Development and initial application of an in vitro model of
apoptosis in rodent cholangiocytes.
Quea, Florencia G., Gregory J. Goresb, Nicholas F. Larussob.
Center for Basic Research in Digestive Diseases, Division of
Gastroenterologic and General Surgery, Division of Gastroenterology
and Internal Medicine
APStracts 3:0151G, 1996.
While histologic data suggest that cholangiocytes die by apoptosis in
human liver diseases, no information exists on the mechanisms of
cholangiocyte apoptosis. Thus, our aims were to establish an in vitro
model of cholangiocyte apoptosis and to test the hypothesis that
changes in intracellular ions would cause apoptosis in cholangiocytes
by a protease-sensitive pathway. A large number of pro-apoptotic
agents were ineffective in inducing apoptosis in rat or human
cholangiocytes in culture; in contrast, beauvericin, a K+ ionophore,
caused apoptosis in both cell lines despite their expression of Bcl
-2. Although beauvericin decreased intracellular K+ and increased
intracellular Ca+2, abolishing the K+ gradient did not prevent
beauvericin-induced apoptosis; in contrast, omission of extracellular
Ca+2 inhibited apoptosis by 42%. The ICE family protease inhibitor,
Cbz-Val-Ala-Asp-cmk, inhibited apoptosis in a concentration-dependent
manner. By Northern blot analysis, cholangiocytes expressed the mRNAs
for three members of the ICE protease family: ICE, Ich-1, and CPP-32.
Cleavage of a substrate for CPP-32-like protease activity, but not a
substrate for ICE and Ich-1, increased following beauvericin
treatment. In summary, we have established an in vitro model of
apoptosis in cholangiocytes. Our data suggest that beauvericin
-induced apoptosis occurs by a Ca+2-dependent, CPP-32 protease
-sensitive pathway despite cholangiocyte expression of Bcl-2.
Received 26 April 1996; accepted in final form 28 July 1996.
APS Manuscript Number G162-6.
Article publication pending Am. J. Physiol. (Gastrointest. Liver
Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 21 August 1996