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Juan Mo, Ming Zhang, Brendan Marshall, Jason Covar, Sally Atherton; Autophagy Is Anti-apoptotic during Murine Cytomegalovirus (MCMV) Infection of the Retina or RPE Cells. Invest. Ophthalmol. Vis. Sci. 2013;54(15):130. doi: https://doi.org/.
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The contribution of apoptosis and autophagy to the pathogenesis of cytomegalovirus infection has not been explored. The purpose of this study was to determine if MCMV infection affects apoptosis and autophagy during MCMV retinitis, and if so, how these processes influence the pathogenesis of MCMV infection.
In vitro, RPE cells were mock-infected or infected with MCMV at an MOI of 1 PFU/cell and treated with 10-6M of rapamycin or 10-5M of chloroquine. Mock-infected and MCMV-infected RPE cells were collected and examined by western blot, electron microscopy, plaque assay and trypan blue exclusion assay. In vivo, immunosuppressed (IS) Atg5flox/+, nestin-Cre mice or Atg5+/+, nestin-Cre mice were inoculated with 1×104 PFU of MCMV K181 via a supraciliary injection. Inoculated eyes were collected at several times p.i., examined by MCMV staining, H&E staining, TUNEL assay and western blotting.
In vitro, the levels of LC3B-II were increased in MCMV infected RPE cells, which was consistent with EM results showing increased accumulation of autophagic vacuoles in MCMV infected cells compared to uninfected control cells. Autophagy was increased while caspase 3-mediated apoptosis was decreased by rapamycin treatment (blocks mTOR signaling pathway). In contrast, autophagy was decreased and caspase 3-mediated apoptosis was increased by chloroquine treatment. In vivo, When the number of MCMV and TUNEL positive cells was compared in the retina of Atg5flox/+, nestin-Cre infected mice with that in Atg5+/+, nestin-Cre infected mice, Atg5flox/+, nestin-Cre infected mice had more MCMV and TUNEL positive cells in the retina than Atg5+/+, nestin-Cre infected mice at early stage of infection. Disruption of the architecture of the retinas of infected eyes in Atg5flox/+, nestin-Cre mice was more severe than that observed in Atg5+/+, nestin-Cre mice.
Rapamycin induced autophagy through blocking mTOR signaling pathway and decreased caspase 3-mediated apoptosis while chloroquine treatment had the opposite effects. The retinas of infected eyes in Atg5flox/+, nestin-Cre mice had more severe disruption than that of Atg5+/+, nestin-Cre mice. Taken together, these results suggest that autophagy during MCMV infection is anti-apoptotic and contributes to maintenance of the retinal structure.
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