Chronic ethanol increases adeno-associated viral transgene expression in rat liver via oxidant and NF kappa B-dependent mechanisms

Recombinant adeno-associated virus (rAAV) transduction is limited in vivo, yet can be enhanced by hydroxyurea, ultraviolet-irradiation, or adenovirus coinfection, possibly via mechanisms involving stress in the host cell. Bec ause chronic ethanol induces oxidative stress, it was hypothesized that chr onic ethanol would increase rAAV transduction in vivo. To test this hypothe sis, rAAV encoding beta -galactosidase was given to Wistar rats that later received either ethanol diet or high-fat control diet via an enteral-feedin g protocol for 3 weeks. Expression and activity of beta -galactosidase in t he liver were increased nearly 5-fold by ethanol, The increase in transgene expression was inhibited by antioxidant diphenylene iodonium (DPI), which is consistent with the hypothesis that ethanol causes an increase in rAAV t ransduction via oxidative stress. Ethanol increased DNA synthesis only slig htly; however, it increased the nuclear transcription factor kappaB (NF kap paB) 4-fold, a phenomenon also sensitive to DPI. Moreover, a 6-fold increas e in rAAV transgene expression was observed in an acute ischemia-reperfusio n model of oxidative stress. Transgene expression was transiently increased 24 hours after ischemia-reperfusion 3 days and 3 weeks after rAAV infectio n. Further, adenoviral expression of superoxide dismutase or I kappaB alpha superrepressor inhibited rAAV transgene expression caused by ischemia-repe rfusion. Therefore, it is concluded that ethanol increases rAAV transgene e xpression via mechanisms dependent on oxidative stress, and NF kappaB likel y through enhancement of cytomegaloviral (CMV) promoter elements. Alcoholic liver disease is an attractive target for gene therapy because consumption of ethanol could theoretically increase expression of therapeutic genes (e .g., superoxide dismutase), Moreover, this study has important implications for rAAV gene therapy and potential enhancement and regulation of transgen e expression in liver.