miR-181a诱导慢性淋巴细胞白血病细胞周期阻滞及细胞凋亡的分子机制探讨

Study on the molecular mechanism of miR-181a induceing cell cycle arrest and apoptosis of chronic lymphocytic leukemia

目的:探讨miR-181a在慢性淋巴细胞白血病细胞周期阻滞及细胞凋亡中的作用并初步阐明其调控机制。方法:利用荧光定量PCR检测慢性淋巴细胞白血病患者肿瘤细胞中miR-181a的表达水平;利用双荧光素酶试验验证miR-181a的靶基因MCL-1和BCL-2,利用免疫印迹试验检测过表达miR-181a对p53通路中关键蛋白MCL-1和BCL-2的影响,利用流式细胞仪检测过表达miR-181a对慢性淋巴细胞白血病细胞凋亡和周期的影响。结果:RT-qPCR结果显示,慢性淋巴细胞白血病细胞中miR-181a的表达下调;双荧光素酶试验验证发现,p53通路下游基因MCL-1和BCL-2均是miR-181a的靶基因;免疫印迹试验结果显示,过表达miR-181a抑制了MCL-1和BCL-2蛋白的表达;流式细胞仪检测结果显示,过表达miR-181a阻滞细胞周期G1/S期转换,促进细胞凋亡。结论:miR-181a通过靶向作用p53通路中关键基因MCL-1和BCL-2,诱导细胞周期阻滞及细胞凋亡,可能在慢性淋巴细胞白血病中扮演着抑癌基因的角色。

Objective:To explore the role of miR-181 a induces cell cycle arrest and apoptosis of chronic lymphoid leukemia and to elucidate its regulatory mechanism.Methods:The expression level of miR-181 a in tumor cells of patients with chronic lymphocytic leukemia was detected by real-time qPCR.The target genes MCL-1 and BCL-2 of miR-181 a were verified by double luciferase assay.The effect of overexpression of miR-181 a on the key proteins MCL-1 and BCL-2 in the p53 pathway was detected by Western-blot assay.The effect of overexpression of miR-181 a on cell apoptosis and cycle of chronic lymphocytic leukemia was detected by flow cytometry.Results:RT-qPCR showed that the expression of miR-181 a was downregulated in chronic lymphocytic leukemia cells.The double luciferase assay confirmed that the downstream genes of p53 pathway MCL-1 and BCL-2 were the target genes of miR-181 a.Western-blot showed overexpression of miR-181 a inhibited the expression of MCL-1 and BCL-2 proteins.Flow cytometry showed that overexpression of miR-181 a blocked cell cycle G1/S conversion and promoted apoptosis.Conclusion:miR-181 a induces cell cycle arrest and apoptosis by targeting the key genes MCL-1 and BCL-2 in p53 pathway,which may play the role of anti-oncogene in chronic lymphoid leukemia.