MicroRNA-34a抑制葡萄膜黑色素瘤细胞增殖的研究

The Mechanism of MicroRNA-34a-mediated Inhibition of Human Uveal Melanoma Cell Proliferation

该文采用阳离子脂质体Lipofectamine介导的方法将microRNA-34a转染入体外培养的人葡萄膜黑色素瘤细胞M23和SP6.5。应用BrdU法、细胞平板克隆形成实验检测转染microRNA-34a后对细胞增殖的影响,发现M23和SP6.5细胞增殖明显被抑制(P<0.01);并利用流式细胞技术检测转染microRNA-34a后细胞周期的变化,发现细胞停滞于G_1期;同时检测转染microRNA-34a后细胞caspase-3/7酶的活性,发现无明显改变。另外,Real-time PCR检测表明阿霉素处理后M23、SP6.5细胞中microRNA-34a的表达量上调(P<0.01)。用阿霉素处理转染microRNA-34a的M23、SP6.5细胞,检测caspase-3/7酶活性的改变,发现caspase-3/7酶活性显著增加(P<0.01)。本研究表明microRNA-34a通过抑制细胞周期来抑制体外培养的人葡萄膜黑色素瘤细胞的增殖,能够增加细胞对阿霉素的敏感性,但不直接诱导细胞凋亡。

MicroRNA-34a was transfected into uveal melanoma cells M23 and SP6.5 by lipofectamine. The proliferation of uveal melanoma cells was examined by BrdU and colony-forming assay, respectively, and transfection of microRNA-34a into uveal melanoma cells led to a significant decrease in cell growth （P〈0.01）. Flow cytometry was applied to analyze cell cycle and these cells were found to have a higher proportion of cell cycle arrest at the G1 phase. The activity of caspase-3/7 had no significant changes. In addition, the expression of microRNA- 34a in uveal melanoma cells after treatment with adriamycin was upregulated based on real-time PCR （P〈0.01）. The activity of caspase-3/7 increased significantly after microRNA-34a transfection and treatment with adriamycin （P〈0.01）. These results indicate that microRNA-34a inhibits proliferation of uveal melanoma cells by cell cycle arrest. Furthermore, microRNA-34a increases cell sensitivity to adriamycin, but doesn＇t induce apoptosis directly.