动脉粥样硬化中CARHSP1基因的表达对缺氧诱导的血管内皮细胞活力凋亡及免疫因子的影响

Effects of CARHSP1 gene expression on viability,apoptosis and immune factors of vascular endothe-lial cells induced by hypoxia in atherosclerosis

目的:探讨动脉粥样硬化斑块中钙调节热稳定蛋白1(CARHSP1)基因的表达对缺氧诱导的血管内皮细胞活力、凋亡及白细胞介素6(IL-6)和C-反应蛋白(CRP)表达的影响。方法:用Western blot检测动脉粥样硬化斑块中CARHSP1的蛋白表达;缺氧处理人脐静脉内皮细胞(HUVECs),将细胞分为正常培养组、缺氧组、缺氧+CARHSP1-siRNA组和缺氧+pc DNA3. 1-CARHSP1组,用CCK-8法及流式细胞术分别检测各组细胞活力及凋亡率; RT-PCR检测IL-6和CRP的表达; Western blot检测凋亡蛋白caspase-3、cleaved caspase-3、Bcl-2和Bax的蛋白水平。结果:CARHSP1在动脉粥样硬化斑块中的蛋白表达显著高于对照组(P <0. 05);缺氧可明显增加CARHSP1的表达。缺氧组HUVECs活力及Bcl-2表达显著低于正常培养组,细胞凋亡率及IL-6、CRP、cleaved caspase-3和Bax的蛋白水平显著高于正常培养组(P <0. 05);与缺氧组比较,缺氧+CARHSP1-siRNA组的活力及Bcl-2表达显著升高,细胞凋亡率及IL-6、CRP、cleaved caspase-3和Bax的蛋白水平显著降低(P <0. 05),pc DNA3. 1-CARHSP1组的细胞活力及Bcl-2表达显著降低,细胞凋亡率及IL-6、CRP、cleaved caspase-3和Bax表达显著升高(P <0. 05)。结论:CARHSP1在动脉粥样硬化斑块中表达升高,抑制CARHSP1表达可提高HUVECs的活力,降低细胞凋亡,下调免疫因子IL-6和CRP的表达,而过表达CARHSP1则反之。

AIM:To investigate the effect of calcium-regulated heat stable protein 1(CARHSP1)gene expression on the viability,apoptosis and expression of interleukin-6(IL-6)and C-reactive protein(CRP)in vascular endothelial cells induced by hypoxia.METHODS:The protein expression of CARHSP1 was detected by Western blot in atherosclerotic plaques.Human umbilical vein endothelial cells(HUVECs)were treated with hypoxia,and the cells were divided into normal culture group,hypoxia group,hypoxia+CARHSP1-siRNA group and hypoxia+pcDNA3.1-CARHSP1 group.The viability and apoptotic rate of the HUVECs were measured by CCK-8 assay and flow cytometry,respectively.The mRNA expression of IL-6 and CRP was detected by RT-PCR.The protein levels of caspase-3,cleaved caspase-3,Bcl-2 and Bax were determined by Western blot.RESULTS:The protein expression of CARHSP1 in atherosclerotic plaques was significantly higher than that in control group(P<0.05).Hypoxia significantly increased the expression of CARHSP1.The cell viability and the protein expression of Bcl-2 were significantly lower in hypoxia group than those in normal culture group(P<0.05).The apoptotic rate and the protein levels of IL-6,CRP,cleaved caspase-3 and Bax were significantly higher than those in normal culture group(P<0.05).Compared with hypoxia group,the cell viability and protein expression of Bcl-2 were significantly increased in hypoxia+CARHSP1-siRNA group,while the apoptotic rate and the protein levels of IL-6,CRP,cleaved caspase-3 and Bax were decreased significantly(P<0.05).The cell viability and protein expression of Bcl-2 were decreased significantly in hypoxia+pcDNA3.1-CARHSP1 group,while the apoptotic rate and the protein levels of IL-6,CRP,cleaved caspase-3 and Bax were increased significantly(P<0.05).CONCLUSION:The expression of CARHSP1 is increased in atherosclerotic plaques,and inhibition of CARHSP1 expression improves the viability,reduces the apoptosis,and down-regulates the expression of IL-6 and CRP in the HUVECs.Over-expression of CARHSP1 exerts the opposite effect.