利奈唑胺影响MEG-01巨核细胞增殖及血小板前体生成的代谢组学研究-中国药房

利奈唑胺影响MEG-01巨核细胞增殖及血小板前体生成的代谢组学研究

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篇名：	利奈唑胺影响MEG-01巨核细胞增殖及血小板前体生成的代谢组学研究
TITLE：	Metabolomics study on the effect of linezolid on the proliferation of MEG-01 megakaryocytes and the generation of platelet precursors
摘要：	目的研究利奈唑胺（LZD）对MEG-01巨核细胞代谢及细胞增殖和血小板前体生成的影响，为阐明LZD诱导的血小板减少症的发病机制提供参考。方法将MEG-01细胞接种于增殖培养基中，分为空白对照组（无处理）、溶剂对照组（4‰二甲基亚砜）和100、200、400、800μg/mLLZD组，培养4d后光镜下观察各组细胞增殖状态，进行细胞计数并检测细胞增殖活力。将细胞接种于分化培养基中，分为空白对照组（无处理）、溶剂对照组（4‰二甲基亚砜）和400μg/mLLZD组，培养14d后光镜下观察细胞的伪足生成情况，统计生成伪足的细胞比例及测量伪足相对长度，测定细胞中CD41、CD42bmRNA的表达水平。提取分化培养基中培养14d的溶剂对照组和400μg/mLLZD组细胞，运用液相色谱-串联质谱技术进行非靶向代谢组学和靶向能量代谢组学检测；在此基础上测量和观察添加丙酮酸（10mmol/L）、LZD（400μg/mL）+丙酮酸（10mmol/L）培养14d后细胞中丙酮酸相对含量及其对细胞增殖和伪足生成的影响。结果400μg/mLLZD可显著抑制MEG-01细胞增殖和伪足的生成（P＜0.01）。400μg/mLLZD处理后，MEG-01细胞的非靶向代谢组学分析结果显示，mTOR信号通路，丙氨酸、天冬氨酸和谷氨酸代谢以及癌症的中心碳代谢等与能量代谢相关的途径发生了显著变化；进一步的靶向能量代谢组学分析结果显示，400μg/mLLZD处理后MEG-01细胞中三磷酸腺苷、鸟苷三磷酸、丙酮酸相对含量较溶剂对照组显著减少（P＜0.01），乳酸相对含量较溶剂对照组显著增加（P＜0.01）。与LZD组相比，LZD+丙酮酸组细胞中丙酮酸相对含量、细胞计数、生成伪足的细胞比例和伪足相对长度均显著升高/增加（P＜0.01）。结论LZD可能通过抑制线粒体能量代谢导致丙酮酸生成减少，进而抑制巨核细胞增殖和血小板前体生成，最终导致血小板减少症的发生。
ABSTRACT：	OBJECTIVE To investigate the metabolic changes in MEG-01 megakaryocytes after treatment with linezolid （LZD） from metabolomic perspective and its impact on the the proliferation of cells and generation of platelet precursors. METHODS MEG-01 cells were seeded in proliferation medium and divided into blank control group （untreated）， solvent control group （4‰DMSO）， and 100， 200， 400， 800 μg/mL LZD groups. The proliferation status of cells in each group was observed under the microscope； cell proliferation and viability were assessed. Cells were also seeded in differentiation medium and divided into blank control group （untreated）， solvent control group （4‰DMSO）， and 400 μg/mL LZD group； after 14 days of culture， platelet precursor generation was observed under the microscope； immunofluorescence staining was performed to count the proportion of cells producing pseudopodia， the relative length of pseudopodia was measured， and the expression levels of CD41 and CD42b mRNA were assessed. Cells from the solvent control group and the 400 μg/mL LZD group， cultured in differentiation medium for 14 days， were extracted and subjected to non-targeted metabolomics and targeted energy metabolomics analysis using liquid chromatography-tandem mass spectrometry. The relative content of pyruvate in cells， after being cultured for 14 days with the addition of pyruvate （10 mmol/L） or LZD （400 μg/mL）+pyruvate （10 mmol/L）， was measured and observed， as well as its effects on cell proliferation and platelet precursor generation. RESULTS 400 μg/mL LZD could significantly inhibit the proliferation of MEG-01 cells and the generation of platelet precursors （P＜0.01）. Non-targeted metabolomic analysis of MEG-01 cells after 400 μg/mL LZD treatment revealed significant changes in energy metabolism-related pathways such as mTOR signaling pathway， alanine， aspartate and glutamate metabolism， and central carbon metabolism in cancer. Targeted energy metabolomic analysis further showed that the relative contents of adenosine triphosphate， guanosine triphosphate， and pyruvate in MEG-01 cells were significantly reduced （P＜0.01）， while the relative contents of lactate were significantly increased （P＜0.01）. Compared with the LZD group， the relative content of pyruvate， cell count， the proportion of cells producing pseudopodia and the relative length of pseudopodia were significantly increased in the LZD+pyruvate group （P＜0.01）. CONCLUSIONS LZD may reduce pyruvate production by inhibiting mitochondrial energy metabolism， thereby suppressing megakaryocyte proliferation and platelet precursor production， ultimately leading to the occurrence of thrombocytopenia.
期刊：	2024年第35卷第23期
作者：	王宁；杨雅；熊丽蓉；孙凤军；田衍平；夏培元
AUTHORS：	WANG Ning，YANG Ya，XIONG Lirong，SUN Fengjun，TIAN Yanping，XIA Peiyuan
关键字：	利奈唑胺；血小板减少症；血小板前体；能量代谢；丙酮酸
KEYWORDS：	linezolid； thrombocytopenia； platelet precursor； energy metabolism； pyruvate
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