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Brendan Marshall, Jason Covar, Sally Atherton, Ming Zhang; Prevention of Cytomegalovirus-Induced Retinitis By Intravenous Administration of Virus-specific siRNA. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1732. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
CMV is a ubiquitous betaherpesvirus which is present in a latent form throughout much of the human population without apparent clinical symptoms. However, during episodes of immune suppression, CMV may escape immune control and induce significant pathology of the eye, including retinal destruction and subsequently, blindness. Retinitis is characterized by both apoptosis and necrosis of the inner and outer retina and a characteristic feature of the apoptotic process is that most of the cells which undergo apoptosis are not infected with virus. We hypothesized that the delivery of virus-inhibiting compounds to the retinal pigment epithelium (RPE) layer of the eye might be beneficial in terms of limiting virus replication and also retinal damage caused by apoptosis of uninfected cells.
Using a murine cytomegalovirus (MCMV) model of infection we injected immunosuppressed Balb/c mice with MCMV via the supraciliary route. Two days following infection, we intravenously injected 1 nanomole of either control siRNA or siRNA specific for the MCMV immediate early protein-3 (IE-3). We then harvested eyes at several subsequent days for a period of two weeks in order to measure virus titers and levels of the IE-3 protein as well as the number of apoptotic cells in the retina, using both TUNEL assays and immunoblots specific for the cleaved caspase 3 protein.
IE-3 specific siRNA produced a substantial decrease in levels of the IE-3 protein as well as MCMV titers over the 14 day time course of virus infection, compared to control siRNA. The amount of apoptosis in anti IE-3 siRNA treated mice, as measured by TUNEL assay, was also greatly reduced compared to control siRNA treated mice. Furthermore, consistent with the results of TUNEL assays, levels of cleaved caspase 3 were also greatly reduced in IE-3 siRNA treated mice compared to those treated with control siRNA. Overall, retinitis was substantially decreased as a result of treatment with IE-3 specific siRNA. Our data indicate that an siRNA, which targets the MCMV IE-3 protein, is effective in substantially reducing the levels of both virus and apoptosis in MCMV infected eyes of immunosuppressed mice.
Intravenous injection of small RNA molecules helps to alleviate ocular damage induced by cytomegaloviruses and provides a route for alteration of host and/or virus gene expression in the retina.
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