摘要
目的构建OSKM诱导的过表达c-Jun肝癌细胞(C3A-c-Jun)重编程细胞系,探讨外源性c-Jun对肝癌细胞重编程的影响。方法通过C3A细胞稳定过表达c-Jun后进行OSKM诱导重编程,通过碱性磷酸酶染色,Real-time PCR,免疫印迹法,免疫荧光等技术对细胞进行鉴定,建立C3A-c-Jun诱导肿瘤干细胞系C3A-c-Jun-i CSCs。结果 C3A-c-Jun-i CSCs克隆呈圆顶状,边缘圆钝,克隆内细胞小且排列紧密,多层分布,碱性磷酸酶染色阳性,mRNA和蛋白水平均可表达多潜能性标记物OCT4、SOX2;C3A-c-Jun-i CSCs组外源性和内源性Sox2的表达量均明显高于C3A-i CSCs对照组;在重编程过程中c-Jun持续表达。结论 OSKM诱导C3A、C3A-c-Jun肝癌细胞重编程,分别建立C3A-i CSCs和C3A-c-Jun-i CSCs细胞系,发现外源性c-Jun通过上调Sox2基因的表达,启动下游一系列反应,对肝癌细胞重编程过程起促进作用。
Objective To establish an OSKM induced C3A-c-Jun of liver cancer reprogrammed cell line, and to explore the effects of exogenous c-Jun on liver cancer cell reprogramming. Methods OSKM was utilized to induce C3A-c- Jun to reprogram. Alkaline phosphatase staining, Real-time fluorescent quantitative polymerase chain reaction（ Real-time PCR）, Western blotting, and immunofluorescence technology were used to identify cells, and establish the C3A-c-Jun- iCSCs. Results C3A-c-Jun-iCSCs cells formed dome-shaped clones, in which ceils were small, closely packed and multi- layered. Alkaline phosphatase staining was positive. They expressed pluripotent markers Oct4, Sox2, Nanog at the mRNA level and express OCT4, and SOX2 at the protein level. C3A-c-Jun-iCSCs had obviously higher gene expression of Sox2 compared with C3A-iCSCs, whatever in exogenous expression or endogenous expression. C-Jun continually expressed in the process of reprogramming. Conclusion OSKM induces C3A/C3A-c-Jun of liver cancer reprogramming and built the C3A- i-CSCs/C3A-c-J-un-iCSCs. Exogenous c-Jun has a promoting effect on liver cancer cell reprogramming process by upregulating Sox2, which then can activate downloaded genes.
作者
苏铭
韩烁
张明智
李睿智
魏示若
曾沃坦
刘进稳
沈丽
SU Ming HAN Shuo ZHANG Ming-zhi LI Rui-zhi WEI Shi-ruo ZENG Wo-tan LIU Jin-wen SHEN Li(Department of Cell Biology, Peking University Health Science Center, Beijing 100191, China Beijing Oriental Yamei Gene Science and Technology Research Institute Co. LTD, Beijing, 100078, China)
出处
《解剖学报》
CAS
CSCD
北大核心
2017年第3期296-302,共7页
Acta Anatomica Sinica
基金
国家自然科学基金(81572313)
广东省省级科技计划项目(2015B020229002)
