丹参酮ⅡA对体外兔血管平滑肌细胞增殖和迁移的影响

目的观察丹参酮A对体外兔血管平滑肌细胞(VSMC)增殖和迁移的影响,并初步探讨丹参酮A的作用机制。方法采用兔胸主动脉,用组织贴块法培养,设立空白对照组,加入不同浓度的丹参酮A共同孵育24 h、48 h和72 h,采用MTT法观察该药对细胞增殖的影响;划痕法观测细胞迁移情况;采用流式细胞仪分析细胞周期和DNA含量;TRITC-鬼笔环肽标记F-actin,观察该药物对细胞骨架微丝结构的影响。结果1同一时间点,丹参酮A各浓度组显著抑制体外培养兔VSMC增殖和迁移。细胞的A570值(24 h、48 h、72 h分别为r=-0.762,P=0.000;r=-0.837,P=0.000;r=-0.944,P=0.000)、迁移距离(24 h、48 h、72 h分别为r=-0.966,P=0.000;r=-0.980,P=0.000;r=-0.966,P=0.000)与浓度呈负相关;2作用24 h后,处于G0/G1期的VSMC百分比增加,细胞DNA含量减少,凋亡率增加。兔VSMC在G0/G1期所占比例(r=0.962,P=0.000)、凋亡率(r=0.982,P=0.000)和浓度呈正相关;3药物干预组和对照组相比,兔VSMC的细胞骨架结构有所改变,对照组中细胞骨架呈极性分布,定向伸展,可见伪足;药物干预组细胞骨架呈非极性分布,未见伪足。结论丹参酮A对体外兔VSMC的增殖和迁移有抑制作用,上述作用可能是通过阻滞兔VSMC通过细胞周期的限制点,使其停滞于G0/G1期,诱导细胞凋亡以及影响细胞骨架微丝结构来实现。

Tanshinone Ⅱ A Inhibits in vitro Cellular Proliferation and Migration of Vascular Smooth Muscle Cell of Rabbit

OBJECTIVE: To study the influence of Tanshinone II A on the proliferation and migration of rabbit vascular smooth muscle cell in vitro and to primarily probe into the mechanism of how Tanshinone II A works. METHODS: Rabbit aortic artery was chosen for primary culture of vascular smooth muscle cells (VSMC), which were used and divided into control and Tanshinone II A groups with different drug concentrations. After cells, according to the protocol, incubated with drug for 24 h, 48 h or 72 h respectively, MTT assay was used to study the effect of Tanshinone II A on the VSMC proliferation; sterile scraper and contrast phase microscope were used to measure the distance of VSMC migration; analyses of cell cycle and DNA content were performed by flow cytometry. Then after TRITC-phalloidin labeling for cytoskeleton filament (F-actin), the influence of Tanshinone II A on the cytoskeleton was revealed by laser confocal microscope. RESULTS: (1) At the same time point, Tanshinone II A inhibited the proliferation and migration of rabbit VSMCs in vitro significantly in a dose-dependent manner; (2) After 24 h, compared with control group Tanshinone II A increased the ratio of rabbit VSMCs in G0/G1 phase, and decreased DNA content significantly in a dose-dependent manner; (3) Tanshinone II A influenced rabbit VSMCs cytoskeleton filament, including relatively short filaments arranged within nonpolarized cells and no lamellipodial protrusions, while in control group formation of lamellipodial protrusions and actin filaments parallel to long axis were seen. CONCLUSION; Tanshinone II A can significantly inhibit the proliferation and migration of rabbit VSMCs in vitro, which may result from arresting rabbit VSMCs through the check point of cell cycle, accumulating them in G0/G1 phase, inducing cell apoptosis and influencing the cytoskeleton filament.