Survival of liver failure pigs by transplantation of reversibly immortalized human hepatocytes with Tamoxifen-mediated self-recombination>

Toshinori Totsugawa, Chen Yong, Jorge David Rivas-Carrillo, Alejandro Soto-Gutierrez, Nalú Navarro-Alvarez, Hirofumi Noguchi, Teru Okitsu, Karen A. Westerman, Michinori Kohara, Michael Reth, Noriaki Tanaka, Philippe Leboulch, Naoya Kobayashi

Research output: Contribution to journalArticle

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Abstract

Background/Aims: Hepatocyte transplantation and bioartificial liver treatment are attractive alternatives to liver transplantation. The availability of well-characterized human hepatocyte lines facilitates such cell therapies. Methods: Human hepatocytes were immortalized with a retroviral vector SSR#197 expressing catalytic subunit of human telomerase reverse transcriptase (hTERT) and enhanced green fluorescent protein (EGFP) cDNAs flanked by a pair of loxP recombination targets. Then, Tamoxifen-dependent Cre recombinase was expressed in SSR#197-immortalized hepatocytes. Cre/LoxP recombination was performed in the established cells by simple exposure to 500 nM Tamoxifen for a week. Then, the reverted population of the cells was recovered by EGFP-negative cell sorting and characterized in vitro and in vivo using a pig model of acute liver failure (ALF) induced by d-galactosamine (0.5 g/kg) injection. Results: A human hepatocyte cell line 16T-3 was established. Reverted 16-T3 cells showed the increased expression of hepatic markers in association with enhanced levels of transcriptional factors. Compared to normal human hepatocytes, albumin production and lidocaine-metabolizing activities of reverted 16-T3 cells were 0.32 and 0.50-fold, respectively. Transplantation of reverted 16T-3 cells significantly prolonged the survival of ALF pigs. Conclusions: Here we demonstrate the usefulness of Cre/LoxP -mediated reversible immortalization of human hepatocytes with Tamoxifen-mediated self-recombination.

Original languageEnglish (US)
Pages (from-to)74-82
Number of pages9
JournalJournal of Hepatology
Volume47
Issue number1
DOIs
StatePublished - Jul 2007
Externally publishedYes

Fingerprint

Liver Failure
Tamoxifen
Genetic Recombination
Hepatocytes
Swine
Transplantation
Survival
Acute Liver Failure
Artificial Liver
Galactosamine
Cell- and Tissue-Based Therapy
Lidocaine
Liver Transplantation
Albumins
Catalytic Domain
Complementary DNA
Cell Line
Injections
Liver
Population

Keywords

  • Acute liver failure
  • Hepatocyte transplantation
  • Hepatocytes
  • Reversible immortalization

ASJC Scopus subject areas

  • Gastroenterology

Cite this

Totsugawa, T., Yong, C., Rivas-Carrillo, J. D., Soto-Gutierrez, A., Navarro-Alvarez, N., Noguchi, H., ... Kobayashi, N. (2007). Survival of liver failure pigs by transplantation of reversibly immortalized human hepatocytes with Tamoxifen-mediated self-recombination> Journal of Hepatology, 47(1), 74-82. https://doi.org/10.1016/j.jhep.2007.02.019

Survival of liver failure pigs by transplantation of reversibly immortalized human hepatocytes with Tamoxifen-mediated self-recombination> / Totsugawa, Toshinori; Yong, Chen; Rivas-Carrillo, Jorge David; Soto-Gutierrez, Alejandro; Navarro-Alvarez, Nalú; Noguchi, Hirofumi; Okitsu, Teru; Westerman, Karen A.; Kohara, Michinori; Reth, Michael; Tanaka, Noriaki; Leboulch, Philippe; Kobayashi, Naoya.

In: Journal of Hepatology, Vol. 47, No. 1, 07.2007, p. 74-82.

Research output: Contribution to journalArticle

Totsugawa, T, Yong, C, Rivas-Carrillo, JD, Soto-Gutierrez, A, Navarro-Alvarez, N, Noguchi, H, Okitsu, T, Westerman, KA, Kohara, M, Reth, M, Tanaka, N, Leboulch, P & Kobayashi, N 2007, 'Survival of liver failure pigs by transplantation of reversibly immortalized human hepatocytes with Tamoxifen-mediated self-recombination>', Journal of Hepatology, vol. 47, no. 1, pp. 74-82. https://doi.org/10.1016/j.jhep.2007.02.019
Totsugawa, Toshinori ; Yong, Chen ; Rivas-Carrillo, Jorge David ; Soto-Gutierrez, Alejandro ; Navarro-Alvarez, Nalú ; Noguchi, Hirofumi ; Okitsu, Teru ; Westerman, Karen A. ; Kohara, Michinori ; Reth, Michael ; Tanaka, Noriaki ; Leboulch, Philippe ; Kobayashi, Naoya. / Survival of liver failure pigs by transplantation of reversibly immortalized human hepatocytes with Tamoxifen-mediated self-recombination>. In: Journal of Hepatology. 2007 ; Vol. 47, No. 1. pp. 74-82.
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abstract = "Background/Aims: Hepatocyte transplantation and bioartificial liver treatment are attractive alternatives to liver transplantation. The availability of well-characterized human hepatocyte lines facilitates such cell therapies. Methods: Human hepatocytes were immortalized with a retroviral vector SSR#197 expressing catalytic subunit of human telomerase reverse transcriptase (hTERT) and enhanced green fluorescent protein (EGFP) cDNAs flanked by a pair of loxP recombination targets. Then, Tamoxifen-dependent Cre recombinase was expressed in SSR#197-immortalized hepatocytes. Cre/LoxP recombination was performed in the established cells by simple exposure to 500 nM Tamoxifen for a week. Then, the reverted population of the cells was recovered by EGFP-negative cell sorting and characterized in vitro and in vivo using a pig model of acute liver failure (ALF) induced by d-galactosamine (0.5 g/kg) injection. Results: A human hepatocyte cell line 16T-3 was established. Reverted 16-T3 cells showed the increased expression of hepatic markers in association with enhanced levels of transcriptional factors. Compared to normal human hepatocytes, albumin production and lidocaine-metabolizing activities of reverted 16-T3 cells were 0.32 and 0.50-fold, respectively. Transplantation of reverted 16T-3 cells significantly prolonged the survival of ALF pigs. Conclusions: Here we demonstrate the usefulness of Cre/LoxP -mediated reversible immortalization of human hepatocytes with Tamoxifen-mediated self-recombination.",
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AU - Soto-Gutierrez, Alejandro

AU - Navarro-Alvarez, Nalú

AU - Noguchi, Hirofumi

AU - Okitsu, Teru

AU - Westerman, Karen A.

AU - Kohara, Michinori

AU - Reth, Michael

AU - Tanaka, Noriaki

AU - Leboulch, Philippe

AU - Kobayashi, Naoya

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