摘要
目的 探讨 PTEN基因对人脑胶质瘤 SHG- 4 4细胞凋亡及凋亡相关基因 bcl- 2表达的影响 ,阐明 PTEN基因抑制肿瘤细胞增殖的机制 .方法 PTEN基因体外转染 SHG- 4 4细胞 ,筛选阳性细胞克隆 ,以原位杂交、免疫组化方法检测PTEN基因的表达情况 ;采用透射电镜、流式细胞仪和核DNA琼脂糖凝胶电泳检测细胞的凋亡情况 ;以免疫荧光法检测 bcl- 2基因的表达 .结果 PTEN基因转染的 SHG- 4 4细胞有 PTEN蛋白的表达 .透射电镜下可见转染 PTEN基因后细胞核染色质浓缩边集、胞质浓缩、核碎裂及凋亡小体形成等典型的凋亡表现 ;流式细胞仪显示细胞周期从 G1 期到 S期发生抑制 ,并且在 G1 期峰前出现一明显的凋亡峰 (15 .9% ) ;细胞核 DNA琼脂糖凝胶电泳呈现凋亡细胞特有的梯状条带 ;bcl-2的表达也显著下调 .结论 PTEN基因可诱导 SHG- 4 4细胞凋亡 ,并下调 bcl- 2蛋白的表达 ,这可能是
AIM To investigate the effects of exotic PTEN on the apoptosis and expression of bcl 2 gene in human brain glioma SHG 44 cells and to probe into the mechanism thatPTEN gene inhibits the proliferation of the tumor cells. METHODS Human glioma SHG 44 cells were transfected with a plasmid vector containing PTEN gene in vitro , and then positive cell clones were selected and amplified. The expression of PTEN gene was examined by in situ hybridization and immunohistochemistry methods. To detect the apoptosis, examinations of transfected cells were performed by transmission electronic microscope, flow cytometry and nucleus DNA agarose gel electrophoresis. The expression of bcl 2 was also detected by immunofluorescent methods. RESULTS SHG 44 cells stably transfected with PTEN gene were obtained by selection, and expression of PTEN gene was able to be detected. Apoptosis body and changes of ultrastructure, including the concentration, side accumulation of the nuclear chromatin and the fragmentation of the nuclear, were detected by transmission electron microscope. Cell cycle detected by flow cytometry showed that the percentage of cells increased in G1 and decreased in S phase. One visible apoptosis peak (15.9%) prior to the peak of G1 also was found by FCM. The characteristic DNA ladder was found in the agarose gel electrophoresis of the nucleus DNA of transfected cells. The expression of bcl 2 gene was down regulated in SHG 44 cells. CONCLUSION PTEN gene SHG 44 cells can induce apoptosis and down regulation of expression of bcl 2 gene in SHG 44 cells, which may be one of the reasons that PTEN gene inhibits the proliferation of human tumor cells.
出处
《第四军医大学学报》
北大核心
2002年第8期681-684,共4页
Journal of the Fourth Military Medical University
基金
国家自然科学基金资助项目 (3 9970 75 2 )
