人21．5kDaMBP基因沉默对神经胶质瘤细胞增殖与凋亡的作用研究

摘要

目的：研究人21．5kDa人脑髓鞘碱性蛋白（MBP）基因沉默对神经胶质瘤U251细胞增殖与凋亡的影响。方法将21．5kDaMBP序列特异性短发夹RNA（shRNA）重组质粒pGenesil‐1‐MBP‐3转染神经胶质瘤细胞株（U251）作为MBP基因沉默组，以空质粒转染为阴性对照组，脂质体转染为脂质体空转组；用实时定量PCR（RT‐PCR）和Westernblot方法检测各组21．5kDaMBPmRNA和蛋白的表达水平，CCK‐8法测定细胞增殖曲线，流式细胞法分析细胞凋亡率。结果与阴性对照组相比，MBP基因沉默组细胞中21．5kDaMBP在mRNA和蛋白水平的表达均显著下降（P＜0．05）、细胞增殖活性明显降低（P＜0．05）、细胞凋亡率显著增高（P＜0．05）。结论人21．5kDaMBP基因沉默对神经胶质瘤细胞U251具有显著的抑制增殖和促进凋亡的双重调节作用。%Objective To investigate the effects of silencing of the human cerebral 21 .5 kDa myelin basic protein (MBP) gene on the proliferation and apoptosis of the glioma U251 cells .Methods The 21 .5 kDa MBP sequence‐specific short hair‐pin RNA (shR‐NA) recombinant plasmids pGenesil‐1‐MBP‐3 were transfected into the human glioma cell line(U251) ,the cells of U251 was used as MBP silencing group ,the cells transfected with negative control plasmids used as negative control group ,and the cells transfected with liposomes used as blank control group .Real‐time PCR and Westernblot were used to detect the expression levels of the 21 .5 kDa MBP mRNA and protein in each group ,and the cell proliferation curve was measured by CCK‐8 assay ,the apoptosis rate was a‐nalysised by Flow cytometry .Results Both the mRNA and the protein expression levels of the 21 .5 kDa MBP of MBP silencing group were significantly lower than those in the control groups (P<0 .05);the cellular proliferation activity of the MBP silencing group decreased significantly (P<0 .05)while the cellular apoptotic rate increased significantly (P<0 .05) .Conclusion Silencing of the human cerebral 21 .5 kDa MBP gene may playa dual role in the inhibition of proliferation and the promotion of apoptosis of the glioma U251 cells .