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用黄芩苷处理体外培养的血管平滑肌细胞(VSMC),采用流式细胞分析、免疫细胞化学染色、Western印迹及免疫共沉淀等方法,探讨不同浓度的黄芩苷对血小板源性生长因子(PDGF)诱导的VSMC增殖的影响及其作用机制。结果表明,与单独PDGF刺激组比较,黄芩苷预处理可明显抑制PDGF诱导的细胞增殖和迁移活性,同时伴有增殖和迁移相关蛋白PCNA、VCAM-1、ICAM-1、cyclinE、CDK2的表达减少;细胞周期分析显示,黄芩苷处理可明显减少处于细胞周期S时相的细胞数,而G_0/G_1期细胞增加,黄芩苷抑制增殖和迁移、阻滞细胞周期进程的作用具有浓度依赖性。免疫共沉淀分析证实,黄芩苷可抑制cyclinE-CDK2复合物的形成,上调p27蛋白水平,二者变化程度具有反相关关系。结果提示,黄芩苷是一种具有抗VSMC增殖和迁移活性的天然单体,其作用机制可能与抑制cyclinE-CDK2复合物的形成和上调p27水平有关。
Vascular smooth muscle cells (VSMCs) cultured in vitro were treated with baicalin and flow cytometric analysis, immunocytochemistry staining, Western blotting, and immunoprecipitation were used to investigate the effects of different concentrations of baicalin on platelet-derived growth factor (PDGF) induction. The effect of VSMC proliferation and its mechanism of action. The results showed that baicalin pretreatment significantly inhibited PDGF-induced cell proliferation and migration compared with PDGF-stimulated group alone, accompanied by decreased expression of proliferation and migration-related proteins PCNA, VCAM-1, ICAM-1, cyclinE, and CDK2. Cell cycle analysis showed that baicalin treatment significantly decreased the number of cells in the cell cycle S phase, while G_0 / G 1 phase cells increased, baicalin inhibit proliferation and migration, blocking the cell cycle progression in a concentration-dependent manner. Co-immunoprecipitation analysis confirmed that baicalin can inhibit the formation of cyclinE-CDK2 complex and up-regulate the level of p27 protein, and there is an inverse correlation between them. The results suggest that baicalin is a natural monomer with anti-VSMC proliferation and migration activity, and its mechanism may be related to inhibition of cyclinE-CDK2 complex formation and up-regulation of p27 levels.