高表达Twist基因促进人乳腺癌细胞MCF-7干细胞样细胞富集和迁移.DOCVIP

高表达Twist基因促进人乳腺癌细胞MCF-7干细胞样细胞富集和迁移 周明莉，杨佳佳，唐石伏，杨 丽，张海龙，杜燕娥，文思阳，柳满然（400016重庆，重庆医科大学检验医学院，临床检验诊断学教育部重点实验室） 【摘要】目的 在人乳腺癌细胞系MCF-7中高表达Twist基因，研究其对MCF-7干细胞特性的细胞的影响。方法 构建重组质粒pBABE-puro-myc-Twist，HEK293T细胞包装逆转录病毒，并感染MCF-7细胞，通过筛选获得成功转入Twist的MCF-7/Twist细胞和MCF-7/Vector对照细胞，再通过长程无血清悬浮培养方法富集具有干细胞特性的乳腺细胞小球(Mammosphere)，并通过Western blot法检测myc-Twist及Twist诱导后富集的Mammosphere相关CSC蛋白标记的表达，Transwell?法检测细胞的迁移能力。结果 成功构建逆转录病毒质粒pBABE-puro-myc-Twist；Twist基因在靶细胞内稳定表达；长程无血清悬浮培养富集出具有干细胞特性的Mammosphere； MCF-7/Twist细胞肿瘤干细胞样细胞富集能力增强(P0.05)； myc-Twist、Twist及SOX2、OCT4蛋白在MCF-7/Twist Mammosphere细胞中表达上调；MCF-7/Twist Mammosphere细胞迁移能力增加(P0.05)。结论 Twist促进MCF-7肿瘤干细胞样细胞富集能力增强，并促进其迁移。 【关键词】Twist；乳腺细胞小球Mammosphere；人乳腺癌细胞MCF-7 【中图分类号】Q782，R737.9， R329.2+8，Q291 【文献标志码】A Overexpression Twist gene in MCF-7 human breast cancer cells and its promotion in CSCs enrichment and cell migration ability. Zhou Mingli，Yang Jiajia，Tang Sshifu，YangLi，Zhang Hailong，Du Yane，Weng Siyang， Liu Manran（College of Laboratory Medicine, Key Laboratory of Laboratory Medical Diagnostics, Ministry of Education, Chongqing Medical University, Chongqing 400016, China） [Abstract] Objective To overexpress Twist gene in MCF-7 human breast cancer cells and investigate its effect on cancer stem-like cell in MCF-7. Methods Construct a recombinant plasmid pBABE-puro-myc-Twist and package of retrovirus in HEK293T cells. MCF-7 human breast cancer cells were infected with the retrovirus carrying Twist gene or the controls and MCF-7/Twist cells and MCF-7/Vector control cells were selected. Mammospheres with characteristics of stem-like cells were collected by long term serum-free suspension cultivation. The expressions of myc-Twist and cancer stem-like cells (CSCs) marker proteins induced by Twist in MCF-7/Twist Mammosphere and MCF-7/Vector Mammosphere were determined by qRT-PCR and Western Blotting. Cell migration abilities were analyzed by Transwell assay. Results The recombinant plasmids (pBABE-puro-myc-Twist) were identified. The Twist gene was eff