目的 通过乳腺癌细胞系及其干细胞的培养,化疗药干预和流式细胞仪筛选鉴定,探讨不同乳腺癌细胞系中CD44+CD24-/low细胞比例,及富集乳腺癌干细胞相关亚群的方法.方法 通过细胞培养乳腺癌细胞系MCF-7、MDA-MB-231,观察生长曲线,比较化疗药物干预下生长情况;利用流式细胞仪检测两种乳腺癌细胞系中CD44+CD24-/low细胞比例;无血清悬浮培养,化疗药(多西紫杉醇、表阿霉素)干预这两种乳腺癌细胞系,观察其是否形成细胞球.结果 (1)MDA-MB-231细胞系倍增时间短,生长速率高于MCF-7细胞系;(2)MCF-7细胞系中可能存在较大比例肿瘤干细胞,其对化疗抵制,能自我更新;(3)化疗敏感性用两独立样本t检验,MCF-7细胞,差异没有统计学意义;MDA-MB-231细胞,差异有统计学意义,提示MDA-MB-231细胞系对该方案化疗较敏感;(4)无血清悬浮培养,MDA-MB-231细胞系未发现明显细胞球;MCF-7细胞系初次无血清培养约6天出现细胞球.加入化疗药筛选后两种细胞,见大部分肿瘤细胞逐渐死亡,未发现明显细胞球;(5)流式细胞仪检测,MCF-7、MDA-MB-231两种细胞系中主要是CD44+CD24+亚群和CD44-CD24+亚群,CD44+CD24-/low细胞比例分别2.07%和0.20%.结论 (1)MDA-MB-231细胞系增值较快,恶性度相对较高,其对TA联合化疗药物较敏感;MCF-7细胞系中可能存在少量肿瘤干细胞,对化疗抵制,能自我更新;(2)无血清培养液培养MCF-7细胞系能形成悬浮细胞球;流式细胞仪检测两种细胞系中CD44+CD24-/low细胞比例小;(3)CD44+CD24-/low表型可能不是乳腺癌干细胞唯一特异性的表面标志.%Objective Under e'lilturing different human breast cancer cell lines and the related content of cancer stem -like cells,we disease CD44 + CD24~'W cell proportion in breast cancer cell lines and related subsets enrichment method of breast cancer stem - like ceils through effective intervention of the commonly chem otherapy and flow eytomctrie analysis. Methods Through the ceil culture of breast cancel ceil linc"MCF - 7, MDA - MB -231" ,wc observed the growth curve and compared grow under conditions with chemotherapy drugs intervention, we detected the expression proportion of CD44 * CD24""? ceil in the two breast cancer ceil lines by flow eytomctry. Under suspension culturing the above ceil lines in scrum -free medium,we observed clonal non adherent spherical clusters by combining writh or without chemotherapy scheme of TA scheme( doeetaxel + table doxorubicin ). Results 1. The multiplied time: elf MDA - MB - 231 ceils weis shorten, anel its greiwth rate; was high er than MCF - 7 ceils. 2. The MCF - 7 cells might exist the larger proportion of cancer stem - like ceils with chemeithe:rapy re:sistance and self - update .3. Chemethe:rapy sensitivity with two ineiepeneie:nt sample t - test,the result in MCF - 7 ceils suggested no statistical significanco( P > 0. 05 ). The result in MDA - MB - 231 cells showed obvious positive significancc( P <0. 05 ). MDA - MB -231 cells were more: sensitive: to chemotherapy from them. 4. Under sonim -free suspension cultures of two hreast ceil lines, MDA-MB-231 ceils gelt no clear cell halls. MCF -7 ceils wore found ceil halls at the: first 6 day. With chemotherapy,the majority of the two tumor cells gradually died,and there were no living ceil halls. 5. Measured the: content of CD44+ CD24 - low eytometry cells by flow eytomctry, the two ahem cell lines mainly contain CD44 + CD24 + snhscts( 74. 2% ,95. 7% )and CD44-CD24+ snhscts( 23.3% ,3-95% ).Tk: CD44 + CD24-low proportion of MCF-7 and MDA-MB-231 was 2.07% and 0.20%,rcspcetircly. Conclusion 1. The MDA - MB -231 calls line which growed faster had higher malignant degree and showed more sensitive than MCF -7 cells line to TA chemotherapy scheme. 2. Under serum -free suspension cultures,MCF -7 cells grow clonal monadherent spherical clusters,and the two lines had small proportion of CD44 + CD24 -low cell with flow eytomctrie detection. 3. The CD44+ CD24-low phenotype may not he the only specific surface markers of breast cancer stem cells.
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