Cre/loxP-based reversible immortalization of human hepatocytes

N. Kobayashi; H. Noguchi; K. A. Westerman; T. Watanabe; T. Matsumura; T. Totsugawa; T. Fujiwara; P. Leboulch; N. Tanaka

doi:10.3727/000000001783986558

Cre/loxP-based reversible immortalization of human hepatocytes

N. Kobayashi, H. Noguchi, K. A. Westerman, T. Watanabe, T. Matsumura, T. Totsugawa, T. Fujiwara, P. Leboulch, N. Tanaka

Research output: Contribution to journal › Article › peer-review

19 Citations (Scopus)

Abstract

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.

Original language	English
Pages (from-to)	383-386
Number of pages	4
Journal	Cell Transplantation
Volume	10
Issue number	4-5
DOIs	https://doi.org/10.3727/000000001783986558
Publication status	Published - 2001

Keywords

Cre/loxP system
Human hepatocytes
Reversible immortalization
Simian virus 40 large T antigen

ASJC Scopus subject areas

Biomedical Engineering
Cell Biology
Transplantation

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title = "Cre/loxP-based reversible immortalization of human hepatocytes",

abstract = "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.",

keywords = "Cre/loxP system, Human hepatocytes, Reversible immortalization, Simian virus 40 large T antigen",

author = "N. Kobayashi and H. Noguchi and Westerman, {K. A.} and T. Watanabe and T. Matsumura and T. Totsugawa and T. Fujiwara and P. Leboulch and N. Tanaka",

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doi = "10.3727/000000001783986558",

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AU - Kobayashi, N.

AU - Noguchi, H.

AU - Westerman, K. A.

AU - Watanabe, T.

AU - Matsumura, T.

AU - Totsugawa, T.

AU - Fujiwara, T.

AU - Leboulch, P.

AU - Tanaka, N.

PY - 2001

Y1 - 2001

N2 - 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.

AB - 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.

KW - Cre/loxP system

KW - Human hepatocytes

KW - Reversible immortalization

KW - Simian virus 40 large T antigen

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Cre/loxP-based reversible immortalization of human hepatocytes

Abstract

Keywords

ASJC Scopus subject areas

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