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转录因子c-Jun对人血管平滑肌细胞中线粒体融合基因2表达调控作用的实验研究 预览

Experimental study on regulation of transcription factor c-Jun on the expression of mitofusin 2 in human vascular smooth muscle cells
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摘要 目的:探讨转录因子c-Jun对培养的人血管平滑肌细胞(hVSMCs)中线粒体融合基因2(Mfn2)表达的调控作用。方法:①siRNA干扰实验:设正常对照组、阴性siRNA对照组和siRNA干扰组,用c-Jun siRNA转染hVSMCs,培养26 h后提取细胞mRNA和总蛋白。通过RT-PCR和Western blot检测Mfn2基因和蛋白质的表达。②过表达实验:设正常对照组、阴性病毒对照组和慢病毒过表达组。用慢病毒载体感染细胞,培养96 h后提取细胞mRNA和总蛋白。通过RT-PCR和Western blot检测Mfn2基因和蛋白质的表达。结果:siRNA敲减c-Jun后,与阴性siRNA对照和正常对照组相比,siRNA干扰组Mfn2 mRNA明显降低[(0.671±0.061)vs.(1.110±0.080),P<0.01;(0.671±0.061)vs.(1.000±0),P<0.01],蛋白质的表达也显著降低[(0.571±0.049)vs.(1.042±0.039),P<0.01;(0.571±0.049)vs.(1.000±0),P<0.01],hVSMCs的数量明显增加。相反,用慢病毒表达载体过表达c-Jun后,与阴性病毒对照和正常对照组相比,慢病毒过表达组Mfn2 mRNA明显增加[(1.414±0.063)vs.(1.056±0.020),P<0.01;(1.414±0.063)vs.(1.000±0),P<0.01],蛋白质的表达也显著增加[(1.449±0.044)vs.(1.089±0.026),P<0.01;(1.449±0.044)vs.(1.000±0),P<0.01],hVSMCs的数量则明显下降。结论:c-Jun能显著正调控Mfn2的表达,并影响hVSMCs的增殖。因此,它可能是Mfn2表达的转录调控因子。 Objective:To investigate the regulation of transcription factor c-Jun on the expression of Mfn2 in cultured human vascular smooth muscle cells(hVSMCs).Methods:1)siRNA interference experiment:Normal control group,negative siRNA control group and siRNA interference group were set up,and hVSMCs were transfected with c-Jun siRNA.After 26 hours of transfection,the cell RNA and total protein were extracted.The expression of Mfn2 gene and protein was detected by RT-PCR and Western blot.(2)Overexpression experiment:Normal control group,negative virus control group and lentivirus overexpression group were set up.Lentivirus vector was used to infect the cells.After 96 hours of incubation,the cell RNA and total protein were extracted.The expression of Mfn2 gene and protein was detected by RT-PCR and Western blot.Results:After knocking down c-Jun in cultured hVSMCs with siRNA,compared with negative siRNA control group and normal control group,the expression of Mfn2 mRNA in siRNA interference group decreased significantly[(0.671±0.061)vs.(1.110±0.080),P<0.01;(0.671±0.061)vs.(1.000±0),P<0.01],and the expression of protein decreased significantly[(0.571±0.049)vs.(1.042±0.039),P<0.01;(0.571±0.049)vs.(1.000±0),P<0.01],while the number of hVSMCs increased significantly.On the contrary,after overexpression of c-Jun with lentivirus expression vector,the Mfn2 mRNA expression in lentivirus overexpression group was significantly increased[(1.414±0.063)vs.(1.056±0.020),P<0.01;(1.414±0.063)vs.(1.000±0),P<0.01],and the protein expression was also significantly increased[(1.449±0.044)vs.(1.089±0.026),P<0.01;(1.449±0.044)vs.(1.000±0),P<0.01],while the number of hVSMCs decreased signifi-cantly.Conclusions:c-Jun can significantly and positively regulate the expression of Mfn2 and affect the proliferation of hVSMCs.Therefore,it may be a transcriptional regulator of Mfn2 expression.
作者 李笑 罗兴才 王佐广 吴琼 辛毅 刘洁琳 刘雅 魏永祥 LI Xiao;LUO Xingcai;WANG Zuoguang;WU Qiong;XIN Yi;LIU Jielin;LIU Ya;WEI Yongxiang(Department of Hypertension,Beijing Anzhen Hospital,Capital Medical University,Beijing Institute of Heart,Lung and Blood Vessel Diseases,Beijing 100029,China)
出处 《心肺血管病杂志》 2019年第5期538-543,共6页 Journal of Cardiovascular and Pulmonary Diseases
基金 国家自然科学基金(81370229,81270216,81870335) 北京市自然科学基金(7102045) 北京市卫生系统高层次卫生技术人才培养计划(2014-3-040)资助.
关键词 线粒体融合基因2 C-JUN 转录因子 转录调控 血管平滑肌细胞 Mitofusin 2 C-Jun Transcription factor Transcript regulation Vascular smooth muscle cells
作者简介 通信作者:王佐广,医学博士,硕士生导师,主要从事高血压的基础与临床等研究。E-mail:wzg70@hotmail.com;通信作者:魏永祥,医学博士,博士生导师,主要从事阻塞性睡眠呼吸暂停低通气综合征与高血压等研究。E-mail:wyhhzq818@sina.com.
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