Cre/loxP-based reversible immortalization of human hepatocytes.
An ideal alternative to the primary human hepatocytes for hepatocyte transplantation would be to use a clonal cell line that grows economically in culture and exhibits the characteristics of differentiated, nontransformed hepatocytes following transplantation. The purpose of the present studies was to establish a reversibly immortalized human hepatocyte cell line. Human hepatocytes were immortalized with a retroviral vector SSR#69 expressing simian virus 40 large T antigen (SV40Tag) gene flanked by a pair of loxP recombination targets. One of the resulting clones, NKNT-3, showed morphological characteristics of liver parenchymal cells and expressed the genes of differentiated liver functions. NKNT-3 cells offered unlimited availability. After an adenoviral delivery of Cre recombinase and subsequent differential selection, efficient removal of SV40Tag from NKNT-3 cells was performed. Here we represent that elimination of the retrovirally transferred SV40Tag gene can be excised by adenovirus-mediated site-specific recombination.
Kobayashi
N
N
First Department of Surgery, Okayama University Medical School, Japan. immortal@md.okayama-u.ac.jp
Noguchi
H
H
Westerman
K A
KA
Watanabe
T
T
Matsumura
T
T
Totsugawa
T
T
Fujiwara
T
T
Leboulch
P
P
Tanaka
N
N
D002460:Cell Line; D017690:Cell Transplantation; D016895:Culture Media, Serum-Free; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D022781:Hepatocytes / Q000502:physiology* / Q000637:transplantation; D006801:Humans; D019426:Integrases / Q000378:metabolism*; D008099:Liver / Q000502:physiology; D011995:Recombination, Genetic; D012190:Retroviridae / Q000235:genetics / Q000378:metabolism; D014161:Transduction, Genetic; D014764:Viral Proteins / Q000378:metabolism*
D016895:Culture Media, Serum-Free; D014764:Viral Proteins; C045073:Cre recombinase; D019426:Integrases