Methods for Monitoring Gene Gun-Induced HBV- and HCV-Specific Immune Responses in Mouse Models

Abstract

The hepatitis B and C viruses (HBV/HCV) are major causes for chronic liver disease globally. For HBV new antiviral compounds can suppress the viral replication for years, but off-therapy responses are rare. Current therapies based on interferon and ribavirin can cure 45–85% of the treated HCV-infected patients largely depending on the viral genotype. New regimens including protease inhibitors will be introduced during 2011 and these will increase the cure rates for the hardest to treat HCV genotype 1 from 45 to 65%. Here a major need is to replace the immunomodulatory effects of interferon and/or ribavirin. Thus, therapeutic vaccines have a place in both chronic HBV and HCV infection. Unfortunately, none of these viruses can infect mice whereby substitute models are needed. We have used several types of murine models to predict the clinical efficacy of therapeutic vaccines for chronic HBV and HCV infections. In this chapter we describe transdermal delivery of genetic vaccines using the Helios Gene Gun device. A central role is that the model should have generally functional immune response, but with selective defects towards HBV and/or HCV. Thus, mice with stable integrated transgenes are useful. However, as a simple model to study the hepatic entry and functionality of a HBV- and/or HCV-specific immune responses other models are needed, where a killed transgenic hepatocyte is replaced by a healthy non-transgenic hepatocyte. Here we can effectively apply a technique termed hydrodynamic injection, which makes 10–30% of hepatocytes transiently transgenic for any plasmid. Within this chapter the methods used to characterize transiently transgenic mice are described. The main methods are the hydrodynamic injection technique, detection of transgene expression by immuno-precipitation, western blot, and immunohistochemistry. Finally, the in vivo functionality of T cells can be determined by using stably transfected syngeneic tumor cell lines expressing HBV and/or HCV proteins. The tumor challenge model enables studies of in vivo T cell function, whereas the cytotoxicity assay is used to determine T cell function in vitro. Overall, these models effectively reveal the efficiency by which various vaccine technologies, including biolistic DNA vaccination can kill the “infected” hepatocyte.