6-溴异香兰素诱发人宫颈癌上皮细胞纺锤体损伤及有丝分裂灾变死亡

The Spindle Damage and Mitotic Catastrophe Induced by 6-bromoisovanillin in HeLa Cells

研究香兰素衍生物中的6-溴异香兰素(6-溴-5-羟基-4-甲氧基苯甲醛,BVAN08)对人宫颈癌上皮(HeLa)细胞纺锤体结构、有丝分裂的影响,及杀死癌细胞的相关机制,为开发新的抗癌药物提供理论依据。研究发现BVAN08对体外培养的人HeLa细胞具有剂量依赖性致死效应。通过抗α-微管蛋白抗体和抗β-微管蛋白抗体免疫荧光标记和激光共聚焦显微镜观察发现其机制是:BVAN08对细胞纺锤体结构造成破坏,诱发产生大量的多中心体细胞,使细胞阻滞在有丝分裂期。异常多极纺锤体细胞的百分率与BVAN08呈浓度依赖效应关系。荧光染料Hoechst33258染色观察到多微核或细胞核割裂和染色质凝集现象,也证实BVAN08诱发Hela细胞有丝分裂灾变死亡。同时免疫印迹实验还发现,BVAN08可导致细胞周期转录调节因子FoxM1(Forkhead box)及其下游靶分子CyclinB1的降解,参与有丝分裂的FoxM1蛋白失活,这表明BVAN08通过抑制FoxM1蛋白的表达是导致细胞纺锤体装配异常、细胞发生有丝分裂灾变死亡的分子机制之一。研究结果提示BVAN08诱发Hela细胞新的死亡方式,具有被开发为新型抗癌药物的潜力。

This paper studies the effect of vanillin derivative 6-bromoisovanillin （6-bromine-5-hydroxy-4-methoxy-benzaldehyde,BVAN08） on the spindle structure damage and mitotic catastrophe of HeLa cells,to provide a solid experimental basis for development of new anticancer drugs. BVAN08 was shown to induce death of HeLa cells in a dose-dependent manner. By laser confocal microscopic observation of BVAN08 -treated cells immunostained with the anti -α -tubulin antibody and anti -γ -tubulin antibody,the following mechanisms were found. It is indicated that BVAN08 destroys the spindle structure （aberrant multi-polar spindles）,induces aberrant multi-polar spindles and multinucleated cells,and causes cell arrest in the mitosis stage. The percentage of abnormal multipolar spindle cells was dose -dependent. Hoechst 33258 fluorescent staining observation of BVAN08 -treated cells revealed multiple micronuclei or multinucleated and chromatin aggregation. Western blot analysis indicated that the cell cycle control associated transcription regulation factor FoxM1 （Forkhead box） and its downstream target molecule CyclinB1 protein were down-regulated. Inactivation of FoxM1 protein indicated that the degradation of FoxM1 protein induced by BVAN08 is one of the mechanisms of abnormal spindle assembly and mitotic catastrophe. The result indicates that BVAN08 has the potential to be developed as a new anticancer drug.