木蝴蝶苷A对肝细胞癌细胞侵袭增殖的抑制作用及机制

Inhibitory effect of oroxin A on invasion and proliferation of hepatocellular carcinoma cells and its mechanism

目的 探讨木蝴蝶苷A(OA)抑制肝细胞癌(HCC)细胞恶性行为的作用机制。方法 购入HCC细胞系Hep3B、MHCC97-L,设置OA梯度浓度(0、30、60、90、120μM组),通过CCK-8检测细胞活力确定OA对HCC细胞的半抑制浓度(IC50),随后采取IC50进行后续实验,将细胞分为对照组(Control组)与OA处理组(OA组),采取试剂盒检测铁死亡相关指标二价铁离子(Fe^(2+))含量、脂质活性氧(Lipid-ROS)含量、还原型谷胱甘肽(GSH)、活性氧脂质过氧化产物丙二醛(MDA)、超氧化物歧化酶(SOD)活性,使用Tri-Carb^■液体闪烁分析仪检测胱氨酸摄取水平。在添加OA的HCC细胞中同时转染pcDNA3.1-TXNDC5(OA+oe-TXNDC5组),以pcDNA3.1-NC为对照(OA+oe-NC组)检测过表达硫氧还蛋白结构域蛋白5(TXNDC5)对OA诱导HCC细胞铁死亡的影响。结果 OA呈一定浓度依赖抑制HCC细胞MHCC97-L与Hep3B活力,对Hep3B半抑制浓度为30μM。OA处理Hep3B细胞使得Fe^(2+)含量显著上升,抑制胱氨酸摄取,Lipid-ROS含量显著增加,升高MDA水平,降低GSH含量、SOD活性,诱导Hep3B细胞铁死亡。过表达TXNDC5可部分逆转OA对细胞铁死亡的诱导作用。结论 OA通过下调TXNDC5诱导HCC细胞铁死亡由此抑制HCC细胞恶性行为。

Objective The aim of this study was to investigate the mechanism of action of oroxin A in inhibiting the malignant behavior of hepatocellular carcinoma cells.Methods The hepatoma cell lines Hep3B and MHCC97-L were purchased.We set gradient concentration of oroxin A(0,30,60,90,120μM),and the semi-inhibitory concentration(IC50)of oroxin A on hepatoma cells was determined by CCK-8 assay of cell viability.IC50 concentration was subsequently taken for subsequent experiments.The cells were grouped as Control and OA.Kits were taken to detect iron death-related indexes Fe^(2+) content,lipid reactive oxygen species(Lipid-ROS)content,glutathione(GSH),malondialdehyde(MDA),superoxide dismutase(SOD)activity,and cystine uptake levels using the Tri-Carb Liquid Scintillation Analyzer.We transfected pcDNA3.1-TXNDC5 into HCC cells induced by oroxin A(OA+oe-TXNDC5),with pcDNA3.1-NC as control(OA+oe-NC).The effect of overexpression of TXNDC5 on ferroptosis in HCC cells induced by oroxin A was examined.Resluts A concentration-dependent inhibition of hepatocellular carcinoma cell MHCC97-L and Hep3B viability was observed.The semi-inhibitory concentration of Hep3B was 30μM.Treatment of Hep3B cells with oroxin A resulted in a significant increase in Fe^(2+)content,inhibition of cystine uptake,significant increase in Lipid-ROS content and MDA level,decrease in GSH content and SOD activity,which mean a induction of ferroptosis in Hep3B cells.Overexpression of TXNDC5 partially reversed the induction of ferroptosis by Oroxin A.Conclusion Oroxin A induced ferroptosis in HCC cells by downregulating TXNDC5,thereby inhibiting the malignant behavior of hepatocellular carcinoma cells.