Beauvericin-induced cholangiocyte apoptosis is mediated by activation of the ice-homologue CPP-32

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Abstract

Cholangiocytes (the epithelial cells that line the bile ductular apparatus) die in a variety of important liver diseases termed the cholangiopamies. We have recently developed an in vitro model of cholangiocyte apoptosis using beauvericin, a K+ ionophore. Because the interleukin IB converting enzyme (ICE) family of proteases have been implicated in apoptosis, our AIM was to determine if the ICE family of proteases participates in beauvericin-induced apoptosis of cholangiocytes. METHODS: In primary cultures of rat cholangiocytes, apoptosis was induced with beauvericin. Apoptosis was evaluated by morphologic (fluorescence microscopy) and biochemical (DNA agarose gel electrophoresis) criteria. Expression of three ICE protease family members (ICE, ICH-1, CPP-32) was evaluated using Northern blot analysis. Fluorogenic substrates were used to measure protease activity in cell lysates obtained from beauvericin-treated cells. RESULTS: Beauvericin caused apoptosis in cultured cholangiocytes in a time- (0-4 hrs) and concentration-dependent manner (0-25 μM). Inhibition of the interleukm 1-B converting (ICE) protease by the ICE inhibitor, cbz-VAD-cmk, inhibited apoptosis in a concentration-dependent (0-200 μM) manner with complete inhibition at 200 μM. Cholangiocytes expressed the message for ICE, ICH-1, CPP-32. Cleavage of the fluorogenic CPP-32 substrate, DEVD-AFC, increased following addition of beauvericin suggesting CPP-32 is activated. In contrast, no increase in ICE activity was observed using the ICE substrate YVAD-AMC. CONCLUSION: These data suggest that beauvericininduced apoptosis of cholangiocytes occurs by a CPP-32-dependent pathway.

Original languageEnglish (US)
Pages (from-to)A441
JournalFASEB Journal
Volume10
Issue number3
StatePublished - Dec 1 1996

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

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