LncRNA MEG3对人脑胶质瘤细胞增殖、侵袭及凋亡的影响

The influence of LncRNA MEG3 on proliferation,invasion and apoptosis of human glioma cells

目的应用靶向长链非编码RNA母系表达基因3(LncRNA MEG3)的过表达重组质粒来转染体外培养的人脑胶质瘤细胞,从而观察其对人脑胶质瘤细胞增殖、侵袭和凋亡的影响。方法应用qRT-PCR法检测正常脑组织细胞(N)和胶质瘤细胞系U251中LncRNA MEG3 mRNA的表达情况。构建pCI-MEG3真核过表达载体,转染体外培养的人脑胶质瘤细胞系U251,作为pCI-MEG3过表达质粒组(pCIMEG3组);空pCI质粒转染体外培养的人脑胶质瘤细胞系U251,作为空载体pCI组(pCI组);无质粒转染的人脑胶质瘤细胞系U251,作为空白对照组(Control组)。采用qRTPCR分析LncRNA MEG3基因表达变化,Western blot法分析LncRNA MEG3相关蛋白MDM2和p53表达变化,MTT法和细胞克隆技术分析胶质瘤细胞增殖能力,Transwell技术分析胶质瘤细胞侵袭能力,流式细胞技术分析胶质瘤细胞凋亡率。结果与正常脑组织细胞比较,胶质瘤细胞中LncRNA MEG3 mRNA表达水平降低。体外实验显示,与Control组和pCI组比较,pCI-MEG3组LncRNA MEG3 mRNA及p53蛋白表达水平均有上调(P<0.05),MDM2蛋白表达下调(P<0.05),细胞增殖能力受到抑制(P<0.01),细胞侵袭能力也受到抑制(P<0.01),细胞凋亡率升高(P<0.01)。结论人脑胶质瘤细胞中LncRNA MEG3的表达水平低于正常人脑组织细胞,靶向LncRNA MEG3基因的过表达重组质粒能够增加人脑胶质瘤细胞内LncRNA MEG3的表达,从而抑制胶质瘤细胞的增殖和侵袭能力,同时增加其凋亡率。

Objective To observe its effect on the proliferation,invasion and apoptosis of human glioma cells by the overexpression recombinant plasmid targeting to long non-coding maternally expressed gene 3(LncRNA MEG3)gene transfects the human glioma cells in vitro.Methods The expression of LncRNA MEG3 mRNA in normal brain tissue cells(N)and glioma cell line U251 cells was detected by qRT-PCR.Eukaryotic overexpression vector ofpCIMEG3 was constructed to transfect the human glioma cell line U251 cells as pCI-MEG3 overexpression plasmid group(pCI-MEG3 group).Those transfected with empty pCI plasmid were divided into empty vector pCI group(pCI group)and those without any transfection were divided into Control group.The expression of LncRNA MEG3 gene was analyzed by qRT-PCR.The expression of MDM2 and P53 of LncRNA MEG3-related protein was confirmed by Western blot.The proliferation ability of glioma cells was analyzed by techniques of MTT and cell cloning.Transwell was used to analyze the invasion ability of glioma cells,and flow cytometry was used to analyze the apoptosis rate of glioma cells.Results Compared with normal brain tissue cells,the expression level of LncRNA MEG3 mRNA decreased in glioma cells.The results of vitro experiments showed that both the mRNA of LncRNA MEG3 and protein expression levels of p53 increased,and the expression levels of MDM2 protein were down-regulated(P<0.05)in pCI-MEG3 group when compared with the control group and the pCI group(P<0.05).Cell proliferation and cell invasion were inhibited(P<0.01).The apoptosis rate increased(P<0.01).Conclusion The expression of LncRNA MEG3 in human glioma cells is lower than that in the normal human brain tissue cells.Overexpression of LncRNA MEG3 in human glioma cells can increase the expression of LncRNA MEG3 thereby inhibiting the proliferation and invasion of human glioma cells and increasing the apoptosis rate.