Stable gene expression with VSV-G pseudotyped-retrovirus vector in the rat liver.
BACKGROUND
Pseudotyped-retrovirus-mediated gene transfer to the regenerating rat liver was investigated in vivo and the findings were compared with those for retrovirus-mediated gene transfer.


METHODS
Four weeks prior to gene transfer, the spleen was transpositioned to the left subcutaneous position to develop a port-splenic shunt. Twenty-four hours after a partial hepatectomy (68%) was performed, the 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 were sacrificed at various points after gene transfer, and X-gal staining, reversed polymerase chain reaction (RT-PCR), and ONPG assay were performed to detect the transferred LacZ cDNA.


RESULTS
In X-gal staining, the transferred LacZ cDNA started to show a strong beta-galactosidase activity in 30 to 50% of the hepatocytes at 3 days after gene transfer. Positive staining continued to be recognized until 28 days 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 thereafter. 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 showed a negative band in Groups 2 at 3 days after gene transfer.


CONCLUSIONS
In conclusion, the pseudotyped-retrovirus vector was useful in establishing a stable and strong expression of the in vivo gene transfer, while targeting the regenerating liver.
Shiraishi
M
M
First Department of Surgery, University of the Ryukyus, Okinawa, 903-0215, Japan.
Tomori
H
H
Nagahama
M
M
Taira
K
K
Nozato
E
E
Sugawa
H
H
Ishida
A
A
Muto
Y
Y
D000818:Animals; D002863:Chromogenic Compounds; D005697:Galactosides; D015870:Gene Expression / Q000502:physiology*; D005822:Genetic Vectors / Q000235:genetics*; D007211:Indoles; D007763:Lac Operon / Q000235:genetics; D008099:Liver / Q000378:metabolism / Q000502:physiology*; D008562:Membrane Glycoproteins; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D012190:Retroviridae / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013194:Staining and Labeling; D014759:Viral Envelope Proteins / Q000235:genetics*; D001616:beta-Galactosidase / Q000378:metabolism
D002863:Chromogenic Compounds; C028391:G protein, vesicular stomatitis virus; D005697:Galactosides; D007211:Indoles; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D014759:Viral Envelope Proteins; D001616:beta-Galactosidase; C044888:5-bromo-4-chloro-3-indolyl beta-galactoside