Cyclic adenosine monophosphate-mediated protection against bile acid-induced apoptosis in cultured rat hepatocytes.
Cyclic adenosine monophosphate (cAMP) has been shown to modulate apoptosis. To evaluate the role of cAMP in bile acid-induced hepatocyte apoptosis, we studied the effect of agents that increase cAMP on the induction of apoptosis by glycochenodeoxycholate (GCDC) in cultured rat hepatocytes. GCDC induced apoptosis in 26.5%+/-1.1% of hepatocytes within 2 hours. Twenty-minute pretreatment of hepatocytes with 100 micromol/L 8-(4-chlorothiophenyl) cAMP (CP-cAMP) resulted in a reduction in the amount of apoptosis to 35.2%+/-3.8% of that seen in hepatocytes treated with GCDC alone. Other agents that increase intracellular cAMP, including dibutyryl cAMP (100 micromol/L), glucagon (200 nmol/L), and a combination of forskolin (20 micromol/L) and 3-isobutyl-1-methylxanthine (20 micromol/L), also inhibited GCDC-induced apoptosis to a similar extent. Pretreatment with the protein kinase A (PKA) inhibitor, KT5720, prevented the protective effect of CP-cAMP and inhibited CP-cAMP-induced activation of PKA activity. Inhibitors of phosphatidylinositol 3-kinase (PI3K), wortmannin (50 nmol/L), or Ly 294002 (20 micromol/L) also prevented the cytoprotective effect of cAMP. PI3K assays confirmed that wortmannin (50 nmol/L) inhibited PI3K activity, while CP-cAMP had no effect on the activity of this lipid kinase. GCDC increased mitogen-activated protein kinase (MAPK) activity, but had no effect on stress-activated protein kinase (SAPK) activity in hepatocytes. cAMP decreased basal and GCDC-induced MAPK activity and increased SAPK activity. The MAPK kinase inhibitor, PD 98059, inhibited both GCDC-mediated MAPK activation and GCDC-induced apoptosis.


CONCLUSIONS
1) agents that increase intracellular cAMP protect against hepatocyte apoptosis induced by hydrophobic bile acids; 2) activation of MAPK by GCDC may be involved in bile acid-induced apoptosis; and 3) cAMP-mediated cytoprotection against bile acid-induced apoptosis appears to involve PKA, MAPK, and PI3K.
Webster
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Tufts University School of Veterinary Medicine, North Grafton, MA, USA.
Anwer
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D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001647:Bile Acids and Salts / Q000737:chemistry / Q000494:pharmacology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000502:physiology; D002227:Carbazoles; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D000242:Cyclic AMP / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D048031:JNK Mitogen-Activated Protein Kinases; D008099:Liver / Q000166:cytology*; D008297:Male; D020928:Mitogen-Activated Protein Kinases; D009025:Morpholines / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011758:Pyrroles / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction; D012995:Solubility; D000077191:Wortmannin
D000730:Androstadienes; D001647:Bile Acids and Salts; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002227:Carbazoles; D002867:Chromones; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D004791:Enzyme Inhibitors; D007211:Indoles; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011758:Pyrroles; D000077191:Wortmannin; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C057416:KT 5720