In vivo hepatocyte retrovirus-mediated gene transfer through the rat biliary tract

Delivering retroviruses targeted to hepatocytes in vivo involves the inject ion of retroviruses directly into the blood stream of the portal vein. The aim of this work was to delineate the conditions for delivering retroviruse s in vivo by perfusing in situ the bile duct of the regenerating rat liver, and to study the hepatocyte transgene expression. At 24 hr after partial h epatectomy, during the S phase of the cell cycle, regenerating livers were perfused for 2.8 +/- 0.5 hr through the bile duct with 36.2 +/- 6.8 ml (0.3 +/- 01 ml/min) of fresh culture supernatant containing amphotropic recombi nant retroviruses encoding the beta-galactosidase gene. The virus total tit er was 1.5 x 10(8) ffu (group I) or 6.5 x 10(8) ffu (groups II and III). Th e hepatic artery blood flow was either maintained (groups I and II) or inte rrupted (group III) during bile duct perfusion. Liver biopsies taken 7 days later showed that 31.4 +/- 24.2% (group I), 58.7 +/- 23.6% (group II), and 45.1 +/- 21.4% (group III) of hepatocytes expressed beta-galactosidase act ivity, predominantly in the periportal and mediolobular zones, This study d emonstrates that hepatocytes of regenerating rat livers that have entered t he S phase of the cell cycle as a result of partial hepatectomy can be tran sduced in vivo by retroviral vectors delivered in situ by bile duct perfusi on. Furthermore, the number of transduced hepatocytes closely correlated wi th the viral total titer and was diminished by hepatic artery blood flow oc clusion during perfusion.