组蛋白去乙酰酶抑制剂MS-275抑制膀胱癌的实验研究

Experimental study in vitro on histone deacetylase inhibitor MS-275 inhibiting bladder cancer

背景与目的:染色质核心组蛋白的乙酰化水平受组蛋白乙酰转移酶和组蛋白去乙酰酶(HDAC)的控制,它与基因表达调控密切相关、HDACs功能异常与肿瘤的发生发展有关,组蛋白去乙酰酶抑制剂(HDAC Is)通过抑制HDACs而抗肿瘤。本文研究HDAC I类药物之一MS-275对人膀胱癌T24细胞的抑制作用,为临床应用提供有价值的实验依据。方法:将不同浓度的MS-275作用于T24细胞,用MTT法检测生长抑制作用,用平板克隆形成试验测定细胞增殖能力的影响,分别采用光学显微镜、透射电子显微镜及AnnexinV、PI双标流式细胞术观察和检测细胞的凋亡情况。结果:MS-275对人膀胱癌T24细胞的生长抑制作用存在明显的剂量、时间依赖关系,0.5μmol/L作用72 h的抑制率为9.34%±3.57%,而8μmol/L作用120 h达94.84%±1.33%。克隆形成率从阴性对照组的55.67%±6.51%降至4μmol/L组的2.33%±0.58%。显微镜下可观察到大量的凋亡形态特征的细胞。流式细胞术检测示,作用48 h后的凋亡率在1μmol/L组、2μmol/L组和4μmol/L组分别为24.19%、30.77%和38.51%,明显高于阴性对照组(2.49%)。结论:组蛋白去乙酰酶抑制剂类药物具有体外抗膀胱癌作用,其重要作用机制之一是诱导膀胱癌细胞凋亡,并有望成为新的抗膀胱肿瘤药物。

Background and purpose： The acetylation level in nucleosomal histones, which is controlled by interactions between histone aeetyl transferases and histone deacetylases（ HDACs）, is closely involved in regulation of gene expression. The abnormal function of HDACs is related to development of neoplasms. Histone deacetylase inhibitors（HDACIs） have antitumor effects through inhibiting HDACs. The purpose of this paper is to investigate the antitumor effect of MS-275, a histone deacetylase inhibitor, on human bladder cancer T24 cell, and provide a valuable research basis for clinical application. Methods： Cultured T24 cells were exposed to different concentrations of MS-275 for different durations. The cell growth inhibition was evaluated by MTF assay. The effect on proliferating ability of T24 cell was determined by clonogenic assay on plate. The cell apoptosis was qualitatively detected by optical microscope, electronic microscope and AnnexinV, Pl-labeled flow cytometry, respectively. Results： The MTT assay revealed that MS-275 inhibited the growth of T24 cell lines not only in a concentration-dependent but also in a time-dependent manner, the growth inhibition ratio was （ 9.34± 3. 57） % when treated with 0.5μmol/L MS-275 for 72h, and （94.84±1.33） % with 8μmol/L for 120 h. The cloning efficiency was decreased from （55.67±6.51）% in negative control group to （2.33±0.58）% at concentration of 4μmol/L. Typical morphological change of apoptotic cells were observed by microscope. The apoptotic percentage at concentration of 1 μmol/L,2μmol/L and 4μmol/L were 24. 19% ,30.77% and 38.51% quantitativel measured by which were significantly higher than that of negative control group（ 2.49% ）. Conclusions： MS-275 can inhibit bladder cancer in vitro through indu cing apoptosis and mitotic death of cells. The results of this study suggest that histone deacetylase inhibitors are hopeful to become new effective drugs to treat bladder neoplasms.