文章摘要

YAP对卵巢癌干细胞增殖的影响

作者: 1许 静, 1郭 哲, 1王 秋宇, 1梁 殿迅
1 南阳市中心医院妇产科,河南 南阳 473000
通讯: 许 静 Email: 3111685775@qq.com
DOI: 10.3978/j.issn.2095-6959.2017.12.004

摘要

目的:探讨YAP基因在卵巢癌干细胞中的表达和作用。方法:取对数期人卵巢细胞株OVCAR-3,胰酶消化后制备单细胞悬液,利用流式细胞术鉴定卵巢癌干细胞。将YAP 5个磷酸化位点突变(YAP-5SA)及C末端缺失(YAP-△C)载体构建在慢病毒载体上,并且将YAP-5SA,YAP-△C载体的病毒转染卵巢癌干细胞,筛选稳定转染的细胞进行MTT细胞增殖实验、平板克隆实验。取卵巢癌干细胞,YAP-5SA和YAP-△C稳定转染的卵巢癌干细胞,接种在15只裸鼠中,观察4周,每周记录1次瘤体体积。结果:分离制备的细胞接种在DMEM培养基中培养24 h后细胞数量开始增多,生长5 d时不同细胞长满瓶底,细胞成干细胞球形,流式细胞仪测定该细胞CD44阳性率为99.8%,CD117为0.76%。YAP在卵巢癌干细胞高表达且YAP-5SA促进卵巢癌干细胞增殖和平板细胞克隆形成,YAP-△C抑制卵巢癌干细胞增殖和平板细胞克隆形成。YAP-△C稳转卵巢癌干细胞促进瘤体生长,YAP-△C抑制卵巢癌干细胞瘤体生长。结论:YAP-5SA促进卵巢干细胞增殖,参与卵巢癌的发生、发展。
关键词: 卵巢干细胞;增殖;克隆形成;YAP信号;人卵巢细胞株OVCAR-3

Effect of YAP gene on proliferation of ovarian cancer stem cells

Authors: 1XU Jing, 1GUO Zhe, 1WANG Qiuyu, 1LIANG Dianxun
1 Department of Obstetrics and Gynecology, Nanyang City Center Hospital, Nanyang Henan 473000, China

CorrespondingAuthor:XU Jing
Email: 3111685775@qq.com

Abstract

Objective: To investigate the role of YAP in ovarian cancer stem cells and its influence on biologic function. Methods: A single cell suspension was prepared after digestion of pancreatic enzymes with OVCAR-3 cells. And then, the ovarian cancer stem cells were identified by flow cytometry. The YAP five phosphorylation site mutation (YAP-5SA) and C terminal deletion (YAP-△C) in the construction of vector lentiviral packaging vector, and the virus core particles transfected with YAP-5SA, YAP-△C vector in ovarian cancer stem cells, then the cell proliferation and stability were examined by MTT assay and colony-forming assay. The ovarian cancer stem cells, YAP-5SA and YAP-△C transfected ovarian cancer stem cells were inoculated in 15 the nude mice, the volume of tumors were measurand for 4 weeks, once a week. Results: After the ovarian cancer stem cells inoculated in DMEM culture medium for 24 h, the number of cells were increased, and 5 days the cells presenting stem cell globules and covering the bottom of the bottle. And the cells had 99.8% CD44 and 0.76% CD117 examined by flow cytometry. YAP level was higher in ovarian cancer stem cells than ovarian cancer cells. And YAP-5SA promoted stem cell proliferation and tablet cell cloning in ovarian cancer stem cells, however, YAP-△C showed the opposite phenomenon. YAP-5SA transfected ovarian cancer stem cells promoted the growth of the tumor, however, YAP-△C transfected ovarian cancer stem cells suppressed the growth of the tumor. Conclusion: These results suggest that YAP contribute to the proliferation of ovarian stem cells and participate in the development and progression of ovarian cancer.
Keywords: ovarian cancer stem cells; proliferation; colony-forming; YAP signaling; human ovarian cell line OVCAR-3

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