In Vivo Hepatocyte Retrovirus-Mediated Gene Transfer through the Rat Biliary Tract

Abstract

Delivering retroviruses targeted to hepatocytes in vivo involves the injection of retroviruses directly into the blood stream of the portal vein. The aim of this work was to delineate the conditions for delivering retroviruses 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 hepatectomy, 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 recombinant retroviruses encoding the beta -galactosidase gene. The virus total titer was 1.5 X 108 ffu (group I) or 6.5 X 108 ffu (groups II and III). The hepatic artery blood flow was either maintained (groups I and II) or interrupted (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 activity, predominantly in the periportal and mediolobular zones. This study demonstrates that hepatocytes of regenerating rat livers that have entered the S phase of the cell cycle as a result of partial hepatectomy can be transduced in vivo by retroviral vectors delivered in situ by bile duct perfusion. Furthermore, the number of transduced hepatocytes closely correlated with the viral total titer and was diminished by hepatic artery blood flow occlusion during perfusion.