Objective:To explore the mechanism of tumulosic acid inhibiting oxygen-glucose deprivation/reoxygenation(OGD/R)-induced cardiomyocyte ferroptosis.Methods:Rat cardiomyocytes H9c2 were cultured in vitro and treated with 0,1.0,2.5,5.0,10.0,20.0 μmol/L tumulosic acid for 24 h immediately after the establishment of the cell OGD/R model.The cell viability of each treatment group was detected by CCK-8 method,and the appropriate concentration of tumulosic acid was screened out.H9C2 cells were randomly divided into control group,model group,tumulosic acid group,ML385 group[nuclear factor erythroid-2-related factor 2(Nrf2)inhibitor group],and tumulosic acid+ML385 group,except for the control group,the rest of the groups were treated with tumulosic acid and ML385 immediately after the establishment of the cell OGD/R model.The viability and apoptosis rate of H9c2 cells in each group were detected by CCK-8 method and flow cytometry respectively.The kits were applied to detect the levels of lactate dehydrogenase(LDH),pro-inflammatory factors[prostaglandin E2(PGE2),and tumor necrosis factor-α(TNF-α)],anti-inflammatory factor interleukin(IL)-10 levels,cellular antioxidant factors[catalase(CAT)and glutathione(GSH)],ferroptosis-related indicators[iron content and malondialdehyde(MDA)]in the supernatant of H9c2 cell culture medium in each group.The apoptosis of H9c2 cells in each group and the expression of Nrf2/solute carrier family 7 member 11(SLC7A11)/glutathione peroxidase 4(GPX4)signal-related proteins were detected by Western Blotting.Results:Compared with control group,the apoptosis rate,LDH release,PGE2 and TNF-α levels in cell culture supernatant,cellular iron content,MDA level,and Bax and Caspase-3 protein expression increased in model group(P<0.05),the cell viability,IL-10 level in cell culture supernatant,cellular CAT and GSH levels,Bcl-2 and Nrf2,SLC7A11,GPX4 protein expression decreased(P<0.05).Compared with model group,the apoptosis rate,LDH release(PGE2)and TNF-αlevels in cell culture supernatant,cellular iron content,MDA level,and Bax and Caspase-3 protein expression decreased in tumulosic acid group(P<0.05),the cell viability,IL-10 level in cell culture supernatant,cellular CAT and GSH levels,Bcl-2 and Nrf2,SLC7A11,GPX4 protein expression increased(P<0.05);the apoptosis rate,LDH release,PGE2,and TNF-αlevels in cell culture supernatant,cellular iron content,MDA level,and Bax and Caspase-3 protein expression increased in ML385 group(P<0.05),the cell viability,IL-10 level in cell culture supernatant,cellular CAT and GSH levels,Bcl-2 and Nrf2,SLC7A11,GPX4 protein expression decreased(P<0.05).Compared with tumulosic acid group,the apoptosis rate,LDH release,PGE2,and TNF-αlevels in cell culture supernatant,cellular iron content,MDA level,and Bax and Caspase-3 protein expression increased in tumulosic acid+ ML385 group(P<0.05),the cell viability,IL-10 level in cell culture supernatant,cellular CAT and GSH levels,Bcl-2 and Nrf2,SLC7A11,GPX4 protein expression decreased(P<0.05).Conclusion:Tumulosic acid can inhibit OGD/R-induced myocardial cell inflammation,lipid peroxidation,and ferroptosis by activating Nrf2/SLC7A11/GPX4 signaling,enhance antioxidant activity and cell viability,and ultimately alleviate apoptosis injury.
oxygen glucose deprivation/reoxygenationcardiomyocytestumulosic acidnuclear factor erythroid-2-related factor 2/solute carrier family 7 member 11/glutathione peroxidase 4ferroptosisexperimental study
NETL
NSTL
万方数据
维普
