【摘要】 目的　利用流式细胞仪 (flowcytometry ,FCM)探讨反义IGF IR基因片段诱导胶质瘤细胞凋亡的作用。方法　根据IGF IRcDNA序列设计其正义、反义基因片段 ,并对部分碱基进行硫代磷酸修饰。实验分为空白对照组、正义核酸组和反义核酸组 ,应用流式细胞技术检测DNA含量 ,根据DNA直方图分析凋亡细胞情况。应用透射电镜观察了凋亡细胞的形态学变化。结果　经反义寡核苷酸 5、10、15、2 0μmol·L-1处理的胶质瘤细胞其凋亡率分别为 16 0 6 %±3 86 %、2 4 5 2 %± 4 84%、36 6 9%± 5 5 3%和 43 85 %±5 72 % ,而用正义寡核苷酸 5、10、15、2 0 μmol·L-1处理的胶质瘤细胞其凋亡率分别为 3 76 %± 1 2 2 %、3 14%±1 15 %、2 5 1%± 0 93%和 3 2 6 %± 1 15 % ,空白对照组(野生型 )自然凋亡率为 1 0 1%± 0 89%。反义核酸组与对照组相比有显著性差异 (P <0 0 5 ) ,而正义核酸组与空白对照组无明显差异 (P >0 0 5 )。凋亡细胞形态上表现为细胞体积缩小 ,染色质凝聚 ,并聚集于核膜下成新月状或块状 ,胞膜内馅形成膜包裹的核碎片或细胞器 ,最后出泡形成凋亡小体。结论　反义IGF IR基因片段诱导胶质瘤细胞凋亡是其抑制细胞增殖机制之一。通过FCM分析可较准确地判定肿瘤细胞凋亡情况更多还原

【Abstract】 Objective The apoptosis induced by antisense phosphorothioate oligodeoxynucleotides (antisense ODNs) was investigated by flow cytometry (FCM). Methods Antisense ODNs was designed to be complimentary to the 5′ portion of IGF IR mRNA a few bp of rat cells downstream of the initiation codon. Cultured glioma cells were divided into antisense, sense and untreated control groups according to different agents added to the media of cultured C6 cells, the DNA content was examined by FCM. The morphology of cell apoptosis was investigated by the transmission electron microscopy. Results There were cell apoptosis in antisense ODNs treated C6 cell. Flow cytometric analysis showed that apoptotic rate of C6 cell treated with antisense ODNs at concentration of 5 μmol·L -1 , 10 μmol·L -1 , 15 μmol·L -1 , 20 μmol·L -1 was 16 06%±3 86%, 24 52%±4 84%, 36 69%±5 53% and 43 85%±5 72% respectively, whereas the apoptotic rate of C6 cell treated with sense ODNs at same concentration was 3 76%±1 22%, 3 14%±1 15%, 2 51%±0 93% and 3 26%±1 15% respectively. There was significant different between antisense and untreated control group ( P <0 05). The morphological feature of apoptotic cell was chromatin condensation, aggregation to the nuclear margin and cytoplasmic condensation, the nuclear fragmentation and apoptotic body was observed in latter stage. Conclusion The apoptosis induced is the one of mechanism of inhibitory proliferation in malignant glioma cell by antisense ODNs. With FCM analysis, the apoptosis of tumor cell could be judged.更多还原