| 引用本文: | 张华敏,程龙慧,董雪漫,叶璐,徐玉欣,陈霖,武璞,周建良.基于代谢组学的柠檬烯抗非小细胞肺癌作用机制研究[J].中国现代应用药学,2024,41(2):192-202. |
| ZHANG Huamin,CHENG Longhui,DONG Xueman,YE Lu,XU Yuxin,CHEN Lin,WU Pu,ZHOU Jianliang.Investigating the Mechanistic Insights of Limonene's Anti-non-small Cell Lung Cancer Effect Through Metabolomics Analysis[J].Chin J Mod Appl Pharm(中国现代应用药学),2024,41(2):192-202. |
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| 基于代谢组学的柠檬烯抗非小细胞肺癌作用机制研究 |
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张华敏1, 程龙慧1, 董雪漫1,2, 叶璐1, 徐玉欣1, 陈霖1, 武璞1,2, 周建良1
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1.杭州师范大学，杭州 311121;2.中南大学，长沙 410083
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| 摘要: |
| 目的 采用非靶向代谢组学等方法，探究柠檬烯抑制非小细胞肺癌增殖的作用机制。方法 以肺癌A549细胞为研究对象，通过CCK-8法测定柠檬烯抑制A549细胞活力及IC50；通过集落形成、流式细胞检测、铁含量测定及线粒体染色等实验，分别评价柠檬烯的体外抗肺癌及诱导铁死亡作用；代谢组学分析发现柠檬烯的潜在作用通路；最后采用Western blotting对相关通路主要蛋白进行验证。结果 与对照组相比，柠檬烯给药组可以显著抑制A549细胞的增殖及集落的形成，且呈剂量依赖性；光学显微镜观察发现，柠檬烯给药后A549细胞出现脱落现象，并可显著改变其形态；同时柠檬烯具有诱导A549细胞凋亡作用，并阻滞在G0-G1期；共聚焦显微镜发现柠檬烯作用后，A549细胞线粒体荧光减弱，同时细胞内铁含量亦显著增加，呈现典型的铁死亡表现；代谢组学研究发现谷胱甘肽(glutathione，GSH)代谢、精氨酸生物合成、D-谷氨酰胺和D-谷氨酸代谢及半胱氨酸和蛋氨酸代谢等多条差异代谢通路，这些通路与细胞内GSH合成密切相关；Western blotting实验发现，柠檬烯给药后细胞中SLC40A1、SLC7A11(xCT)及GPX4蛋白含量显著减少。结论 柠檬烯抗肺癌作用机制可能与降低肺癌细胞中GSH合成及增加Fe2+含量诱导其铁死亡有关。 |
| 关键词: 柠檬烯 非小细胞肺癌 代谢组学 铁死亡 |
| DOI：10.13748/j.cnki.issn1007-7693.20233146 |
| 分类号:R966 |
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| Investigating the Mechanistic Insights of Limonene's Anti-non-small Cell Lung Cancer Effect Through Metabolomics Analysis |
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ZHANG Huamin1, CHENG Longhui1, DONG Xueman1,2, YE Lu1, XU Yuxin1, CHEN Lin1, WU Pu1,2, ZHOU Jianliang1
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1.Hangzhou Normal University, Hangzhou 311121, China;2.Central South University, Changsha 410083, China
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| Abstract: |
| OBJECTIVE To elucidate the mechanisms responsible for the inhibitory effects of limonene on the proliferation of non-small cell lung cancer(NSCLC) by non-targeted metabolomics and additional approaches. METHODS The CCK-8 assay was utilized to evaluate the inhibitory effects of limonene on NSCLC A549 cell viability and to ascertain the IC50. In vitro experiments, encompassing colony formation, flow cytometry, iron content assessment, and mitochondrial staining, were conducted to assess the anti-lung cancer and iron-induced cell death effects of limonene. Metabolomic analysis was employed to identify potential pathways influenced by limonene, and Western blotting was carried out to validate pivotal proteins within these pathways. RESULTS In comparison to the control group, the limonene-treated group demonstrated a significant, dose-dependent reduction in A549 cell proliferation and colony formation. Optical microscopy revealed cellular detachment and pronounced changes in cellular morphology following exposure to limonene. Limonene induced apoptosis in A549 cells and arrested them in the G0-G1 phase of the cell cycle. Confocal microscopy unveiled diminished mitochondrial fluorescence and an augmented intracellular iron content, indicative of the classical phenomenon of ferroptosis. Metabolomic investigations unveiled divergent metabolic pathways, including glutathione(GSH) metabolism, arginine biosynthesis, D-glutamine and D-glutamate metabolism, as well as cysteine and methionine metabolism, with many of them intricately linked to intracellular GSH synthesis. Western blotting experiments underscored a marked reduction in the levels of SLC40A1, SLC7A11(xCT), and GPX4 proteins within the cells post-limonene treatment. CONCLUSION Limonene may induce ferroptosis in lung cancer cells by reducing GSH synthesis and increasing Fe2+ levels. |
| Key words: limonene non-small cell lung cancer metabolomics ferroptosis |
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