【摘要】 目的:探讨白藜芦醇治疗高血压的作用靶点及相关作用机制。方法:利用血管紧张素Ⅱ诱导大鼠胸大动脉平滑肌细胞A7r5模拟高血压时平滑肌细胞增殖细胞模型,并使用白藜芦醇与miR-21-5p mimic进行干预。利用CCK-8实验检测A7r5细胞活力;利用细胞划痕实验检测A7r5细胞的迁移能力;利用real-time PCR技术检测miR-21-5p的表达;利用Western blot技术检测细胞增殖相关蛋白PCNA、Cyclin D1、CDK4与细胞迁移相关蛋白MMP2、MMP9、PDCD4的表达。此外,选取对数生长期293T细胞,利用双荧光素酶报告基因检测技术验证miR-21-5p与PDCD4的靶向关系。结果:与正常对照组相比,模型组细胞活力、迁移能力及增殖和迁移相关蛋白PCNA、Cyclin D1、CDK4、MMP2、MMP9与miR-21-5p的表达水平显著上调,PDCD4蛋白的表达水平显著下降,且差异具有统计学意义(P <0.01);与模型组相比,白藜芦醇处理组的细胞活力、迁移能力及增殖和迁移相关蛋白PCNA、Cyclin D1、CDK4、MMP2、MMP9与miR-21-5p的表达水平显著下调,PDCD4蛋白的表达水平显著上调,且差异具有统计学意义(P<0.001);而miR-21-5p mimic转染后细胞活力、迁移能力及增殖和迁移相关蛋白PCNA、Cyclin D1、CDK4、MMP2、MMP9与miR-21-5p的表达水平显著上调,PDCD4蛋白的表达水平显著下降且差异具有统计学意义(P<0.001),双荧光素酶检测显示miR-21-5p可显著抑制野生型PDCD4 3’UTR报告基因载体的荧光素酶活性(P <0.01)。结论:白藜芦醇通过调节miR-21-5p/PDCD4通路,抑制血管平滑肌细胞增殖与迁移,进而对高血压起到一定的治疗作用。更多还原

【Abstract】 Objective:To explore the target and related mechanism of resveratrol in the treatment of hypertension. Methods:Using angiotensin Ⅱ to induce rat aortic smooth muscle cells A7r5 to establish a hypertensive cell model and interventing with resveratrol and miR-21-5p mimic on this basis. CCK-8 assay was used to detect cell viability of A7r5 cells,wounding healing assay was used to detect migration ability of A7r5 cells,Real-time PCR was used to detect expression of miR-21-5p,Western blot was used to detect the expression of cell proliferation-related proteins PCNA,Cyclin D1 and CDK4,and cell migration associated proteins MMP2,MMP9 and PDCD4. In addition,the targeting relationship between miR-21-5p and PDCD4 was verified by dual luciferase reporter gene detection technology in logarithmic growth phase 293T cells. Results:Compared with the NC group,the cell viability,migration ability and the expression level of proliferation and migration-related proteins PCNA,Cyclin D1,CDK4,MMP2,MMP9 and miR-21-5p were significantly up-regulated,and the expression level of PDCD4 protein was significantly decreased in the NC+ model group,and the difference was statistically significant(P < 0.01);Compared with the NC+ model group,the cell viability,migration ability,and the expression level of proliferation and migration-related proteins PCNA,Cyclin D1,CDK4,MMP2,MMP9 and miR-21-5p were significantly down-regulated,and the expression level of PDCD4 protein was significantly up-regulated in the NC+ model+resveratrol group,and the difference was statistically significant(P < 0.01);Compared with the NC+ model+resveratrol group,cell viability,migration ability and the expression level of proliferation and migration related proteins PCNA,Cyclin D1,CDK4,MMP2,MMP9 and miR-21-5p was significantly up-regulated,and the expression level of PDCD4 protein was significantly decreased in the miR-21-5p mimic+model+resveratrol group,and the difference was statistically significant(P < 0.01). The dual luciferase assay miR-21-5p significantly inhibited the luciferase activity of the wild-type PDCD4 3’UTR reporter vector(P < 0.01). Conclusion:Resveratrol inhibits the proliferation and migration of vascular smooth muscle cells by regulating the miR-21-5p/PDCD4 pathway,which in turn has a certain therapeutic effect on hypertension.更多还原