附子通过AMPK-自噬-铁死亡途径抑制Hela细胞活性OA

Objective To investigate the effect of Aconiti Lateralis Radix Praeparata on the viability of Hela cells and to study the mechanism by which Aconiti Lateralis Radix Praeparata induces ferroptosis in Hela cells.Methods A water decoction of Aconiti Lateralis Radix Praeparata was administered to mice by gavage to prepare drug-containing serum.This Aconiti Lateralis Radix Praeparata-containing serum was used to replace fetal bovine serum for treating human embryonic kidney epithelial cells(HEK 293T)and human cervical cancer cells(Hela).Cell viability was determined by colorimetric assay.Concentrations of malondialdehyde(MDA)and glutathione(GSH)in the cells were measured by enzyme-linked immunosorbent assay(ELISA).Protein levels of glutathione peroxidase 4(GPX4),AMP-activated protein kinase(AMPK),and phosphorylated AMPK(p-AMPK,specifically at Thr172)were determined by Western Blot.Hela cells were pretreated with the AMPK inhibitor BML275 and the autophagy inhibitor 3-methyladenine(3MA)to further explore the roles of AMPK and autophagy in Aconiti Lateralis Radix Praeparata-induced ferroptosis.Lipid peroxidation levels were measured using the BODIPY 581/591 C11 fluorescent probe.Results Compared with the control group,Aconiti Lateralis Radix Praeparata-containing serum did not affect the viability of HEK 293T cells but significantly reduced the viability of Hela cells(P＜0.05,P＜0.000 1).In Hela cells of the Aconiti Lateralis Radix Praeparata group,the level of the ferroptosis marker MDA increased(P＜0.001),GSH content decreased(P＜0.001),GPX4 level was significantly reduced(P＜0.001),and the protein contents of AMPK and p-AMPK were significantly elevated(P＜0.01,P＜0.05).Pretreatment with either BML-275 or 3MA significantly increased the GPX4 content in the Aconiti Lateralis Radix Praeparata-treated cells(P＜0.001,P＜0.01).Lipid peroxidation was significantly increased in the Aconiti Lateralis Radix Praeparata group(P＜0.000 1),and pretreatment with BML-275 or 3-MA significantly reduced this lipid peroxidation level(P＜0.000 1).Conclusion Aconiti Lateralis Radix Praeparata may reduce Hela cell viability by activating AMPK and inducing autophagy to promote ferroptosis,playing a significant role in the ferroptosis process of Hela cells.