EFHD2调节NOX4/ROS通路启动糖代谢重编程促进乳腺癌进展的机制研究
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1.川北医学院基础医学与法医学院人体解剖学教研室,四川 南充 637000;2.川北医学院基础医学与法医学院免疫学教研室,四川 南充 637000;3.川北医学院临床医学院,四川 南充 637000

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R-33

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EFHD2 regulates the NOX4/ROS pathway to initiate glucose metabolism reprogramming and promote breast cancer progression
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1. Department of Human Anatomy, Institute of Basic Medicine and Forensic Medicine, North Sichuan Medical College, Nanchong 637000, China. 2. Department of Immunology, Institute of Basic Medicine and Forensic Medicine, North Sichuan Medical College, Nanchong 637000. 3. School of Clinical Medicine, North Sichuan Medical College, Nanchong 637000

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    摘要:

    目的 基于NOX4/ROS信号通路探讨EFHD2影响乳腺癌发生进展的作用机制。 方法 将细胞分为NC-shRNA组和EFHD2-shRNA组,构建沉默EFHD2表达的慢病毒载体及其对照载体后转染乳腺癌细胞MDA MB-23、MCF-7,qRT-PCR验证转染效率;CCK8检测细胞增殖活性;平板克隆实验检测细胞集落形成能力;划痕实验检测细胞迁移;Transwell实验检测细胞侵袭;流式细胞术检测细胞凋亡及ROS水平;qRT-PCR检测GLUT1、PDK1、PFK1、PKM2、PDH、LDH mRNA的表达;Western blot检测Cleaved caspase-3、MMP-2、NOX4蛋白的表达。 结果 与NC-shRNA组相比,EFHD2-shRNA组细胞中EFHD2的表达明显降低,细胞存活及其集落形成能力减弱;细胞凋亡率及促凋亡蛋白Cleaved caspase-3的表达升高;细胞迁移距离缩短,细胞侵袭数及促迁移、侵袭蛋白MMP-2的表达降低;乳酸及GLUT1、PDK1、PFK1、PKM2、LDH的水平降低,ATP及PDH的水平升高;流式结果表明沉默EFHD2后ROS水平降低,NOX4蛋白下调。 结论 EFHD2可通过调节NOX4/ROS信号通路,抑制ROS生成,引起乳酸及葡萄糖堆积从而促进乳腺癌细胞凋亡,抑制细胞增殖、迁移及侵袭。

    Abstract:

    Objective To investigate the mechanism by which EFHD2 affects the occurrence and progression of breast cancer based on the NOX4/ROS signaling pathway. Methods Cells were divided into an NC-shRNA group and EFHD2-shRNA group. A lentiviral vector for EFHD2 silencing and a control vector were constructed and used to transfect MDA-MB-23 and MCF-7 breast cancer cells. The transfection efficiency was verified by qRT-PCR. A CCK8 assay was used to detect cell proliferation activity. A plate cloning assay was employed to measure cell colony formation ability. A scratch test was used to detect cell migration, and a Transwell assay used to assess cell invasion. Flow cytometry was applied to detect apoptosis and ROS levels. qRT-PCR was used to analyze the mRNA expression of GLUT1, PDK1, PFK1, PKM2, PDH, and LDH, while Western blot was applied to detect the expression of Cleaved caspase-3, MMP-2, and NOX4 proteins. Results Compared with the NC-shRNA group, the EFHD2-shRNA group’s EFHD2 expression was significantly decreased and its cell survival and colony formation ability were weakened. The apoptosis rate and the expression of the pro-apoptotic protein Cleaved caspase-3 increased. The cell migration distance was shortened, while the number of invading cells and the expression of MMP-2, which promotes migration and invasion, were decreased. The levels of lactic acid and GLUT1, PDK1, PFK1, PKM2, and LDH decreased, while the levels of ATP and PDH increased. Streaming result showed that ROS levels were reduced and NOX4 protein was down-regulated after silencing EFHD2. Conclusions EFHD2 inhibits ROS production by regulating the NOX4/ROS signaling pathway, causing lactic acid and glucose accumulation, promoting the apoptosis of breast cancer cells, and inhibiting cell proliferation, migration, and invasion.

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徐明飞,张 怡,康如意,刘超越. EFHD2调节NOX4/ROS通路启动糖代谢重编程促进乳腺癌进展的机制研究[J].中国比较医学杂志,2024,34(9):66~75.

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  • 收稿日期:2024-03-05
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  • 在线发布日期: 2024-10-30
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