摘要
目的探讨SP600125对人宫颈癌HeLa细胞的增殖周期、凋亡以及侵袭的影响。方法采用CCK-8法检测不同时间点不同浓度的SP600125作用后HeLa细胞的增殖状态。确定20μmol/L的SP600125用于后续实验。利用平板克隆形成实验检测细胞增殖能力,DAPI染色观察细胞核形态,流式细胞仪检测细胞周期和凋亡,细胞划痕和Transwell法检测细胞迁移和侵袭能力,qRT-PCR和Western blot分别检测SP600125作用不同时间点后各组细胞的p53、Mad2L1和CDC20 mRNA和蛋白水平的变化。结果与对照组(0.1%DMSO)相比,10、20、30、40、50μmol/L SP600125作用24 h均能使细胞的增殖活性降低。与对照组相比,各SP600125处理组的细胞凋亡率明显增加,且G_(2)/M期细胞比例增加(P<0.001),而SP600125处理HeLa细胞24和48 h的G_(0)/G_(1)期比例减少(P<0.001),其细胞的克隆数、迁移和侵袭能力明显下降(P<0.001);qRT-PCR和Western blot结果显示Mad2L1 mRNA和蛋白表达明显下降(P<0.05),p53和CDC20 mRNA和蛋白则呈上升趋势(P<0.01)。结论SP600125可通过上调p53和CDC20以及下调Mad2L1的表达诱导宫颈癌HeLa细胞周期阻滞于G_(2)/M期并促进细胞凋亡来抑制细胞的增殖、迁移和侵袭。
Objective To investigate the effect of SP600125 on the proliferation,cell cycle,apoptosis and invasion of human cervical cancer HeLa cells.Methods CCK-8 method was used to detect the proliferation of HeLa cells treated with different concentrations of SP600125 at different time points.The 20μmol/L of SP600125 was determined for subsequent experiments.Cell proliferation ability was detected using plate clone formation assay;nuclear morphology was observed by DAPI staining;cell cycle and apoptosis were measured by flow cytometry;cell migration and invasion were detected by cell scratch and Transwell methods;the mRNA and protein levels of p53,Mad2L1 and CDC20 were measured by qRT-PCR and Western blot after SP600125 treatment at different time points.Results Compared with control group(0.1%DMSO),cells proliferative activity were reduced by 10,20,30,40 and 50μmol/L SP600125 treatment for 24h.Compared with control group,the rate of apoptosis was significantly increased in SP600125 treatment groups,and the cell proportion in G_(2)/M phase increased(P<0.001),while the cell proportion in G_(0)/G_(1) phases cells was reduced after SP600125 treatment for 24h and 48h(P<0.001),and the clonal number,migration and invasion ability of HeLa cells also decreased significantly(P<0.001).qRT-PCR and Western blot results showed a significant decrease in Mad2L1 mRNA and protein expression(P<0.05)and a significant increase in p53 and CDC20 mRNA and protein expression(P<0.01).Conclusion SP600125 can induce cell cycle arrest of cervical cancer HeLa cells in G_(2)/M phase by upregulating p53 and CDC20 and downregulating Mad2L1 expression,and promote cell apoptosis to inhibit cell proliferation,migration and invasion.
作者
莫艳秀
姚飞虹
刘峻彤
胡紫昂
李木兰
MO Yanxiu;YAO Feihong;LIU Juntong;HU Ziang;LI Mulan(Department of Histology and Embryology,School of Basic Medical Science,Xiangnan University,Chenzhou 423000,China)
出处
《肿瘤防治研究》
CAS
CSCD
2022年第4期304-313,共10页
Cancer Research on Prevention and Treatment
基金
湖南省教育厅资助科研项目(19A465)
湖南省大学生创新创业训练计划【2020】191(3665)
湘南学院2020高层次人才科研启动基金(20A20)。
