Puma抑制卵巢癌细胞生长及其机制研究

Function and significance of Puma inhibiting ovarian carcinoma growth through inducing cell apoptosis

目的探讨Puma对于卵巢癌细胞生长及体内成瘤的影响,了解其行使功能的分子机制。方法采用腺病毒载体实现Puma在卵巢癌细胞中的过表达,并通过CCK-8检测细胞增殖情况,通过Annexin-V染色测定细胞凋亡情况,通过小鼠荷瘤实验确定卵巢癌细胞体内成瘤能力。并进一步通过Western blot的方法检测了细胞凋亡相关基因Caspase-9和Bax蛋白的表达及定位情况。结果成功构建hTERT基因启动子介导的Puma基因腺病毒表达载体,并实现Puma基因在卵巢癌细胞中的过表达,过表达Puma后,卵巢癌细胞增殖被明显抑制(P<0.05),细胞凋亡增加,从对照组细胞的7.2%增加为过表达组的29%(P<0.05),且在小鼠体内的成瘤能力下降,所产生肿瘤组织的平均质量由0.53 g降至0.22 g(P<0.05)。结论 Puma通过激活细胞内Caspase-9的活性、促使Bax从胞质转移至线粒体、促进细胞凋亡信号传导诱导细胞凋亡,从而抑制卵巢癌细胞增殖及体内成瘤能力。

Objective To investigate the function and mechanism of p53 up-regulated modulator of apoptosis（Puma） in inhibiting ovarian carcinoma cell growth in vitro and in vivo.Methods Overexpression of Puma in ovarian carcinoma cell line COC1 was achieved by adenovirus vectors.Cell proliferation was detected using CCK-8 kit,meanwhile the percentage of apoptotic cells was identified by Annexin V staining.The mouse xenograft model was established to detect the inhibition on ovarian carcinoma cell growth in vivo.The expression of caspase 9 and Bax was detected by Western blotting.Results Puma was successfully over-expressed in COC1 cells using the established adenovirus expression vector.Over-expression of Puma inhibited the cell proliferation of COC1 cells evidently（P 0.05） and increased the percentage of apoptotic cells to 29% as compared to 7.2% in the control（P 0.05）.Meanwhile,Puma inhibited the tumorigenicity of ovarian carcinoma cells in vivo.The average weight of xenografts was 0.22 g in the Puma overexpression group,significantly lower than 0.53 g in the control group（P 0.05）.Conclusion Puma plays a tumor-suppressor role in ovarian carcinoma.Puma inhibits cell proliferation,induces cell apoptosis partially through activating caspase 9 and inducing Bax translocation to mitochondria and also inhibits the tumorigenicity in vivo.