目的:Ferroptosis是一种新发现的程序性细胞死亡形式;在胃癌中,调节铁凋亡的具体机制尚未完全阐明。在这项研究中,我们旨在研究微粒体谷胱甘肽S-转移酶1(MGST1)如何调节胃癌细胞的铁凋亡。
方法:过表达MGST1或仅表达低水平MGST1的胃腺癌(SGC7901)细胞用特定化合物(erastin,索拉非尼,RSL3、MK-2206和SC79)。然后,我们检测到丙二醛(MDA)的水平,谷胱甘肽(GSH),铁和活性氧(ROS)。通过蛋白质印迹测定非经典自噬和蛋白激酶B(Akt)/糖原合酶激酶-3β(GSK-3β)途径的蛋白质表达水平,并通过细胞计数试剂盒-8(CCK-8)测定分析细胞活力。使用qRT-PCR检测目标基因的表达。
结果:我们评估了一系列诱导铁凋亡的化合物,发现MGST1表达在SGC7901细胞的铁凋亡过程中下调。铁凋亡诱导剂RSL3在经典的铁凋亡事件中起作用,而MGST1的过表达削弱了这些作用。有趣的是,MGST1的过表达通过抑制ATG16L1的表达和LC3-I向LC3-II的转化而导致自噬失活。ATG16L1的上调消除了MGST1对铁凋亡的抑制作用。值得注意的是,MGST1的过表达诱导Akt/GSK-3β途径的激活。Akt抑制剂拮抗MGST1对自噬和铁凋亡的抑制作用。
结论:总的来说,我们的发现证明了铁凋亡的一个新的分子机制和信号通路。我们还鉴定了MGST1的过表达通过激活Akt/GSK-3β信号通路诱导胃癌细胞增殖。
OBJECTIVE: Ferroptosis is a newly discovered form of programmed cell death; however, the specific mechanisms that regulate ferroptosis have yet to be fully elucidated in gastric carcinoma. In this study, we aimed to investigate how microsomal glutathione S-transferase 1 (MGST1) regulates ferroptosis in gastric carcinoma cells.
METHODS: Gastric adenocarcinoma (SGC7901) cells that overexpressed MGST1 or expressed only low levels of MGST1, were treated with specific compounds (erastin, sorafenib, RSL3, MK-2206 and SC79). Then, we detected the levels of malondialdehyde (MDA), glutathione (GSH), iron and reactive oxygen species (ROS). Protein expression levels of the non-classical autophagy and protein kinase B (Akt)/glycogen synthase kinase-3β (GSK-3β) pathways were determined by western blotting and cell viability was analyzed by Cell Counting Kit-8 (CCK-8) assays. The expressions of target genes were detected using qRT-PCR.
RESULTS: We evaluated a range of ferroptosis-inducing compounds and found that MGST1 expression was down-regulated during ferroptosis in SGC7901 cells. The ferroptosis inducer RSL3 played a role in classical ferroptotic events while the overexpression of MGST1 impaired these effects. Interestingly, the overexpression of MGST1 resulted in the inactivation of autophagy by repressing the expression of ATG16L1 and the conversion of LC3-I to LC3-II. The upregulation of ATG16L1 eliminated the inhibitory action of MGST1 on ferroptosis. Notably, the overexpression of MGST1 induced the activation of the Akt/GSK-3β pathway. An Akt inhibitor antagonized the inhibitory effects of MGST1 on autophagy and ferroptosis.
CONCLUSIONS: Collectively, our findings demonstrate a novel molecular mechanism and signaling pathway for ferroptosis. We also characterized that the overexpression of MGST1 induces gastric carcinoma cell proliferation by activating the Akt/GSK-3β signaling pathway.