Evaluation of promoter strength in mouse and rat primary hepatocytes using adenovirus vectors.
Primary cultured hepatocytes are widely used in the studies of basic and clinical hepatology. Finding an efficient method for gene transfer into primary hepatocytes will be an important advance for these studies. In the present study, we evaluated the activity of an adenovirus vector including promoters for the Rous sarcoma virus (RSV), elongation factor 1alpha, and cytomegalovirus (CMV) as well as the beta-actin promoter/CMV enhancer (CA) using beta-galactosidase as a reporter gene. Although RSV and elongation factor 1alpha promoters had low transcriptional activity in hepatocytes, the CA and CMV promoters had high activity. The CA promoter was the most active, mediating 50.3- and 204.4-fold more activity than the RSV promoter in mouse and rat hepatocytes, respectively. Dose-response studies revealed that transgene activity can be controlled by as much as 1000-fold, by selection of the promoter and the number of infectious particles per cell. These findings should help in the construction of adenovirus vectors for expressing genes of interest in rodent primary cultured hepatocytes.
Arita
Eri
E
Laboratory of Bio-Functional Molecular Chemistry, Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Osaka, Japan.
Kondoh
Masuo
M
Isoda
Katsuhiro
K
Nishimori
Hikaru
H
Yoshida
Takeshi
T
Mizuguchi
Hiroyuki
H
Yagi
Kiyohito
K
D000199:Actins / Q000235:genetics; D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000235:genetics; D017930:Genes, Reporter; D005822:Genetic Vectors; D022781:Hepatocytes / Q000378:metabolism*; D007763:Lac Operon; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D020648:Peptide Elongation Factor 1 / Q000235:genetics; D011401:Promoter Regions, Genetic; D051381:Rats; D017207:Rats, Sprague-Dawley; D052736:Rous sarcoma virus / Q000235:genetics; D014158:Transcription, Genetic; D014161:Transduction, Genetic; D019076:Transgenes; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D000199:Actins; D020648:Peptide Elongation Factor 1; D001616:beta-Galactosidase