| Background:Ursolic Acid(UA)is a pentacyclic triterpenoid compound existing in natural plants,which has a variety of pharmacological effects,especially in anti-tumor,anti-cardiovascular disease and liver protection.Studies have shown that ursolic acid has a protective effect on cholestatic liver injury,but the specific mechanism of its liver protective effect has not been reported.Bile acid in vivo is mainly transferred and excreted by biliary duct BSEP and MRP2 after UGT2B7 metabolism in the liver,while the expression of UGT2B7,BSEP,MRP2 is regulated by Nrf2,and the literature has shown that Ursa acid(UA)has a clear activation effect on the nuclear receptor Nrf2.Therefore,ursolic acid is likely to play the role of liver protection by activating the nuclear receptor Nrf2 and upregulating the expression of UGT2B7,BSEP,MRP2 to promote bile acid effluviation,thus reducing liver cell damage.Previous studies by the research group also showed that UA significantly up-regulated the mRNA and protein levels of nuclear receptor Nrf2,metabolic enzyme UGT2B7,transporters BSEP and MRP2 in HepG2 cells,and the expression of MRP2 was regulated by Nrf2.Therefore,it is of practical significance to further explore the mechanism of ursolic acid in liver protection in rats.Objectives:This experiment adopted alpha naphthyl isothiocyanates(ANIT)on the liver injury induced by male SD rats as models,explore the ursolic acid Nrf2 nuclear receptors in liver injury in rats,and its metabolic enzymes UGT2B7,transporter BSEP and MRP2 pathways.It provides a theoretical basis for the clinical application of ursolic acid in liver protection.Methods:1.Thirty male SD rats were randomly divided into five groups: Control group;ANIT(60mg/kg)model group;UA(10mg/kg)low dose group;UA(20mg/kg)medium dose group;UA(40mg/kg)high-dose group.After 14 days of intragastric administration,anesthesia and dissection:1)Blood was collected from the inferior vena cava,serum was collected by centrifugation,and serum indicators were detected by automatic biochemical analyzer: alanine aminotransferase(ALT),aspartate aminotransferase(AST)and alkaline phosphatase(ALP)and ?-glutamine transferase(?-GT),total bilirubin(T-BIL),direct bilirubin(D-BIL)and total bile acid(TBA)concentration;2)Take the liver for HE staining and analyze the liver pathological status of different groups.2.Twenty-one male SD rats were randomly divided into three groups: Control group;ANIT(60mg/kg)model group;UA(40mg/kg)group.After 14 days of intragastric administration,the liver was dissected to extract RNA and protein from the liver tissue.By Rt-QPCR,Western blotting,the effects of UA on mRNA and protein expression of nuclear receptor Nrf2,metabolic enzyme UGT2B7,transporter BSEP and MRP2 were investigated.3.Twenty-four male SD rats were randomly divided into three groups: Control group;ANIT(60mg/kg)model group;ANIT+UA(40mg/kg)administration group.Each large group is further divided into two groups: the shRNA-Nrf2 group and the shRNA-Blank group.After lentivirus infection 21 days,anesthetize and dissect the liver:1)Intravenous injection of lentivirus;2)Extract RNA and protein from liver tissue,by Rt-QPCR,Western blotting to further explore the expression of mRNA and protein expression of metabolic enzymes UGT2B7 and transporters BSEP,MRP2 by UA in Nrf2-silent liver injury rats Influence and its association with the nuclear receptor Nrf2.Results:1.Compared with the control group,serum levels of ALT,AST,ALP,?-GT,T-BIL,D-BIL and TBA were significantly increased in the ANIT model group.Compared with the model group,ursolic acid reduced serum ALT,AST,ALP,?-GT,T-BIL,D-BIL and TBA in a dose-dependent manner.The histological evaluation also clearly showed that UA preadministration could significantly reduce the liver pathological damage caused by ANIT.2.Compared with the control group,the mRNA and protein levels of nuclear receptor Nrf2,metabolic enzyme UGT2B7,transporter BSEP and MRP2 were significantly down-regulated in the ANIT model group.After UA preadministration,mRNA and protein levels of nuclear receptor Nrf2,metabolic enzyme UGT2B7,transporter BSEP and MRP2 in ANIT induced liver injury rats were significantly up-regulated.3.In the liver injury rat model with silent Nrf2,compared with the shRNA-Blank group injected with the GFP control lentivirus,the up-regulated effect of UA on nuclear receptor Nrf2,metabolic enzyme UGT2B7,transporter BSEP,MRP2 mRNA and protein levels in the liver injury rats was significantly decreased.Conclutions:1.Ursolic acid has a protective effect on anit-induced liver injury.2.Ursolic acid significantly upregulated the expressions of nuclear receptor Nrf2,metabolic enzyme UGT2B7,transporter BSEP and MRP2 in rats with liver injury.3.The hepatoprotective effect of ursolic acid may be achieved by activating the nuclear receptor Nrf2,thereby up-regulating downstream target genes UGT2B7,BSEP,and MRP2. |
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