In situ perfusion of the liver enhances the efficiency of retrovirus-mediated gene transfer to hepatocytes.
To increase the efficiency of retrovirus-mediated gene transfer targeting an individual's liver in vivo, the liver was perfused in situ with the retrovirus vector during hepatic cold ischemia. Four weeks prior to gene transfer, the spleen was transpositioned to the left subcutaneous position to develop a portosplenic shunt, which was performed in order to prevent intestinal congestion during hepatic ischemia. Traditional retrovirus vectors (1 x 10(5) CFU/ml) which encode genes for the Escherichia coli beta-galactosidase (LacZ) were used in this study. Twenty-four hours after partial hepatectomy (70%), the remnant liver was surgically isolated, perfused with 1 ml of vector solution through the portal vein, and kept in contact with the vector for 30 min under cold ischemia (group 1). Hepatic ischemia could thus be performed without any intestinal congestion, due to the preestablished portosystemic shunt. In group 2, the liver was perfused with 1 ml of vector solution through the portal vein without in situ perfusion of the liver. Animals were sacrificed 1, 3, 7 and 28 days after gene transfer. In X-gal staining, the transferred LacZ was detected positive in 10-15% of the hepatocytes only in group 1, 3 days after gene transfer. Graft histology and a liver function test showed no difference between both groups 24 h after gene transfer. In conclusion, in situ perfusion of the liver greatly enhanced the efficacy of retrovirus-mediated gene transfer, targeting an individual's liver in vivo.
Tomori
H
H
Departments of Surgery I and Clinical Laboratory Medicine, Faculty of Medicine, University of the Ryukyus, Nishihara-cho, Okinawa, Japan.
Shiraishi
M
M
Muto
Y
Y
D000818:Animals; D018014:Gene Transfer Techniques; D008099:Liver / Q000166:cytology / Q000378:metabolism* / Q000503:physiopathology; D010477:Perfusion; D051381:Rats; D017208:Rats, Wistar; D012190:Retroviridae / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction