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目的:观察针对Survivin基因的短发卡RNA(shRNA)对人脑胶质母细胞瘤U251细胞在体外凋亡的影响。方法:对U251细胞,稳定转染Survivin基因shRNA真核表达载体pWH1-SR的U251-SR细胞,以及稳定转染空载体pWH1的U251-P细胞,分别采用相差显微镜、HE染色、Hoechst染色、TUNEL染色以及透射电镜观察各组细胞的形态学特征;采用流式细胞术(FCM)定量测定各组细胞的凋亡细胞数量。结果:相差显微镜、HE染色、Hoechst染色、TUNEL染色以及透射电镜观察显示,U251-SR凋亡细胞明显增多,并呈现典型的细胞凋亡形态学改变;FCM定量分析凋亡细胞数量显示,与U251(2.1%)和U251-P(2.7%)相比,U251-SR凋亡细胞增加约6倍,达14.4%。结论:针对Survivin基因的shRNA能够在体外诱导U251细胞发生大量凋亡。
Objective: To observe the effect of Survivin gene short hairpin RNA (shRNA) on human glioblastoma U251 cells in vitro apoptosis. Methods: U251 cells, U251-SR cells stably transfected with Survivin gene shRNA eukaryotic expression vector pWH1-SR and U251-P cells stably transfected with empty vector pWH1 were detected by phase contrast microscopy, HE staining, Hoechst staining, TUNEL Staining and transmission electron microscopy were used to observe the morphological characteristics of each group. Flow cytometry (FCM) was used to quantitatively determine the number of apoptotic cells in each group. Results: The apoptotic cells of U251-SR were significantly increased by HE staining, Hoechst staining, TUNEL staining and transmission electron microscopy. The morphological changes of typical apoptotic cells were observed. The quantitative analysis of apoptotic cells by FCM showed that U251- (2.1%) compared to U251-P (2.7%), U251-SR apoptotic cells increased about 6-fold to 14.4%. Conclusion: shRNA targeting Survivin gene can induce a large number of apoptosis in U251 cells in vitro.