infiltration and production of pro-inflammatory cytokines and cytotoxic mediators, resulting in the perpetuation of inflammation. Our aim is to evaluate the potential for AAV gene therapy in ocular inflammation. Methods: Immunological consequences of subretinal administration of AAV.GFP were assessed in mice. Local and systemic responses against the vector and GFP transgene were investigated by immunohistochemistry, proliferation assays, cytokine production and antibody production. Murine cDNAs encoding immunoregulatory genes were cloned, using established techniques, into AAV vectors. Expression of each vector product was investigated by ELISA and/or Western Blot and functional activity assessed following 293T cell transfection. Results: Following subretinal injections of AAV.GFP there was no increase in levels of anti-viral or anti-GFP antibodies at d14 to d80 post injection (pi). No neutralising antibodies were produced. There were no significant proliferative responses to viral proteins or GFP at d21pi or d21 post GFP expression. Only occassional leucocytic infiltration and activation were detected by immunohistochemistry. We have developed AAV vectors encoding murine FasL, IL-10, TGFß, IL-4 and IL-1Ra. Transgene expression was detected by ELISA/Western Blot and shown to be immunologically functional by the ability to induce jurkat cell apoptosis and inhibition of T cell proliferation in vitro. Conclusions: We have confirmed there are negligible immune responses in mice following subretinal AAV injection. A thorough investigation has shown that neither the viral vector nor the reporter gene product results in a deleterious immune response that would exacberate inflammation. These results support the use of the AAV vector system in modulating ocular inflammation. We have developed a series of AAV constructs encoding murine ligands and cytokines that are capable of regulating the activation status of immune cells and are now assessing the effect these signals have in murine EAU.
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