TGF-β1 Up-Regulates the Expression of PDGF-β Receptor mRNA and Induces a Delayed PI3K-, AKT-, and p70S6K-Dependent Proliferative Response in Activated Hepatic Stellate Cells

Authors

  • Ruchi Shah,

    1. Lipid Research Laboratory, VA Medical Center, Washington, District of Columbia
    2. Department of Biochemistry and Molecular Biology, The George Washington University Medical Center, Washington, District of Columbia
    Search for more papers by this author
  • Karina Reyes-Gordillo,

    1. Lipid Research Laboratory, VA Medical Center, Washington, District of Columbia
    2. Department of Biochemistry and Molecular Biology, The George Washington University Medical Center, Washington, District of Columbia
    Search for more papers by this author
  • Jaime Arellanes-Robledo,

    1. Lipid Research Laboratory, VA Medical Center, Washington, District of Columbia
    2. Department of Biochemistry and Molecular Biology, The George Washington University Medical Center, Washington, District of Columbia
    Search for more papers by this author
  • Carmen G. Lechuga,

    1. Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, New York
    2. Departamento de Oncología Molecular, Centro Nacional de Investigaciones Oncológicas, Madrid, Spain
    Search for more papers by this author
  • Zamira Hernández-Nazara,

    1. Department of Biochemistry and Molecular Biology, The George Washington University Medical Center, Washington, District of Columbia
    2. Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, New York
    3. Departamento de Fisiología, Universidad de Guadalajara, Guadalajara, Mexico
    Search for more papers by this author
  • Adam Cotty,

    1. Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, New York
    Search for more papers by this author
  • Marcos Rojkind,

    1. Department of Biochemistry and Molecular Biology, The George Washington University Medical Center, Washington, District of Columbia
    2. Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, New York
    3. Experimental Pathology and Immunology Sections, Department of Clinical Investigation, Walter Reed Army Medical Center, Washington, District of Columbia
    Search for more papers by this author
  • M. Raj Lakshman

    Corresponding author
    1. Lipid Research Laboratory, VA Medical Center, Washington, District of Columbia
    2. Department of Biochemistry and Molecular Biology, The George Washington University Medical Center, Washington, District of Columbia
    • Reprint requests: M. Raj Lakshman, PhD, Director of Research Labs, VA Medical Center, 50 Irving Street, NW, Washington, DC 20422; Tel.: 202-745-8330; Fax: 202-462-2006; E-mail: raj.lakshman@va.gov

    Search for more papers by this author

  • RS and KR-G contributed equally to this manuscript.
  • Marcos Rojkind: Deceased.
  • The opinions or assertions contained herein are the private views of the authors and are not to be construed as official or as reflecting the views of the Department of the Army or the Department of Defense.

Abstract

Background

Transforming growth factor beta 1 (TGF-β1) is a pleiotropic cytokine that activates hepatic stellate cell (HSC) proliferation, but inhibits parenchymal cell proliferation. Therefore, we hypothesize that TGF-β1 regulates HSC proliferation and elucidated its molecular action.

Methods

In order to elucidate the molecular mechanism whereby TGF-β1 up-regulates platelet derived growth factor beta (PDGF-β) receptor mRNA and induces a delayed proliferation of HSC, we used proliferation and apoptosis assays as well as RT-PCR, Western blot analysis, immunostaining, and flow cytometry in mouse and rat HSC.

Results

We show that TGF-β1 markedly induces the proliferation of mouse HSC in culture with concomitant 2.1-fold (p < 0.001) stimulation in [3H]-thymidine incorporation into cellular DNA. This induction is maximal between 24 and 36 hours postcytokine exposure that is triggered by 7.6-fold (p < 0.001) up-regulation of PDGF-β receptor mRNA and associated increase in PDGF-β receptor protein after 48 hours. TGF-β1-dependent HSC proliferation is mimicked by H2O2 that is inhibited by catalase, implying that TGF-β1 action is mediated via reactive oxygen species. HSC proliferation is blunted by PDGF-β receptor–neutralizing antibody as well as by specific inhibitors of PI3 kinase (PI3K), AKT, and p70S6K, indicating that the action of TGF-β1 involves the activation of PDGF-β receptor via the PI3K/AKT/p70S6K signaling pathway. TGF-β1 also induces a reorganization of actin and myosin filaments and cell morphology leading to the formation of palisades although their myosin and actin contents remained constant. These findings suggest that TGF-β1-mediated oxidative stress causes the transdifferentiation of HSC and primes them for extracellular matrix (ECM) deposition and scar contraction.

Conclusions

We conclude that liver injury up-regulates TGF-β1 that inhibits parenchymal cell proliferation, but stimulates HSC proliferation leading to the production of ECM and type I collagen resulting in fibrosis.

Ancillary