Camptothecin(CPT)和Cytosine arabinoside(Ara-C)联合应用对MOLT-4细胞株周期特异性凋亡的影响  

Effects of camptothecin and cytosine arabinoside on cell cycle specificity of apoptotic cells during combination treatments in MOLT-4 cell lines in vitro

作　　者：喻春钊[1] 吴丰华[2] 余源[1] 肖薇[1] 李小兰[1] 陶德定[1] 吴剑宏[1] 龚建平[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院分子医学中心/肿瘤研究所/外科,武汉430030 [2]华中科技大学同济医学院生物化学分子生物学系

出　　处：《肿瘤》2004年第4期349-353,共5页Tumor

基　　金：973肿瘤计划 (G19980 5 12 12 );卫生部科研基金项目 (No .P70 70 2 3 9)

摘　　要：目的　以急性T淋巴细胞白血病细胞株MOLT— 4为模型 ,探讨作用于同一细胞周期的化疗药物Camptothecin(CPT)和Cytosinearabinoside(Ara C)联合应用时是否具有抗癌增效作用、引起细胞凋亡的周期时相性是否发生改变及对细胞周期检测点 (ckeckpoint)的影响。 方法　喜树碱 (CPT) ( 0 .2 5 μmol/L) ,阿糖胞苷 (Ara C) ( 1.0 0 μmol/L) ,及喜树碱 (CPT) ( 0 .2 5 μmol/L) +阿糖胞苷 (Ara C) ( 1.0 0 μmol/L)二药共同诱导急性早幼粒白血病细胞株MOLT 4细胞 4h ,分别采用激光共聚焦显微镜 (Confocal)观察其细胞的形态变化 ,采用Sub G1法 ,API法及cyclins/DNA双参数法 ,运用流式细胞术分析凋亡、凋亡细胞的周期时相性及CyclinE的表达规律。以Westernblot印迹法检测Bcl 2蛋白的表达。结果　喜树碱和阿糖胞苷联合应用时 ,凋亡细胞的数目明显增加 ,二者具有协同效应。发生凋亡的细胞大多数仍然是S期细胞。CyclinE的表达升高 ,bcl 2蛋白的表达降低。结论　作用于同一细胞周期的化疗药物CPT和Ara C联合应用时 ,具有协同效应但并不改变细胞周期的作用点(Checkpoint)。CyclinE和Bcl 2蛋白似乎在药物作用时起着关键性的调节作用 ,并对检测点的敏感性产生重要影响。Objective To determine whether Camptothecin(CPT) could augment Cytosine arabinoside(Ara-C) activity and could change the phase of apoptosis in the cell cycle during combination treatments. Methods CPT,Ara-C,alone or in combination were added to a flask of Molt-4 cells in log phase growth, giving a final concentration of CPT (0.25 uM), Ara-C(1.00 uM) or CPT (0.25uM)+Ara-C(1.00uM).Incubation experiments with Ara-C or CPT were performed on MOLT-4 cells that were treated for 4 h.To observe cellular apoptotic shapes with confocal laser scanning microscopy.Furthermore,three different flow cytometric methods,including Sub-G1, API Assay and CyclinE/DNA, were used to analyze the cell cycle specificity of apoptosis and expression of CyclinE. For further analysis, express of Bcl-2 were assayed by western blot. Results CPT could augment Ara-C activity and increase the amount of apoptotic cells 。A synergistic interaction in the two drugs was found.Most apoptotic cells were observed in S phase of the cell cycle.Expression of CyclinE had been elevated and expression of Bcl-2 had been decreased. Conclusion There is a synergistic interaction in the two drugs combinations in S phase of the cell cycle.The interactions still occur at the same checkpoint in the cell cycle.Both CyclinE and Bcl-2 play a critical role in response to chemotherapy agents and checkpoint status has been implicated in cell sensitivity to these drugs.

关 键 词：流式细胞术 细胞周期 凋亡 化学疗法 检测点(Checkpoint) 

分 类 号：R730.231[医药卫生—肿瘤]