Stable gene expression with VSV-G pseudotyped-retrovirus vector in the ratliver

Background Pseudotyped-retrovirus-mediated gene transfer to the regeneratin g rat liver was investigated in vivo and the findings were compared with th ose for retrovirus-mediated gene transfer. Materials and methods. Four weeks prior to gene transfer, the spleen was tr anspositioned to the left subcutaneous position to develop a port-splenic s hunt. Twenty-four hours after a partial hepatectomy (68%) was performed, th e liver was perfused in situ and kept in contact with either a pseudotyped- retrovirus vector encoding LacZ (7 x 10(7) cfu/ml, Group 1) or a retrovirus vector encoding LacZ (1 x 10(4) cfu/ml, Group 2) for 30 min. The animals w ere sacrificed at various points after gene transfer, and X-gal staining, r eversed polymerase chain reaction (RT-PCR), and ONPG assay were performed t o detect the transferred LacZ cDNA. Results. In X-gal staining, the transferred LacZ cDNA started to show a str ong beta-galactosidase activity in 30 to 50% of the hepatocytes at 3 days a fter gene transfer. Positive staining continued to be recognized until 28 d ays with a slight decrease in its intensity thereafter. On the other hand, Group 2 animals showed weak staining, which was observed in about 10 to 15% of the hepatocytes from 3 days after gene transfer and then decreased ther eafter. In RT-PCR, positive mRNA of LacZ was detected constitutively until 28 days after gene transfer in Group 1, whereas two-thirds of the samples s howed a negative band in Groups 2 at 3 days after gene transfer. Conclusion. In conclusion, the pseudotyped-retrovirus vector was useful in establishing a stable and strong expression of the in vivo gene transfer, w hile targeting the regenerating liver. (C) 1999 Academic Press.