Study On Protective Effect And Mechanism Of Fisetin On Hypoxia/Reoxygenation Injury In Rat Hepatocytes

Objective: To explore the protective effect of fisetin(FIS)on rat hepatocytes from hypoxia /reoxygenation(H/R)injury,and whether the mechanism of fisetin protection is related to TLR4/ NF-κB signaling pathways regulation.Methods: Hypoxia/reoxygenation model of BRL-3A cells was established and cells were pretreated with FIS or TLR4 inhibitor HTA125.Survival rate was detected by CCK-8.Cell apoptosis was detected by double staining with flow cytometry.The levels of alanine transaminase(ALT)and aspartate transaminase(AST)were detected by Microplate method.TNF-α and IL-1β levels were detected by ELISA.The mRNA and protein expression levels of TLR4,NF-κB p65 and IκB-α in the cells were determined by quantitative real-time PCR and Western blotting.The translocation of NF-κB p65 was observation by using immunofluorescence technique.Results: Subjected to H/R,cell survival rate decreased and the apoptosis enhanced.The levels of ALT and AST in cell supernate enhanced,as well as the levels of TNF-α and IL-1β.The expression levels of TLR4 and NF-κB p65 increased,while the expression levels of IκB-α decreased.The translocation of NF-κB p65 increased.FIS or HTA125 pretreatment increased the cell survival rate and decreased the apoptosis.ALT,AST,TNF-α and IL-1β levels reduced significantly,the expression levels of TLR4 and NF-κB p65 decreased and the expression level of IκB-α increased.Moreover,fisetin inhibited the translocation of NF-κB p65.Conclusions: FIS can alleviate the hepatocyte injury induced by H/R,which may be related with its regulation effect on TLR4/NF-κB signaling pathway.