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首页> 外文期刊>African Journal of Biotechnology >Impact of pEGFP mediated ING4 gene on growth of glioma U251 cells and its potential molecular mechanism
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Impact of pEGFP mediated ING4 gene on growth of glioma U251 cells and its potential molecular mechanism

机译：pEGFP介导的ING4基因对神经胶质瘤U251细胞生长的影响及其潜在分子机制

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To investigate the impact of the inhibitor of growth family 4 (ING4) on growth of glioma cells (U251 cells) and its potential molecular mechanism, total RNA was extracted from?the?embryonic tissues and ING4 was amplified by RT-PCR and cloned into pEGFP-C2 vector. U251 cells were transfected with eukaryotic expression vector pEGFP-ING4 mediated by cationic polymers polyethylenimine (PEI). Flow cytometry and G418 were used to screen the cells successfully transfected with pEGFP-ING4. PEGFP-ING4 group, pEGFP group and blank control group (without transfection) were included in this study. Morphological examination, MTT assay and Hochest staining were employed to detect the proliferation and apoptosis of U251 cells. RT-PCR, immunohistochemitry and western blot were recruited to determine the mRNA and protein expression of ING4, respectively. ELISA was performed to measure the VEGF level in the supernatant of U251 cells. Sequencing of pEGFP-ING4 showed sequence correctness and the tranfection efficiency was 84% for pEGFP-ING4 and 82% for pEGFP. Results from RT-PCR, immunohistochemistry and western blot revealed significantly increased ING4 expression. When compared with the pEGFP group and blank group, the growth inhibition rate and apoptotic rate 72 h after transfection in the pEGFP-ING4 group were significantly higher (P 0.05). ING4 can be highly expressed in the pEGFP-ING4 group and remarkably suppress the growth of U251 cells through inducing apoptosis but not suppressing VEGF expression.
机译：为了研究生长家族4抑制剂（ING4）对神经胶质瘤细胞（U251细胞）生长的影响及其潜在的分子机制，从纤毛组织中提取总RNA，并通过RT-PCR扩增ING4并克隆到pEGFP-C2载体。用阳离子聚合物聚乙烯亚胺（PEI）介导的真核表达载体pEGFP-ING4转染U251细胞。流式细胞仪和G418用于筛选成功转染pEGFP-ING4的细胞。本研究包括PEGFP-ING4组，pEGFP组和空白对照组（无转染）。采用形态学检查，MTT法和Hochest染色法检测U251细胞的增殖和凋亡。分别采用RT-PCR，免疫组化和western blot方法分别测定ING4的mRNA和蛋白表达。进行ELISA以测量U251细胞上清液中的VEGF水平。 pEGFP-ING4的测序显示序列正确性，pEGFP-ING4的转染效率为84％，pEGFP的转染效率为82％。 RT-PCR，免疫组织化学和蛋白质印迹的结果表明ING4表达显着增加。与pEGFP组和空白组相比，pEGFP-ING4组转染后72 h的生长抑制率和凋亡率显着更高（P 0.05）。 ING4可以在pEGFP-ING4组中高表达，并通过诱导凋亡显着抑制U251细胞的生长，但不抑制VEGF的表达。

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