Preliminary study on cancer stem-like cells in human breast cancer cell lines
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摘要:
目的：检测不同乳腺癌细胞系中肿瘤干细胞相关亚群含量，并探讨使乳腺癌细胞系MCF-7中肿瘤干细胞相关亚群富集的方法。方法：利用流式细胞仪检测3种乳腺癌细胞系MCF-7、MDA-MB-231、MDA-MB-435s中SP细胞（side population cells）比例及CD44+CD24-／low细胞比例。用Percoll不连续密度梯度法分离MCF-7细胞亚群，利用流式细胞仪筛选出SP细胞富集的亚群。用添加生长因子的无血清培养液培养MCF-7，检测其中CD44+CD24-／low细胞的比例。结果：MCF-7、MDA-MB-435s中SP细胞比例分别为3.61%和2.72%，MDA-MB-231中未检测到SP细胞。3种细胞系中CD44+CD24-／low细胞比例分别为0.87%、0.59%和94.04%。MCF-7经Percoll分选，D亚群SP细胞富集，比例高达25.23%。MCF-7中CD44+CD24-／low细胞比例随着无血清培养时间的增加而增加，3周时达29.35%。结论：SP检测与CD44+CD24-／low细胞检测是目前乳腺癌干细胞检测的两种常用方法，但两者在同一乳腺癌细胞系中的检测结果并不一致，两种方法均有一定的局限性。另外，Percoll分选或无血清培养均能使MCF-7中肿瘤干细胞相关亚群（SP细胞或CD44+CD24-／low细胞）富集。
Abstract:
Objective：To investigate the content of cancer stem-like cells in different human breast cancer cell lines and to explore the methods for enriching cancer stem-like cells in MCF-7. Methods：The percentage of SP cells and CD44+CD24-／low cells in human breast cancer cell lines MCF-7， MDA-MB-231 and MDA-MB-435s were detected with flow cytometry. MCF-7 was separated by Percoll gradient centrifugation into five subpopulations. In order to screen which subpopulation contained the highest content of SP cells， each subpopulation was detected with flow cytometry. Then MCF-7 was cultured in serum-free DMEM／F12 containing EGF， bFGF， BSA and insulin. The percentage of CD44+CD24－／low cells was detected with flow cytometry after 1，2，3weeks，respectively. Results：The percentage of SP cells was 3.61% in MCF-7 and 2.72% in MDA-MB-435s. There was no SP cells detected in MDA-MB-231. The percentage of CD44+CD24-／low cells was 0.87% in MCF-7， 0.59% in MDA-MB-231 and 94.04% in MDA-MB-435s. After the separation of Percoll gradient centrifugation， subpopulation D of MCF-7 was rich in SP cells，and reached up to 25.23%. The cancer stem-like cells in MCF-7 grew as clonal nonadherent spherical clusters. CD44+CD24-／low cells in MCF-7 increased after culture in serum-free medium. The percentage of CD44+CD24-／low cancer stem-like cells reached up to 29.23% after 3 weeks culture in serum-free medium. Conclusion：SP cells detection and CD44+CD24-／low cells detection are two commonly used methods in detecting breast cancer stem cells. But the results of two methods often disagree. Each method has its own limitation. The cancer stem-like cells in MCF-7 can enrich after separation by Percoll gradient centrifugation or culture in serum-free medium.
