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本研究通过下调c-Met来观察白血病细胞株K562对抗肿瘤药物硼替佐米的敏感性,以此探索c-Met信号对白血病细胞株的增殖、凋亡等的影响,为临床上寻求治疗白血病新方案提供理论依据。本研究首先采用MTT实验初步判断硼替佐米对K562细胞的增殖能力的影响;再构建c-Met干扰载体转染K562细胞,并采用Real-time PCR检测干扰效果,筛选最适干扰质粒。然后采用筛选的c-Met干扰质粒转染细胞,分别使用MTT法检测细胞增殖能力、软琼脂克隆形成实验检测细胞克隆能力、流式细胞术检测细胞凋亡情况。结果显示硼替佐米对K562细胞的增殖具有显著抑制作用(p<0.05),且这种抑制作用具有浓度依赖性。c-Met干扰质粒构建及干扰效果检测实验中发现,随着转染时间的延长,绿色荧光信号越来越强,表明转染效率越来越高。但Real-time检测结果发现,3个干扰质粒中仅c-Met-Homo-1和c-Met-Homo-3干扰质粒对c-Met的表达具有明显抑制效果(p<0.05),且c-Met-Homo-3干扰质粒干扰效果较好。使用c-Met-Homo-3干扰质粒转染细胞后发现,c-Met下调能够显著增强硼替佐米对K562细胞增殖的抑制作用(p<0.05),且增强硼替佐米对K562细胞克隆形成的抑制能力,同时增强了硼替佐米诱导的K562细胞的凋亡。本研究结论为,下调c-Met能够增加白血病细胞株K562对硼替佐米的化疗敏感性。
This study was to investigate the effect of c-Met signaling on the proliferation and apoptosis of leukemia cell line by investigating the sensitivity of leukemia cell line K562 to bortezomib against leukemia cell line K562 by down-regulating c-Met. Program provides a theoretical basis. In this study, MTT assay was used to determine the effect of bortezomib on the proliferation of K562 cells. The c-Met interference vector was transfected into K562 cells. Real-time PCR was used to detect the interference effect and the optimal interference plasmid was screened. Then, the transfected cells were screened by c-Met interference plasmids, MTT assay was used to detect cell proliferation, colony formation assay was used to detect cell cloning ability, and flow cytometry was used to detect apoptosis. The results showed that bortezomib significantly inhibited the proliferation of K562 cells (p <0.05), and the inhibitory effect was concentration-dependent. c-Met interference plasmid construction and interference effect detection experiments found that with the extension of transfection time, the green fluorescent signal is getting stronger and stronger, indicating that transfection efficiency is getting higher and higher. Real-time results showed that only c-Met-Homo-1 and c-Met-Homo-3 interference plasmids inhibited the expression of c-Met significantly (p <0.05), and c- Met-Homo-3 interference plasmid interference is better. C-Met-Homo-3 interference plasmid transfected cells found that c-Met downregulation can significantly enhance the inhibitory effect of bortezomib on K562 cell proliferation (p <0.05), and enhance the bortezomib on K562 cell clone formation Inhibition, while enhancing the bortezomib-induced apoptosis in K562 cells. This study concluded that downregulation of c-Met can increase chemosensitivity to bortezomib in leukemia cell line K562.