新型微管抑制剂YB-B1S诱导Hela细胞凋亡及其作用机制

New microtubule inhibitor of YB-B1S-induced apoptosis in Hela cells and its mechanism

目的:探讨吲哚3-草酰胺衍生物YB-B1S在体外实验中诱导宫颈癌细胞株(Hela细胞)发生凋亡及其作用机制。方法:采用MTT法、生长曲线法、流式细胞术、荧光显微镜及DNA ladder法检测YB-BIS对Hela细胞凋亡和细胞周期的作用;RT-PCR检测其对凋亡相关基因表达的影响;激光共聚焦显微镜观察YB-B1S对细胞骨架的影响。结果:YB-B1S在体外实验中对于Hela细胞有较强的生长抑制作用,有显著的量效关系,荧光显微镜观察到了凋亡小体;流式细胞术检测到了凋亡峰,同时发现YB-B1S诱导细胞阻滞于细胞周期的G2+M期;DNA ladder法检测到了凋亡时DNA降解形成的梯带;YB-B1S作用细胞24h后,caspase-3mRNA的表达增加,同时Bcl-2mRNA的表达降低(P<0.01);共聚焦显微镜观察到YB-B1S处理后的Hela细胞的细胞骨架遭到破坏。结论:YB-B1S体外可以显著抑制Hela细胞的增殖并诱导其凋亡,其机制可能与影响凋亡相关基因caspase-3及抑癌基因Bcl-2的表达,阻断细胞周期及抑制微管的聚合,破坏细胞骨架有关。

Objective ：To investigate whether the derivative of indol - 3 - glyoxylic acid amide induced apoptosis on human galactophore cancer Hela cells line （ Hela cells） in vitro and the mechanism. Methods： MTr assay , cell growth curve ,flow cytometry （ FCM ） , fluorescence microscopy, DNA ladder, RT - PCR, were carried out , and the changes of cytoskeleton was detected by confocal microscopy. Results： The eytotoxic effecet on Hela cells could be observed by MTT assay and cell growth curve, and this effect was in a dose - and time - dependent manner . After treatment of Hela cells with YB -B1S, typical apoptotic features were observed under the fluorescent microscopes by the Hoeehst staining. We also found the typical ＂sub - G1＂ peak and cells blocked in G2 ＋ M phase by flow cytome- try. RT- PCR showed that YB -B1S significantly up -regulated caspase -3 and down -regulated Bc1-2 mRNA expressions in Hela cells. The disrupted cytoskeleton can be found by the eonfoeal microscopy. Conclusion： YB - B1S showed obvious anticancer activity in vitro by inducing apoptosis and causing cell cycle arrest in the dose - dependent manner and can interfere with microtubule polymerization and disrupting cytoskeleton.