Protective effect of nitric oxide on isolated rat hepatocytes submitted to an oxidative stress

Abstract

We have previously suggested that the nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathway protects both hepatocytes and endothelial cells against liver ischemia-reperfusion injury in rat. We study here the ability of NO to protect isolated hepatocytes against an in vitro oxidative stress induced with hypochlorite solution (ClO [minus ]). The severity of ClO [minus ]-induced stress was quantified by the measurement of total glutathione and membrane lipid peroxidation. Cell damage was assessed by morphologic (cell viability and bleb formation) and biologic (transaminase release) criteria. A 30-minute incubation of hepatocytes with 100 [mu ]mol/L ClO [minus ] maximally decreased cell viability ([minus ]40%) and increased bleb formation (+300%) and release of transaminases activities (aspartate transaminase [AST] = +60% and alanine transaminase [ALT] = +300%). A good correlation was observed between morphologic and biologic criteria. A preincubation of cells with 50 [mu ]mol/L 8-Br-cGMP, did not affect the adverse ClO [minus ] effects on the morphologic criteria. In the presence of 20 [mu ]mol/L spermineNONOate, an NO donor, ClO [minus ] did not decrease cell viability, whereas its deleterious effects on bleb formation was unchanged. A preincubation with a specific inhibitor of the soluble guanylate cyclase, the 1 H-[1,2,4]oxadiazolo[4,3- a]quinoxalin-1-one (ODQ; 1 [mu ]mol/L), did not affect the beneficial effect of NO on the cell viability. Our results suggest that NO protects hepatocytes against oxidative stress by a mechanism, which is cGMP-independent. However, taking into account the cytoprotective effects of cGMP in the liver, it is likely that the rapid effect of NO observed in vitro is relayed in vivo by a more long-lasting mechanism, which would be inhibited by ODQ and mimicked by 8-Br-cGMP.