Li Yi, Zhang Lijun, Zhang Yuke, Zhang Qi, Zhang Lijun
Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Tianjin Medical University, Tianjin 300070, China.
Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Tianjin Medical University, Tianjin 300070, China. *Corresponding author, E-mail:
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2025 Aug;41(8):687-694.
Objective To explore the role and mechanism of mammalian target of rapamycin (mTOR) in oxidized low-density lipoprotein (oxLDL)-induced ferroptosis in vascular smooth muscle cells (VSMCs). Methods A model of oxLDL-induced VSMC ferroptosis was established. VSMCs were co-treated with either the mTOR inhibitor rapamycin or the autophagy inducer carbonyl cyanide m-chlorophenylhydrazone (CCCP), followed by detection of autophagy and ferroptosis-related indexes. Quantitative real-time PCR and Western blot were used respectively to analyze the expression of mTOR, glutathione peroxidase 4 (GPX4), sequestosome 1 (p62), and microtubule-associated protein 1 light chain 3 (LC3). Flow cytometry was employed to assess VSMC death. C11 BODIPY fluorescent staining was used to measure cellular lipid peroxidation levels. Colorimetric assays were performed to determine the contents of malondialdehyde (MDA), ferrous ion (Fe) and glutathione (GSH). Results oxLDL significantly upregulated mTOR expression in VSMCs, while increasing p62 expression and reducing LC3 expression, thereby suppressing VSMC autophagy. Compared with oxLDL treatment alone, rapamycin co-treatment reversed oxLDL-induced VSMC ferroptosis, as characterized by reduced VSMC death, increased GPX4 expression and GSH contents, along with decreased MDA content, Fe content and lipid peroxidation levels. Similarly, CCCP co-treatment activated autophagy characterized by reduced p62 expression and elevated LC3 expression, which subsequently alleviated oxLDL-induced ferroptosis, showing reduced VSMC death, increased GPX4 expressions and GSH contents, and decreased MDA content, Fe content and lipid peroxidation levels. Moreover, mTOR inhibition by rapamycin significantly reversed the oxLDL-induced upregulation of p62 and downregulation of LC3. Conclusion mTOR may promote oxLDL-induced VSMC ferroptosis by suppressing autophagy.
目的 探讨雷帕霉素靶蛋白（mTOR）在氧化型低密度脂蛋白（oxLDL）诱导的血管平滑肌细胞（VSMC）铁死亡中的作用及机制。方法 建立oxLDL诱导的VSMC铁死亡模型。VSMC分别与mTOR抑制剂雷帕霉素或自噬诱导剂羰基氰m-氯苯腙（CCCP）共同处理，随后检测自噬和铁死亡相关指标。分别采用定量实时PCR和蛋白质印迹法分析mTOR、谷胱甘肽过氧化物酶4（GPX4）、聚集体蛋白1（p62）和微管相关蛋白1轻链3（LC3）的表达。采用流式细胞术评估VSMC死亡情况。采用C11 BODIPY荧光染色法检测细胞脂质过氧化水平。采用比色法测定丙二醛（MDA）、亚铁离子（Fe）和谷胱甘肽（GSH）的含量。结果 oxLDL显著上调VSMC中mTOR的表达，同时增加p62表达并降低LC3表达，从而抑制VSMC自噬。与单独oxLDL处理相比，雷帕霉素共同处理可逆转oxLDL诱导的VSMC铁死亡，表现为VSMC死亡减少、GPX4表达和GSH含量增加，同时MDA含量、Fe含量和脂质过氧化水平降低。同样，CCCP共同处理激活了自噬，表现为p62表达降低和LC3表达升高，随后减轻了oxLDL诱导的铁死亡，表现为VSMC死亡减少、GPX4表达和GSH含量增加，以及MDA含量、Fe含量和脂质过氧化水平降低。此外，雷帕霉素抑制mTOR可显著逆转oxLDL诱导的p62上调和LC3下调。结论 mTOR可能通过抑制自噬促进oxLDL诱导的VSMC铁死亡。
