LncRNA MEG3对脑胶质瘤细胞替莫唑胺获得性耐药的影响

The influence of LncRNA MEG3 in temozolomide resistance of glioma cells

目的探讨长链非编码RNA母系表达基因3(LncRNA MEG3)对胶质瘤细胞替莫唑胺(TMZ)获得性耐药的影响。方法构建TMZ耐药胶质瘤细胞U87(U87/TMZ),qRT-PCR法检测LncRNA MEG3表达水平,MTT法检测细胞增殖能力。用脂质体法使U87/TMZ细胞中LncRNA MEG3过表达(pcDNA-MEG3组),转染空载体作为空载体组(pcDNA组),同时以常规培养的U87/TMZ细胞作为空白组(Control组),并用10μg/ml TMZ处理。采用qRT-PCR评估质粒转染效果;Western blot法检测LncRNA MEG3相关蛋白表达;细胞克隆实验检测LncRNA MEG3对细胞增殖的影响;Transwell实验检测LncRNA MEG3对细胞侵袭的影响;流式细胞术检测LncRNA MEG3对细胞凋亡的影响,并构建小鼠移植瘤动物模型进行体内实验。结果U87/TMZ细胞中LncRNA MEG3表达水平低于U87(P<0.01),并且U87/TMZ的细胞活力高于U87(P<0.01)。过表达LncRNA MEG3联合TMZ后,与Control组和pcDNA组比较,pcDNA-MEG3组上调了LncRNA MEG3 mRNA及p53蛋白表达,下调了MDM2蛋白表达,且pcDNA-MEG3组增殖和侵袭能力降低,凋亡能力增加(P<0.01)。构建小鼠移植瘤动物模型,与pcDNA组相比,pcDNA-MEG3组肿瘤体积和质量降低(P<0.01)。结论过表达LncRNA MEG3能减弱TMZ耐药胶质瘤细胞的耐药性。

Objective To explore the effect of LncRNA MEG3 in temozolomide(TMZ)resistance of glioma cells.Methods TMZ-resistant glioma cells U87(U87/TMZ) were constructed,qRT-PCRwas used to detect the LncRNA MEG3 expression level, and MTT was used to detect the cell proliferation ability. The LncRNA MEG3 in U87/TMZ was overexpressed by liposome method (pcDNA-MEG3 group), the empty vectoRwas transfected as the empty vectoRgroup (pcDNA group), and the conventionally cultured U87/TMZ was used as the blank group (Control group),then treated with 10 μg/ml TMZ. The plasmid transfection effect was analyzed by qRT-PCR. The expression of LncRNA MEG3-related protein was confirmed by Western blot. The cell cloning experiment detects the effect of LncRNA MEG3 on cell proliferation. The effect of LncRNA MEG3 on cell invasion was tested by Transwell. The effect of LncRNA MEG3 on cell apoptosis was detected by flow cytometry. And mouse transplant tumoRanimal model was constructed foRin vivo experiments. Results Compared with U87, the expression level of LncRNA MEG3 in U87/TMZ was loweR( P< 0.01), and the cell viability of U87/TMZ was higheRthan that of U87 ( P< 0.01). AfteRoverexpression of MEG3 combined with TMZ, compared with the Control group and the pcDNA group, the pcDNA-MEG3 group up-regulated the expression of MEG3 mRNA and p53 protein, and down-regulated the expression of MDM2 protein, and the proliferation and invasion ability of the pcDNA-MEG3 group decreased while the apoptosis ability increased ( P< 0.01). The construction of mouse transplant tumoRanimal model showed that the tumoRvolume and mass of the pcDNA-MEG3 group were reduced compared with the pcDNA group ( P< 0.01). Conclusion Overexpression of LncRNA MEG3 can attenuate the drug resistance of TMZ-resistant glioma cells.