Nitric oxide ameliorates hydrophobic bile acid-induced apoptosis in isolated rat hepatocytes by non-mitochondrial pathways.
Hydrophobic bile acids are toxic to isolated rat hepatocytes by mechanisms involving mitochondrial dysfunction and oxidative stress. In the current study we examined the role of nitric oxide (NO), a potential mediator of apoptosis, during bile acid-induced apoptosis. Freshly isolated rat hepatocytes and hepatic mitochondria generated NO and peroxynitrite (ONOO(-)) in a concentration- and time-dependent manner when exposed to the toxic bile salt glycochenodeoxycholate (GCDC) (25-500 microm), which was prevented by the nitric-oxide synthase (NOS) inhibitors N(G)-monomethyl-N-arginine monoacetate (l-NMMA) and 1400W. Relationships between hepatocyte NO production and apoptosis were examined by comparing the effects of NOS inhibitors with other inhibitors of GCDC-induced apoptosis. Inhibitors of caspases 8 and 9, the mitochondrial permeability transition blocker cyclosporin A, and the antioxidant idebenone reduced NO generation and apoptosis in GCDC-treated hepatocytes. In contrast, NOS inhibitors had no effect on GCDC-induced apoptosis despite marked reduction of NO and ONOO(-). However, treatment with the NO donors S-nitroso-N-acetylpenicillamine and spermine NONOate [N-(-aminoethyl)N-(2-hydroxy-2-nitrohydrazino)-1,2-ethylenediamine) inhibited apoptosis and caspase 3 activity while significantly elevating NO levels above GCDC-stimulated levels. Neither NO donors nor NOS inhibitors affected GCDC-induced mitochondrial permeability transition or cytochrome c release from liver mitochondria or GCDC-induced mitochondrial depolarization from isolated hepatocytes, suggesting that NO inhibits bile acid-induced hepatocyte apoptosis by a non-mitochondrial-dependent pathway. In conclusion, whereas NO produced from GCDC-treated hepatocytes neither mediates nor protects against bile acid-induced apoptosis, higher levels of NO inhibit GCDC-induced hepatocyte apoptosis by caspase-dependent pathways.
Gumpricht
Eric
E
Pediatric Liver Center and Liver Transplantation Program, Section of Pediatric Gastroenterology, Hepatology, and Nutrition, Department of Pediatrics, University of Colorado School of Medicine, Denver 80262, USA.
Dahl
Rolf
R
Yerushalmi
Baruch
B
Devereaux
Michael W
MW
Sokol
Ronald J
RJ
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D005999:Glycochenodeoxycholic Acid / Q000494:pharmacology*; D022781:Hepatocytes / Q000187:drug effects*; D008297:Male; D008930:Mitochondria, Liver / Q000378:metabolism*; D009569:Nitric Oxide / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D051381:Rats; D017207:Rats, Sprague-Dawley
D004791:Enzyme Inhibitors; D009569:Nitric Oxide; D005999:Glycochenodeoxycholic Acid; D019001:Nitric Oxide Synthase