摘要目的 在无血清培养液悬浮培养条件下筛选人乳腺癌细胞系MCF-7、MDA-MB-231的干细胞相关亚群,并初步探讨其相关基因的表达差异.方法 采用有血清贴壁和无血清悬浮培养方法,扩增出贴壁细胞和悬浮微球体细胞两种肿瘤细胞.采用流式细胞术检测MCF-7、MDA-MB-231贴壁及悬浮微球体细胞中CD44+ CD24-、CD133+表型细胞比例的变化.反转录聚合酶链反应(RT-PCR)检测两组贴壁及悬浮微球体细胞中CD44、CD24、CD133、ALDH3A1、ABCG2、CXCR4基因的表达变化.结果 MCF-7微球体细胞的CD44+ CD24-/ low细胞比例高于其贴壁培养细胞(P＜0.05);CD133+细胞比例同其贴壁培养细胞差异无统计学意义(P＞0.05).MDA-MB-231微球体细胞的CD44+ CD24-/low细胞比例低于其贴壁培养细胞(P＜0.05);CD133+细胞比例高于其贴壁培养细胞(P＜0.05).MCF-7微球体细胞中CD44、ABCG2的表达较其贴壁细胞上调(均P＜0.05),CD24、CD133、ALDH3A1、CXCR4表达与其贴壁细胞相比,差异均无统计学意义(均P＞ 0.05).MDA-MB-231微球体细胞中CD24、CD133、ABCG2、ALDH3A1、CXCR4表达较其贴壁细胞上调(均P＜0.05),CD44表达下调(P＜0.05).MCF-7和MDA-MB-231微球体细胞中CD44、CD24、CD133、ALDH3A1表达差异有统计学意义(均P＜ 0.05).结论 乳腺癌MCF-7和MDA-MB-231细胞系微球体中乳腺癌干细胞相关基因的表达存在差异,提示不同分子亚型乳腺癌干细胞的来源可能不同.

abstractsObjective To enrich breast cancer stem cells of breast cancer cell lines MCF-7 and MDA-MB-231 through culturing mammospheres,and to detect the expression differences of gene of breast cancer stem cells makers.Methods The MCF-7 and MDA-MB-231 cell lines were cultured by serum and serum-free medium.The proportion of CD44+/CD24-and CD133+ cancer stem cells was measured in cells derived from mammosphere cells or monolayer culture cells by flow cytometry,and the expression of CD44,CD24,CD133,ALDH3A1,ABCG2 and CXCR4 mRNA were detected by RT-PCR.Results Flow cytometry analysis indicated that the CD44+/CD24-low proportion of the MCF-7 mammosphere cells was higher than its adherent culture cells (P ＜ 0.05),and CD133+ proportion had no difference between them (P ＞ 0.05).However,the CD44+/CD24-/low proportion of the MDA-MB-231 mammosphere cells was lower than its adherent culture cells (P ＜ 0.05),while CD133+ proportion was significantly higher than its adherent cultured cells (P ＜ 0.05).RT-PCR analysis suggested that the expression of CD44 and ABCG2 increased obviously in MCF-7 microspheres (P＜ 0.05),and the expression of CD24,CD133,ALDH3A1 and CXCR4 had no significant difference between the mammosphere cells and adherent culture cells (P ＞ 0.05).The CD24,CD133,ABCG2,ALDH3A1 and CXCR4 increased obviously in MDA-MB-231 microspheres.On the contrary,the CD44 decreased (P ＜ 0.05).The expression of CD44 and CD24,CD133 and ALDH3A1 had significant differences in the microspheres of MCF-7 and MDA-MB-231 cells (P ＜ 0.05).Conclusion The related cancer stem cells gene expression is different in the microspheres of human breast cancer cell line,which indicates that the different subtypes of breast cancer may be derived from different origins.