甲硫氨酸限制对骨肉瘤细胞增殖、凋亡及铁死亡的影响
Effects of Methionine Restriction on the Proliferation,Apoptosis,and Ferroptosis of Osteosarcoma Cells
张馨丹 1赵正刚 1张文轩 1李芳红 1周素瑾 1赵子建1
作者信息
- 1. 广东工业大学生物医药学院 广东广州 510006
- 折叠
摘要
目的:探讨甲硫氨酸限制对骨肉瘤细胞增殖,凋亡和铁死亡的影响.方法:使用细胞计数检测剥夺甲硫氨酸对三株不同的骨肉瘤细胞系增殖的影响;高通量测序(RNA-seq)分析甲硫氨酸限制后骨肉瘤细胞的转录组学变化;流式细胞术检测细胞周期,凋亡以及ROS,脂质ROS水平;Western blot检测铁死亡关键蛋白谷胱甘肽过氧化物GPX4以及铁死亡标志蛋白前列腺素内过氧化物合成Cox-2的表达.结果:甲硫氨酸限制显著抑制骨肉瘤细胞的增殖(P＜0.001);RNA-seq分析筛选出1719个差异表达基因,基因富集分析发现甲硫氨酸限制可显著激活铁死亡通路并显著抑制细胞周期通路;细胞实验证实甲硫氨酸限制将骨肉瘤细胞阻滞在G2M期,显著诱导凋亡细胞比例增加;同时,细胞内Cox-2的表达增加,GPX4的活性降低,活性氧和脂质活性氧积累,最终导致细胞死亡,且这种作用可以被铁死亡抑制剂Fer-1部分挽救.结论:本研究的结果为甲硫氨酸限制治疗骨肉瘤提供了一定的科学依据.
Abstract
Objective:To investigate the effects of methionine restriction on the proliferation,apoptosis,and ferroptosis of os-teosarcoma cells.Methods:Three human osteosarcoma cell lines,MNNG-HOS,U2OS and Saos-2,were treated with normal or methion-ine-restricted medium,and the effects of methionine restriction on cell growth were assessed by cell counting;High-throughput sequenc-ing(RNA-seq)was employed to analyze the molecular changes in osteosarcoma cells following methionine restriction;Flow cytometry was used to evaluate the cell cycle,cell apoptosis,reactive oxygen species(ROS),and lipid ROS levels;Western blot analysis was con-ducted to determine the protein expression of key ferroptosis biomarkers GPX4 and COX2.Results:Methionine restriction significantly inhibited the proliferation of osteosarcoma cells(P＜0.001);RNA-seq analysis identified 1719 differentially expressed genes.The enrich-ment analysis indicated that the Ferroptosis pathway was significantly activate,and cell cycle pathway were significantly inhibited(P＜0.001);Cytological experiments showed that methionine restriction induced G2/M Cell Cycle arrest and apoptosis in osteosarcoma cells;Besides,methionine restriction could induce ROS,lipid ROS level,and decrease GPX4 protein expression,upregulate COX2 protein ex-pression in osteosarcoma cells.Moreover,methionine restriction Induced ferroptosis could be partially rescued by ferrostatin-1.Conclu-sions:Our findings provide potential novel therapeutic approaches or combination therapies for treating osteosarcoma while offering a theoretical basis for utilizing methionine restriction therapy against osteosarcoma.
关键词
甲硫氨酸限制/骨肉瘤/增殖/铁死亡Key words
Methionine restriction/Osteosarcoma/Proliferation/Ferroptosis引用本文复制引用
基金项目
国家重点计划研究发展项目(2018YFA0800603)
广东省珠江人才计划(2016ZT06Y432)
广东省重点领域研发计划(2019B020201015)
广东省基础与应用基础研究联合基金(2023A1515110036)
出版年
2024
NETL
NSTL
万方数据