A biliary HCO3 umbrella constitutes a protective mechanism against bile acid-induced injury in human cholangiocytes§

Authors

  • Simon Hohenester,

    1. Tytgat Institute for Liver and Intestinal Research, Department of Gastroenterology and Hepatology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands
    2. Department of Internal Medicine III, University of Aachen, Aachen, Germany
    Search for more papers by this author
    • These authors contributed equally.

  • Lucas Maillette de Buy Wenniger,

    1. Tytgat Institute for Liver and Intestinal Research, Department of Gastroenterology and Hepatology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands
    Search for more papers by this author
    • These authors contributed equally.

  • Coen C. Paulusma,

    1. Tytgat Institute for Liver and Intestinal Research, Department of Gastroenterology and Hepatology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands
    Search for more papers by this author
  • Sandra J. van Vliet,

    1. Department of Molecular Cell Biology and Immunology, VU Medical Center, Amsterdam, The Netherlands
    Search for more papers by this author
  • Douglas M. Jefferson,

    1. Tufts University School of Medicine, Boston, MA
    Search for more papers by this author
  • Ronald P. Oude Elferink,

    1. Tytgat Institute for Liver and Intestinal Research, Department of Gastroenterology and Hepatology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands
    Search for more papers by this author
  • Ulrich Beuers

    Corresponding author
    1. Tytgat Institute for Liver and Intestinal Research, Department of Gastroenterology and Hepatology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands
    • Department of Gastroenterology and Hepatology, Tytgat Institute for Liver and Intestinal Research, Academic Medical Center, University of Amsterdam, Meibergdreef 9, 1105 AZ Amsterdam, The Netherlands
    Search for more papers by this author
    • fax: +31-20-6917033


  • Potential conflict of interest: Nothing to report.

  • This work was supported by a research scholarschip from the Deutsche Forschungsgesellschaft (to S.H.; HO 4460/1-1), a Ph.D. scholarship from the University of Amsterdam Academic Medical Center (to L.M.W.), and research grants “Primary sclerosing cholangitis” from the Deutsche Crohn-Colitis-Vereinigung and the Norwegian Primary Sclerosing Cholangitis Foundation (to U.B.).

  • §

    Data were, in part, reported at the Annual Meeting of the American Association for the Study of Liver Diseases in Boston, MA, in November 2010, at the Annual Meeting of the European Association for the Study of the Liver in Berlin, Germany, in April 2011, and at the Falk® International Bile Acid Meeting in Freiburg, Germany, in October 2010 and were published, in part, in abstract form in HEPATOLOGY 2010;52(S1):923A, J Hepatol 2011;54(S1):S279, J Hepatol 2011;54(S1):S280, and Dig Dis 2011;29:62.

Abstract

Human cholangiocytes are continuously exposed to millimolar levels of hydrophobic bile salt monomers. We recently hypothesized that an apical biliary HCOmath image umbrella might prevent the protonation of biliary glycine-conjugated bile salts and uncontrolled cell entry of the corresponding bile acids, and that defects in this biliary HCOmath image umbrella might predispose to chronic cholangiopathies. Here, we tested in vitro whether human cholangiocyte integrity in the presence of millimolar bile salt monomers is dependent on (1) pH, (2) adequate expression of the key HCOmath image exporter, anion exchanger 2 (AE2), and (3) an intact cholangiocyte glycocalyx. To address these questions, human immortalized cholangiocytes and cholangiocarcinoma cells were exposed to chenodeoxycholate and its glycine/taurine conjugates at different pH levels. Bile acid uptake was determined radiochemically. Cell viability and apoptosis were measured enzymatically. AE2 was knocked down by lentiviral short hairpin RNA. A cholangiocyte glycocalyx was identified by electron microscopy, was enzymatically desialylated, and sialylation was quantified by flow cytometry. We found that bile acid uptake and toxicity in human immortalized cholangiocytes and cholangiocarcinoma cell lines in vitro were pH and AE2 dependent, with the highest rates at low pH and when AE2 expression was defective. An apical glycocalyx was identified on cholangiocytes in vitro by electron microscopic techniques. Desialylation of this protective layer increased cholangiocellular vulnerability in a pH-dependent manner. Conclusion: A biliary HCOmath image umbrella protects human cholangiocytes against damage by bile acid monomers. An intact glycocalyx and adequate AE2 expression are crucial in this process. Defects of the biliary HCOmath image umbrella may lead to the development of chronic cholangiopathies. (HEPATOLOGY 2012;55:173–183)

Ancillary