【摘要】 目的利用RNA干扰(RNA interference,RNAi)技术特定沉默胶质瘤U251细胞株的血小板源生长因子-B(PDGF-B)基因,观察其对U251细胞株细胞凋亡和增殖的影响。方法利用脂质体将针对PDGF-B基因的siRNA转染进入U251细胞,利用实时荧光定量多聚核苷酸链式反应(RTPCR)检测PDGF-B基因表达;Western blot检测显示siRNA转染组PDGF-B蛋白表达,采用MTT法检测胶质瘤U251细胞的增殖,应用流式细胞计数观察抑制PDGF-B基因后胶质瘤U251细胞的凋亡情况。结果 RT-PCR检测PDGF-B基因表达明显下降;Western blot检测显示siRNA转染组PDGF-B蛋白表达明显抑制(抑制率>60%),MTT结果显示siRNA转染组U251细胞增殖较对照组明显降低;流式细胞学检测提示降低PDGF-B在胶质瘤细胞的表达能抑制胶质瘤细胞的有丝分裂,促进细胞的凋亡。结论构建针对胶质瘤细胞PDGF-B的RNA干扰质粒并转染人胶质瘤U251细胞株后,可明显抑制U251细胞株PDGF的表达,对人胶质瘤U251细胞株有明显的生长抑制和促进凋亡作用。更多还原

【Abstract】 Objective This experiment used RNA interference technology to silence platelet derived growth factor subunit B( PDGF-B) gene of U251 cell line and its influence on the apoptosis and proliferation of U251 cell line was observed. Methods siRNA was transfected into U251 cells by liposomal and the cells formed a doublestranded siRNA recognition and degradation of PDGF-B mRNA. Real-time quantitative polymerase chain reaction( RT-PCR) was used for the detection of the expression of PDGF-B; Western blot for the detection of expression of PDGF-B protein; methyl thiazolyl tetrazolium( MTT) for the detectino of 251 cell proliferation; flow cytometry for the dection of cell apoptosis. Results RT-PCR showed expression of PDGF-B gene was significantly decreased. Western blot showed that expression of PDGF-B protein in siRNA transfected group was significantly inhibited( inhibition rate>60%). MTT assay showed that in siRNA transfection group the proliferation of U251 cells was significantly lower than that in the control group; flow cytometric showed that siRNA can effectively inhibit the mitosis and promote the apoptosis of glioma cells. Conclusion After siRNA transfection of the PDGF-B gene into U251 cells by liposomal,the expression of PDGF in U251 cell line was signficantly inhibited,and accordingly it inhibits the growth and promotes the apoptosis of human glioma U251 cell line.更多还原