Role of protein kinase C in reduction of hepatic ischemia-reperfusion injury by CO2 preconditioning in rats

Abstract

Objective To investigate the role of protein kinase C (PKC) in reduction of hepatic ischemiareperfusion injury by CO2 preconditioning in rats.Methods Forty-eight male Wistar rats,aged 8-10 weeks,weighing 230-270 g,were randomly divided into 3 groups (n =16 each):hepatic ischemia-reperfusion injury group (group HIRI),CO2 preconditioning group (group P),and c helerythrine (CHE,a specific inhibitor of PKC) group (group CHE).The portal vein,hepatic artery and bile duct of the left lateral and median lobes of the liver were occluded for 1 h,followed by 4 h reperfusion in anesthetized rats.The rats inhaled 50％ O2-50％ N2 for 1 h during mechanical ventilation in group HIRI.In P group,the rats inhaled 50％ O2-45％ N2-5％ CO2 for 1 h during mechanical ventilation and then inhaled 50％ O2-50％ N2 and the hepatic ischemia-reperfusion injury was performed 15 min later.In group CHE,CHE 5 mg/kg was injected intraperitoneally at 10 min before mechanical ventilation,and the other procedures were similar to those previously described in P group.Before mechanical ventilation,immediately before ischemia,and at 0,1,2,3 and 4 h of reperfusion,mean arterial pressure (MAP) was recorded and arterial blood samples were obtained for blood gas analysis.At 4 h of reperfusion,the serum aspartate amino transferase (AST) and alanine amino-transferase (ALT) activities and tumor necrosis factor-α (TNF-α) concentration (by ELISA) were determined and hepatic specimens were obtained for detection of malondialdehyde (MDA) content and superoxide dismutase (SOD) activity (by spectrophotometry),and the expression of activated caspase-3 (by immuno-histochemistry) and PKC (by Western blot) in hepatic tissues.Apoptosis index was calculated by using TUNEL.Results Compared with group HIRI,MAP,PaO2 and PaCO2were significantly increased immediately before ischemia and during reperfusion in group P,MAP and PaCO2 were increased during reperfusion and PaO2 was increased immediately before ischemia and during reperfusion in group CHE,the serum ALT and AST activities,TNF-α concentrations,MDA content and apoptosis index were decreased,and the expression of activated caspase-3 was down-regulated in P and CHE groups,and the SOD activity was increased,and the expression of PKC was up-regulated in group P (P ＜ 0.05 or 0.01),and no significant changes were found in the SOD activity and PKC expression in CHE group (P ＞ 0.05).Compared with group P,MAP was significantly increased immediately after onset of reperfusion,while decreased at 1-4 h of reperfusion,PaO2 was decreased immediately before ischemia and during reperfusion,PaCO2 was decreased at 3 h of reperfusion,the serum ALT and AST activities,TNF-α concentrations,MDA content and apoptosis index were increased,and the expression of activated caspase-3 was up-regulated,and the expression of PKC was downregulated in group CHE (P ＜ 0.05).Conclusion PKC is involved in reduction of hepatic ischemia-reperfusion injury by CO2 preconditioning in rats. Key words: Carbon dioxide; Protein kinase C; Reperfusion injury; Liver