UbcH10基因沉默对多发性骨髓瘤U-1996细胞增殖和凋亡的作用研究

The effects of UbcH10 gene silencing on proliferation and apoptosis research of multiple myeloma U-1996 cell

目的采用脂质体法转染siRNA的方法沉默多发性骨髓瘤U-1996细胞中的UbcH10基因,研究基因沉默后U-1996细胞增殖活性及凋亡等生物学特性的变化情况。方法根据UbcH10基因信息,设计针对UbcH10基因编码序列(CDS)区的siRNA序列。通过脂质体法转染相关siRNA序列至多发性骨髓瘤U-1996细胞。在转染siRNA后48h,采用实时定量聚合酶链反应及蛋白免疫印迹法检测U-1996细胞中UbcH10基因的mRNA和其蛋白水平。选取未转染siRNA序列的空白组和转染阴性siRNA序列组作为试验对照,采用CCK-8法检测转染siRNA序列后24、48、72hU-1996细胞的增殖情况,采用流式细胞术检测48h后U-1996细胞的凋亡情况。结果通过脂质体法成功转染构建siRNA序列至U-1996细胞。转染后的U-1996细胞中UbcH10基因的mRNA及蛋白表达情况较转染前均明显下降,差异有统计学意义(P<0.05)。和对照组相比,沉默UbcH10基因后U-1996细胞的增殖活性下降,差异有统计学意义(P<0.05),细胞凋亡率上升,差异也有统计学意义(P<0.05)。结论干扰UbcH10基因表达可显著抑制多发性骨髓瘤细胞的增殖活性并增加细胞凋亡。

Objective To investigate the effects of UbcH10 gene silencing on proliferation and apoptosis of myeloma cell line U-1996.Methods According to the UbcH10 genetic information,the siRNA sequence targeting UbcH10 was designed.It was transfected into U-1996 cells via lipofectamine2000.UbcH10 mRNA and protein were examined 48 hafter transfection FQ-PCR and western blot,respectively.Cells without transfection were served as blank controls and those transfected with negative sequence as negative controls.The cell proliferation was detected by CCK-8assay 24 h,48hand 72 hafter transfection,and the cell apoptosis was detected by flow cytometry 48 hafter transfection.Results The siRNA sequence was transfected into U-1996 cells via lipofectamine successfully.The expression level of UbcH10 mRNA and protein was significantly decreased than before transfection（P〈0.05）.Compared with control group,the proliferative activity of U-1996 cells after UbcH10 gene silencing was significantly decreased（P〈0.05）,the apoptosis rate was significantly incresased（P〈0.05）.Conclusion UbcH10 gene silencing can significantly inhibit the proliferative activity of U-1996 cells and increase the cells apoptosis.