新生霉素通过诱导DNA损伤抑制慢粒白血病细胞增殖  

Novobiocin inhibits CML cells proliferation by inducing DNA damage

作　　者：黄立森 陈显凌[3,4] 柯方[5] 许建文[1,2] 郑鸣[6] 许建华[1,2] 吴丽贤[1,2] 

机构地区：[1]福建医科大学药学院药理系,福建福州350004 [2]福建省天然药物药理学重点实验室,福建福州350004 [3]福建医科大学附属协和医院血液科,福建福州350001 [4]福建省血液病研究所,福建福州350001 [5]福建医科大学药学院药化系,福建福州350108 [6]福建医科大学解剖系,福建福州350108

出　　处：《中国药理学通报》2014年第3期69-76,共8页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No 30901824;81173096);福建省自然科学基金杰出青年项目(No 2011J06013);福建省高校跨世纪优秀人才项目(No JA11101)

摘　　要：目的研究新生霉素(novobiocin,Nov)在体外抑制慢粒白血病(CML)细胞增殖作用与诱导DNA损伤的关系。方法MTT及羟基荧光素二醋酸盐琥珀酰亚胺脂(CFSE)染色法检测Nov对CML细胞增殖的影响;流式细胞术检测CML细胞反应性氧自由基(ROS)含量、DNA损伤数量、细胞周期阻滞情况和细胞凋亡的比例;Western blot探讨Nov对DNA损伤、细胞周期和细胞凋亡调控通路相关蛋白表达的影响。结果 Nov明显抑制CML细胞增殖,半数抑制率(IC50)分别为(232.50±0.22)μmol·L-1和(237.10±0.13)μmol·L-1,减少CML细胞的增殖分裂代数;Nov增加细胞ROS水平,诱导细胞阻滞在G2/M期并增加凋亡率,呈剂量依赖关系;Nov增加H2AX、ATM、P53的磷酸化水平以及Parp和Caspase-3的切割,而减少CDC25A和CDC25C的表达。结论Nov通过激活ROS产生,诱导CML细胞DNA的损伤和线粒体途径激活的细胞凋亡。Aim To investigate the effect of Novobio- ein（Nov） on its eytotoxieity against CML eells, and to explore its underlying mechanisms by which Nov might induce DNA damage and apoptosis through reactive oxygen species （ROS）. Methods MTT and CFSE were used to measure the proliferation inhibition ratio of K562 and K562/CO1 cells; Flow eytometry （FCM） was used to teste the level of extraeellular ROS, DNA damage, cell cycle progression, mitoehondria mem- brane potential （MPP） and apoptosis; Western blot was used to verify the amount of proteins. Result Nov significantly inhibited the proliferation of CML cells with ICso （232.50 ± 0.22） ·mol · L-1 and （237.10 ±0. 13） ·mol · L-1, respectively. Nov in- creased the generation of intracellular ROS, followed by induction of DNA damage, activation of the ATM- p53-r-H2AX pathway and checkpoint-related signals CHK1/CHK2, which led to increased numbers of cells in the S and G2/M phases of the cell cycle. Further- more, Nov induced apoptosis through decrease of mito- ehondtial membrane potential and activation of caspase- 3 and PARP. The above eft＇eeLs were all prevented by the ROS scavenger N-aeetyleysteine. Conclusion These findings therefore suggest that Nov induced DNA damage and mitoehondria-dependent cell apoptosis in CML cells are mediated via ROS generation.

关 键 词：新生霉素 ROS DNA损伤 r-H2AX 细胞周期检测点 细胞周期阻滞 凋亡 

分 类 号：R730.5[医药卫生—肿瘤]