Establishment of an immortalized human hepatic stellate cell line to develop antifibrotic therapies

Norikuni Shibata, Takamasa Watanabe, Teru Okitsu, Masakiyo Sakaguchi, Michihiko Takesue, Takemi Kunieda, Kenji Omoto, Shinichiro Yamamoto, Noriaki Tanaka, Naoya Kobayashi

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Abstract

Because human hepatic stellate cells (HSCs) perform a crucial role in the progress of hepatic fibrosis, it is of great value to establish an immortalized human cell line that exhibits HSC characteristics and grows well in tissue cultures for the development of antifibrotic therapies. Thus, we engineered an immortalized human hepatic stellate cell (HSC) line TWNT-4 by retrovirally inducing human telomerase reverse transcriptase (hTERT) into LI 90 cells established from a human liver mesenchymal tumor. Parental LI 90 entered replicative senescence, whereas TWNT-4 showed telomerase activity and proliferated for more than population doubling level (PDL) 200 without any crisis. TWNT-4 expressed platelet-derived growth factor-β receptor (PDGF-βR), α-smooth muscle actin (α-SMA), and type I collagen (α1) and was considered to be an activated form of HSCs. Treatment of TWNT-4 cells with either 100 U/ml of IFN-γ or 1 ng/ml of rapamycin (Rapa) for 14 days led to lower expression of type I collagen (α1) at RNA and protein levels. Exposure of TWNT-4 cells to both of IFN-γ (10 U/ml) and Rapa (0.1 ng/ml) for 14 days effectively decreased the expression of type I collagen (α1), PDGF-βR, and α-SMA expression and suppressed TGF-β1 secretion of TWNT-4 cells. We successfully induced apoptosis by transducing TNF-related apoptosis-inducing ligand (TRAIL) into TWNT-4 cells using adenovirus vectors Ad/GT-TRAIL and Ad/PGK-GV-17. These findings suggested that immortalized activated HSC line TWNT-4 would be a useful means to develop antifibrotic therapies.

Original languageEnglish
Pages (from-to)499-507
Number of pages9
JournalCell Transplantation
Volume12
Issue number5
Publication statusPublished - 2003

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Keywords

  • Human hepatic stellate cells
  • Human telomerase reverse transcriptase
  • Interferon-γ
  • Rapamycin
  • Type I collagen

ASJC Scopus subject areas

  • Cell Biology
  • Transplantation

Cite this

Shibata, N., Watanabe, T., Okitsu, T., Sakaguchi, M., Takesue, M., Kunieda, T., Omoto, K., Yamamoto, S., Tanaka, N., & Kobayashi, N. (2003). Establishment of an immortalized human hepatic stellate cell line to develop antifibrotic therapies. Cell Transplantation, 12(5), 499-507.