title: Methyl methanesulfonate induces necroptosis in human lung adenoma A549 cells through the PIG-3-reactive oxygen species pathway.
abstract: Methyl methanesulfonate (MMS) is an alkylating agent that can induce cell death through apoptosis and necroptosis. The molecular mechanisms underlying MMS-induced apoptosis have been studied extensively; however, little is known about the mechanism for MMS-induced necroptosis. Therefore, we first established MMS-induced necroptosis model using human lung carcinoma A549 cells. It was found that, within a 24-h period, although MMS at concentrations of 50, 100, 200, 400, and 800 μM can induce DNA damage, only at higher concentrations (400 and 800 μM) MMS treatment lead to necroptosis in A549 cells, as it could be inhibited by the specific necroptotic inhibitor necrostatin-1, but not the specific apoptotic inhibitor carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone (Z-VAD-fmk). MMS-induced necroptosis was further confirmed by the induction of the necroptosis biomarkers including the depletion of cellular NADH and ATP and leakage of LDH. This necroptotic cell death was also concurrent with the increased expression of p53, p53-induced gene 3 (PIG-3), high mobility group box-1 protein (HMGB1), and receptor interaction protein kinase (RIP) but not the apoptosis-associated caspase-3 and caspase-9 proteins. Elevated reactive oxygen species (ROS) level was also involved in this process as the specific ROS inhibitor (4-amino-2,4-pyrrolidine-dicarboxylic acid (APDC)) can inhibit the necroptotic cell death. Interestingly, knockdown of PIG-3 expression by small interfering RNA (siRNA) treatment can inhibit the generation of ROS. Taken together, these results suggest that MMS can induce necroptosis in A549 cells, probably through the PIG-3-ROS pathway.
authors.affiliation: Suzhou Biological Technology Co. Ltd. of Centre Testing International Corporation, Kunshan, Jiangsu, 215300, China.|College of Pharmaceutical Sciences, Zhejiang University, Hangzhou, Zhejiang, 310003, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Jiang
authors.affiliation: College of Pharmaceutical Sciences, Zhejiang University, Hangzhou, Zhejiang, 310003, China.
authors.forename: Shigang
authors.initials: S
authors.lastname: Shan
authors.affiliation: Department of Toxicology, Zhejiang University School of Public Health, Hangzhou, Zhejiang, 310058, China.
authors.forename: Linfeng
authors.initials: L
authors.lastname: Chi
authors.affiliation: Department of Toxicology, Zhejiang University School of Public Health, Hangzhou, Zhejiang, 310058, China.
authors.forename: Guanglin
authors.initials: G
authors.lastname: Zhang
authors.affiliation: Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, Zhejiang, 310058, China.
authors.forename: Xiangjing
authors.initials: X
authors.lastname: Gao
authors.affiliation: Department of Toxicology, Hangzhou Normal University School of Medicine, Hangzhou, Zhejiang, 310016, China.
authors.forename: Hongjuan
authors.initials: H
authors.lastname: Li
authors.affiliation: Department of Toxicology, Zhejiang University School of Public Health, Hangzhou, Zhejiang, 310058, China. zhuxq@zju.edu.cn.
authors.forename: Xinqiang
authors.initials: X
authors.lastname: Zhu
authors.affiliation: Department of Toxicology, Hangzhou Normal University School of Medicine, Hangzhou, Zhejiang, 310016, China. junyanghzzju@163.com.|Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, The First Affiliated Hospital, Zhejiang University, Hangzhou, Zhejiang, 310003, China. junyanghzzju@163.com.
authors.forename: Jun
authors.initials: J
authors.lastname: Yang
mesh_terms: D000072283:A549 Cells; D000236:Adenoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D004249:DNA Damage; D004305:Dose-Response Relationship, Drug; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D008741:Methyl Methanesulfonate / Q000494:pharmacology*; D009336:Necrosis; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects* / Q000235:genetics
chemical_list: D018906:Antineoplastic Agents, Alkylating; D047908:Intracellular Signaling Peptides and Proteins; D011518:Proto-Oncogene Proteins; D017382:Reactive Oxygen Species; C411296:TP53I3 protein, human; D008741:Methyl Methanesulfonate
keywords: Apoptosis; Methyl methanesulfonate; Necroptosis; PIG-3; ROS