Induction Of Nrf2 Pathway By Dendrobium Nobile Lindl. Alkaloids Protects Against Carbon Tetrachloride Induced Acute Liver Injury

Objective:To observe the protective effect and underlying of Dendrobium nobile Lindl.alkaloids?DNLA?on carbon tetrachloride?CCl₄?induced acute liver injury in mice.Methods:?1?Sixty C57BL/6J mice were randomly divided into normal control group,DNLA administration alone group?20 mg/kg?,CCl₄ model group,DNLA low dose group?10 mg/kg?,DNLA high dose group?20 mg/kg?and bifendate group?150 mg/kg?,10 mice in each group.The mice were given DNLA or bifendate for 7 days,and the normal control group and the CCl₄ model group were given equal volume of distilled water.After 2 hours of the last administration,acute liver injury was induced by intraperitoneal injection of0.2%CCl₄ solution?0.1 mL/10 g?.The mice were sacrificed 24 hours posttreatment and liver and serum samples were collected.The levels of serum ALT and AST,the contents of malondialdehyde?MDA?and activities of superoxide dismutase?SOD?and glutathione?GSH?in mouse liver were determined.The hepatic histopathological changes were observed by hematoxylin-eosin staining?HE?.The ultrastructural changes of liver were observed by transmission electron microscopy.The expression levels of Nrf2 nuclear protein and its downstream target proteins GCLC,NQO1 and HO-1 were detected by Western blotting.?2?Wild-type?WT?and Nrf2 knockout(Nrf2^-/-)mice were randomly divided into normal control group,DNLA normal administration group?20 mg/kg?,CCl₄ model group and DNLA-administered group?20 mg/kg?,6 mice in each group.The methods of administration,disposal and detection are the same as above.Results:?1?Compared with the normal control group,the serum levels of ALT and AST in the CCl₄ model group increased significantly.MDA content in liver tissue increased,while SOD and GSH activity decreased significantly.Pathological examination showed hepatic cell degeneration,necrosis and inflammatory cell infiltration.While hepatocyte mitochondria showed obvious swelling,hepatocyte nuclear membrane shrinkage,incompleteness,heterochromatin increased significantly,concentrated edge set.And the expression level of Nrf2 in the nucleus of hepatocytes decreased,while the expression levels of total proteins NQO1,HO-1 and GCLC increased.Compared with the CCl₄ model group,the serum levels of ALT and AST in the DNLA-administered group were significantly decreased,and the activities of SOD and GSH in the liver tissue were significantly increased,while the MDA content was significantly decreased.The pathological examination showed that the pathological damage of the liver was significantly reduced.The damage of liver mitochondria was significantly improved.The results of Western bolt showed that the expression of Nrf2,GCLC,NQO1 and HO-1 in DNLA low and high dose groups was significantly increased.?2?Compared with the normal control group,the serum levels of ALT and AST in the Nrf2^-/-mice CCl₄ model group were significantly higher than those in the WT mice.Histopathological examination of liver also showed that the degree of liver injury in Nrf2^-/-mice was more severe than that in WT mice.Compared with the model group,the serum levels of ALT and AST in the WT DNLA-administered group were significantly decreased,but those in the Nrf2^-/-DNLA-administered group were not inhibited.For histopathological examination,DNLA could reduce the pathological damage of the liver in the WT mice,but not in the Nrf2^-/-mice.Western blot analysis showed that the expression levels of GCLC,NQO1,GST and GPX1 in the liver of WT group were significantly increased after DLNA administration,but no significant difference was found in Nrf2^-/-mice.Conclusion:DNLA can significantly improve CCl₄-induced acute liver injury in mice,which may be related to the increase of the expression of oxidative stress-related proteins Nrf2,GCLC,NQO1,HO-1.