泛醌氧化还原酶铁硫蛋白4通过影响线粒体功能参与肺炎衣原体感染诱导的血管平滑肌细胞焦亡
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(天津医科大学基础医学院生理学与病理生理学系,天津市 300070)

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张雨珂,硕士研究生,研究方向为动脉粥样硬化发病的分子机制,E-mail:13781155403@163.com。通信作者张丽莙,博士,教授,博士研究生导师,研究方向为动脉粥样硬化发病的分子机制,E-mail:lijunwz@hotmail.com。

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国家自然科学基金资助项目(82070452)


Ubiquinone oxidoreductase iron-sulfur protein 4 participates in vascular smooth muscle cell pyroptosis induced by Chlamydia pneumoniae infection through affecting mitochondrial function
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Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Tianjin Medical University, Tianjin 300070, China)

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    目的]探究肺炎衣原体(C.pn)感染诱导血管平滑肌细胞(VSMC)焦亡及其可能机制。 [方法]组织贴块法培养大鼠原代VSMC,利用C.pn感染VSMC模型,使用倒置相差显微镜观察C.pn感染后VSMC形态学的变化,试剂盒检测C.pn感染VSMC后乳酸脱氢酶(LDH)含量变化,Western blot实验检测GSDMD和Caspase-1的表达,串联质谱标签法定量蛋白质组学实验和GO富集分析检测线粒体氧化磷酸化和氧化呼吸链Complex相关蛋白的变化。 [结果]与对照组相比,倒置相差显微镜下可见C.pn感染后VSMC膜外出现气泡状囊泡,C.pn感染VSMC 36 h、48 h后LDH含量分别增加38.92%和79.54%(均P<0.001),焦亡相关蛋白GSDMD表达增加1.74倍和1.67倍(均P<0.001);C.pn感染VSMC 48 h后Caspase-1表达(pro-Caspase-1)和活性(Caspase-1 p12/p10)分别增加2.69倍和3.47倍(均P<0.001);质谱结果显示,C.pn感染VSMC后有20种差异表达的蛋白质富集在氧化磷酸化通路中,同时发现,ComplexⅠ泛醌氧化还原酶铁硫蛋白4(NDUFS4)下降最为显著。进一步的Western blot实验结果显示,C.pn感染VSMC 36 h、48 h后NDUFS4的表达水平分别下降了57.5%和57%(均P<0.001)。 [结论]C.pn感染可能通过抑制NDUFS4表达影响线粒体功能,从而诱导VSMC焦亡。

    Abstract:

    Aim To explore the pyroptosis of vascular smooth muscle cell (VSMC) induced by Chlamydia pneumoniae (C.pn) infection and its possible mechanisms. Methods Primary rat VSMC were cultured by explant method. After the model of VSMC infected with C.pn was established, the changes in morphology of VSMC were observed under an inverted phase microscope, the lactic dehydrogenase (LDH) content was detected by the kit, and the expression levels of GSDMD and Caspase-1 were determined by Western blot, the changes in mitochondrial oxidative phosphorylation and the expression of complex-related proteins were measured by quantitative proteomic analysis by tandem mass tag technology and gene ontology. Results Compared with the control group, bubble-like vesicles were found outside the membrane of VSMC after C.pn infection under an inverted phase microscope. After C.pn infection of VSMC for 36 h and 48 h, LDH content increased by 38.92% and 79.54% (P<0.001), respectively, and the expression of pyropotosis-related protein GSDMD increased by 1.74 times and 1.67 times (P<0.001). After C.pn infection of VSMC for 48 h, the expression(pro-Caspase-1) and activity(Caspase-1 p12/p10)of Caspase-1 increased by 2.69 times and 3.47 times (P<0.001), respectively. The mass spectrometry results showed that there were 20 differentially expressed proteins enriched in the oxidative phosphorylation pathway after C.pn infection, and at the same time, ComplexⅠubiquinone oxidoreductase iron-sulfur protein 4 (NDUFS4) decreased significantly. Further Western blot results showed that the expression level of NDUFS4 decreased by 57.5% and 57% (P<0.001) after C.pn infection of VSMC for 36 h and 48 h respectively. Conclusion C.pn infection may induce VSMC pyroptosis by affecting mitochondrial function through downregulating NDUFS4 expression.

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张雨珂,赵茜,张利军,王蓓蓓,苗国琳,张琪,张丽莙.泛醌氧化还原酶铁硫蛋白4通过影响线粒体功能参与肺炎衣原体感染诱导的血管平滑肌细胞焦亡[J].中国动脉硬化杂志,2023,(7):573~580.

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  • 收稿日期:2022-10-10
  • 最后修改日期:2023-01-24
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  • 在线发布日期: 2023-09-01