Generation of hydroperoxides in isolated rat hepatocytes and hepatic mitochondria exposed to hydrophobic bile acids.
OBJECTIVE
The mechanisms causing liver injury in cholestatic diseases are unclear. The hypothesis that accumulation of hydrophobic bile acids in hepatocytes during cholestasis leads to generation of oxygen free radicals and oxidative injury was tested. The aim of this study was to determine if hydrophobic bile acid toxicity is associated with increased hydroperoxide generation in isolated rat hepatocytes and mitochondria.


METHODS
Hepatocytes were exposed to taurochenodeoxycholic acid (TCDC; 0-2000 mumol/L) or taurocholic acid (TC; 1000 mumol/L), and cellular injury, intracellular hydroperoxide generation, and thiobarbituric acid-reacting substances (TBARS) were measured. Isolated mitochondria were incubated with 400 mumol/L chenodeoxycholic acid or 400 mumol/L cholic acid, and hydroperoxide generation was measured fluorometrically.


RESULTS
Hepatocyte injury, hydroperoxide generation, and TBARS increased over 4 hours on exposure to TCDC but not TC. Hydroperoxide generation preceded hepatocyte injury and accumulation of TBARS. Preincubation of hepatocytes with the antioxidant, d-alpha-tocopheryl succinate, completely abrogated cellular injury, hydroperoxide, and TBARS generation. Hydroperoxide generation was increased in mitochondria exposed to chenodeoxycholic acid.


CONCLUSIONS
Intracellular generation of hydroperoxides by mitochondria appears to be an early event in hydrophobic bile acid-induced hepatocyte toxicity. Antioxidants may be of benefit in cholestasis.
Sokol
R J
RJ
Section of Pediatric Gastroenterology and Nutrition, University of Colorado School of Medicine, Denver, USA.
Winklhofer-Roob
B M
BM
Devereaux
M W
MW
McKim
J M
JM
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D001647:Bile Acids and Salts / Q000494:pharmacology*; D002478:Cells, Cultured; D002635:Chenodeoxycholic Acid / Q000494:pharmacology; D019826:Cholic Acid; D002793:Cholic Acids / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D008297:Male; D008930:Mitochondria, Liver / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013655:Taurochenodeoxycholic Acid / Q000494:pharmacology; D013656:Taurocholic Acid / Q000494:pharmacology; D017392:Thiobarbituric Acid Reactive Substances / Q000378:metabolism
D000975:Antioxidants; D001647:Bile Acids and Salts; D002793:Cholic Acids; D017392:Thiobarbituric Acid Reactive Substances; D002635:Chenodeoxycholic Acid; D013655:Taurochenodeoxycholic Acid; D013656:Taurocholic Acid; D006861:Hydrogen Peroxide; D019826:Cholic Acid