摘要
目的研究新型Nutlins类似物NL-608在体外诱导乳腺癌MCF-7细胞凋亡的作用，并初步探讨其可能的凋亡通路。方法采用四甲基偶氮唑蓝（MTT）法检测NL-608化合物对MCF-7细胞增殖的影响；采用流式细胞仪（FCM）检测NL-608诱导MCF-7细胞凋亡情况；采用Westernblot方法检测多聚ADP-核糖聚合酶（PARP）、半胱天冬酶（pro-caspase3、pro-caspase8、pro-caspase9）以及Fas和Fas配体（FasL）的变化，并利用活力检测试剂盒检测caspase3、caspase8、caspase9的活力。结果NL-608对MCF-7细胞的增殖具有抑制作用，并呈浓度依赖性，100、33、11、3．7、1．2、0．4、0．14Ixmol／LNL-608作用后，MCF-7细胞的存活率分别为（6．70±1．27）％、（10．90±5．08）％、（24．31±3．77）％、（45．144±5．14）％、（79．16±10．01）％、（97．00±2．31）％和（99．90±0．01）％。随着NL-608浓度的增加，PARP被切割，pro-caspase3和pro—caspase8的Westernblot条带变淡，但pro-caspase9的条带无变化。活力测定结果进一步显示，easpase3和caspase8被激活，其相对活力与对照组相比分别为（3．76±0．35）倍和（2．71±0．27）倍；而caspase9的相对活力为（1．03±0．08）倍，即对easpase9无明显影响。Westernblot结果显示，Fas和FasL的表达量均随NL-608浓度的增加而增加。结论NL-608化合物能够有效地在体外诱导MCF-7细胞凋亡，且可能依赖于easpase3和caspase8相关的死亡受体凋亡途径，为进一步研发乳腺癌治疗药物奠定了实验基础。
Objective To observe the effect of NL-608 （a nutlin analog） on apoptosis induction in human breast cancer MCF-7 cells in vitro, and investigate the relevant molecular mechanism. Methods The effect of NL-608 on proliferation of MCF-7 cells was determined by MTI＂ assay. The apoptosis in MCF-7 cells was determined by flow cytometry with annexin V-FITC and PI. The activity of caspase 3, caspase 8 and caspase 9 was determined with caspase activity assay kit and Western blot, and the proteins of Fas and FasL were determined by Western blot. Results NL-608 showed a dose-dependent inhibitory effect on the proliferation of MCF-7 cells. It induced apoptosis in MCF-7 cells in a dose-dependent manner. The activity of caspase 3 and easpase 8 in MCF-7 cells was increased with the increasing concentration of NL-608, but caspase 9 had no changes. The proteins of Fas and FasL were increased in a dose-dependent manner. Conclusion NL-608 induces apoptosis in MCF-7 cells in vitro through inducing caspase 3 activity and death receotor-mediated signal oathwav.
出处
《中华肿瘤杂志》
CAS
CSCD
北大核心
2012年第2期100-103,共4页
Chinese Journal of Oncology
基金
吉林省科委重点项目（20080932）
长春市科技计划项目（长科合2006224）
