Daxx在卵巢癌细胞中的定位及对凋亡的影响

Location of Daxx in ovarian cancer cells and the impact on cell apoptosis

目的探索Daxx在卵巢癌细胞的定位及对凋亡的影响。方法免疫荧光检测Daxx在卵巢癌细胞C13k的定位,以及化疗药物顺铂(DDP)作用后的蛋白分布。利用细胞核及细胞浆蛋白提取试剂盒,分别提取DDP处理前后的核蛋白及浆蛋白,Western-blot检测Daxx在胞核和胞浆中的变化。裸鼠体内实验进一步验证化疗药DDP对Daxx蛋白分布的影响。结果免疫荧光结果提示,在化疗药DDP处理前,Daxx定位在卵巢癌细胞C13k的胞核中,胞浆内肉眼几乎观察不到Daxx;化疗药处理后,Daxx由胞核部分转移到胞浆;Western-blot结果表明,在化疗药DDP处理前,在卵巢癌细胞C13K中Daxx多分布在胞核,胞浆中表达量较少;化疗药作用后,Daxx在胞核的分布明显下降,胞浆中的分布明显升高;并且伴有Fas相关蛋白FADD表达量的升高;裸鼠肿瘤免疫组化显色提示,用DDP处理过的小鼠肿瘤切片,胞浆中Daxx表达量明显增加,且伴有凋亡指示蛋白Cleaved-parp明显升高。结论在卵巢癌细胞C13K中,化疗药DDP可使Daxx由胞核转入胞浆,并参与Fas凋亡通路的启动。由此表明Daxx在卵巢癌细胞C13K中有促进细胞凋亡的作用,为卵巢癌的治疗提供一个新的方向。

Objective To explore the location of death domain associated protein （Daxx） in ovarian cancer cells and the impact on cell apoptosis, Methods Immunofluorescence was used to detect the location of Daxx protein in ovarian cancer C13k cells before and after DDP treatment. Cell nucleus and cell plasma protein extraction kits were used to extract nucleoprotein and plasma protein before and after DDP treatment. Western blot was used to detect the changes of Daxx in cell nucleus and cell plasma. Nude mice in vivo experiment was performed to further confirm the effect of DDP on location of Daxx protein. Results Immunofluorescence results showed that Daxx located in cell nucleus of ovarian cancer C13k cells before DDP treatment, Daxx was rarely observed in cytoplasm by naked eye; after DDP treatment, Daxx transferred from cell nucleus to cytoplasm patially. Western blot results showed that Daxx mainly located in cell nucleus of ovarian cancer C13k cells before DDP treatment, a little Daxx was observed in cytoplasm; after DDP treatment, location of Daxx in cell nucleus decreased significantly, while location of Daxx in cytoplasm increased significantly, at the same time, the expression of Fas-related protein FADD increased. Immunohistochemical staining of nude mice showed that the expression of Daxx in cytoplasm increased significantly in tumor sections of mice, the expression of apoptosis indicator Cleaved-parp increased significantly at the same time. Conclusion In ovarian cancer C13k cells, DDP can transfer Daxx from cell nucleus to cytoplasm, which is involved in start of Fas apoptosis pathway. The study indicates the promoting effect of Daxx on apoptosis of ovarian cancer C13k cells, which provide a new direction for treatment of ovarian cancer.