The Study On The Mechanism Of Bianliangziyu Ethanol Extract Alleviating Acetaminophen-induced Hepatotoxicity

Objective: In this study,we investigated the hepatoprotective and antioxidative effects of “Bianliang ziyu” extract(BZE)on liver injury and explored its molecular mechanism.Method: Sprague-Dawley rats were administered BZE by intragastric administration for 10 d,and then acetaminophen(APAP)was administered by gavage to induce acute liver injury.The activities of serum Superoxide Dismutase(SOD),Glutathione(GSH),alanine aminotransferase(ALT),aspartate aminotransferase(AST)and superoxide dismutase(ROS)in the rats were measured,the organ index of rats was calculated and the liver of each rat was examined for histopathological changes.In vitro,HL-7702 cells were pretreated with BZE for 24 h and then exposed to 30 mmol/L APAP for 12 h.The survival rate of the cells and ALT,AST,ROS,SOD,GSH activities were determined.Then,we investigated the effects of BZE on oxidative stress,apoptosis and the activation of nuclear factor erythroid-2-related factor 2(Nrf2)signaling in HL-7702 cells induced by APAP.First,we examined the effect of BZE on mitochondrial membrane potential using flow cytometry,and then detected mitochondrial apoptosis-related proteins Bax,Bcl-2 and Caspase-3 expression and oxidative stress signaling pathway Nrf2,HO-1 protein expression using Western Blotting.Finally,the expression of Nrf2 was down-regulated by siRNA,and the expression of mitochondrial apoptosis-related proteins HO-1,Bcl-2,Bax and Caspase-3 were detected by Western blotting.Results: The results showed that BZE prevented APAP-induced liver injury and suppressed hepatic oxidative stress in vitro and in vivo.In vivo,BZE improves the morphology of tissue cells in HE staining,reduces liver damage markers ALT and AST,reduces the amount of ROS produced by oxidative stress and increases the expression of two-phase detoxification enzymes GSH and SOD.In vitro,BZE can significantly improve the survival rate of HL-7702 cells,improve the morphology of APAP-induced damage,significantly reduce ALT,AST,reduce ROS and increase the expression of antioxidant enzymes GSH and SOD.BZE was also observed to significantly inhibit the reduction of mitochondrial membrane potential and regulate the expression of Bcl-2,Bax and Caspase-3 in APAP-induced HL-7702 cells.In addition,BZE significantly promoted nuclear translocation and the expression of Nrf2 as well as its downstream gene hemeoxygenase-1(HO-1)in vitro.Furthermore,our findings showed that siNrf2 reversed the effects of BZE on cell survival and apoptosis-related protein expression in APAP-induced HL-7702 cells.Conclusions: BZE inhibits liver oxidative stress and prevents APAP-induced hepatotoxicity in vitro and in vivo.BZE plays an important role in preventing hepatotoxicity by inhibiting oxidative stress and apoptosis through activation of Nrf2 signaling.