DADLE’s Improvement Of Hepatic Ischemia/Reperfusion Injury In Mice By Activating The Nrf2/HO-1 Pathway

Objective:Hepatic Ischemia/Reperfusion(I/R)injury is a common pathophysiological process in liver surgery,which can cause acute liver injury and may even lead to liver failure if serious.Many studies have shown that DADLE,as a specific delta opioid receptor(DOR)agonist,has a good protective effect in I/R injury of the brain,heart and kidney.The aim of this study was to investigate the protective effect of DADLE on hepatic ischemia-reperfusion injury in mice and its mechanism,and provide a new strategy for clinical prevention of hepatic ischemia-reperfusion injury.Methods :Forty-eight mice underwent 70% of liver ischemia for 45 min,Six mice were sacrificed at each reperfusion time points(0 h,1 h,2 h,4 h,6 h,8 h,12 h,24 h),The changes of serum Alanine aminotransferase(ALT)and aspartate aminotransferase(AST)were measured at different time points after reperfusion in mice.Thirty mice were intraperitoneally injected with different concentrations of DADLE(1 mg / kg,3 mg / kg,5 mg / kg,7 mg / kg,10 mg / kg)at 15 min before ischemia,Only 45 min after ischemia,mice were sacrificed at the same time point according to the previous experiment.The effects of different concentrations of DADLE on serum ALT and AST were observed.Twenty-four C57BL/6 mice were randomly divided into four groups: normal group(Sham group),ischemia/ reperfusion group(I/R group),DADLE agonist group(DADLE group)and naloxone antagonist group(N/D Group).The serum levels of ALT and AST were measured with an automated chemical analyzer.HE staining was used to observe the morphological changes of liver tissue.The levels of SOD,MDA,GSH and CAT in liver homogenates were measured by enzymatic activity and spectrophotometry.The expression of HO-1 mRNA was detected by RT-PCR.WB was used to detect the expression of Nrf2 and its induced protein HO-1.Results:(1)we detected the levels of serum ALT and AST at different time(0 h,1 h,2 h,4 h,6 h,8 h,12 h and 24 h)respectively.The results showed that ALT and AST levels reached the highest at 6 hours after I/R.(2)we detected the levels of serum ALT and AST in different concentrations of DADLE(1 mg/kg,3 mg/kg,5mg/kg,7 mg/kg and 10 mg/kg).The results showed that the ALT and AST levels were significantly lower when DADLE at 5 mg/kg.DADLE can reduce serum ALT and AST levels in ischemia-reperfusion injury group and can improve hepatic sinus blockage,hepatocyte swelling,degeneration,necrosis and inflammatory cell infiltration.The results showed that DADLE could increase the levels of T-SOD,GSH and CAT in liver homogenate,and decrease the level of MDA in liver tissue homogenate.(5)RT-PCR was used to detected the expression of HO-1 mRNA in the liver.The results showed that DADLE could increase the expression of HO-1 mRNA in the liver.(6)Western blotting showed that DADLE could promoted the nuclear transfer of Nrf2 and increase the expression of downstream protein HO-1.Conclusions:DADLE has a good protective effect in liver I/R injury.Part of the mechanism may be that DADLE activates Nrf2 for nuclear translocation to up-regulate the expression of HO-1,the downstream anti-oxidation factor,which may play an anti-oxidative role in hepatic I/R injury in mice.