Suspension culture combined with chemotherapeutic agents for sorting of breast cancer stem cells.
Suspension culture combined with chemotherapeutic agents for sorting of breast cancer stem cells.
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DOI：
10.1186/1471-2407-8-135
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发表时间：
2008-05-14
期刊：
影响因子：
3.8
通讯作者：
Wu ZY
中科院分区：
文献类型：
--
作者：
Li HZ;Yi TB;Wu ZY
肿瘤干细胞（CSC）假说一直没有得到很好的证明，缺乏最令人信服的证据，关于一个单一的细胞能够产生肿瘤。CSCs数量的缺乏和分离纯化方法的不当已成为CSCs大发展的主要障碍。本研究采用悬浮培养结合抗癌方案作为筛选乳腺癌干细胞的策略。BrCSC可以在非贴壁悬浮培养中存活并高度富集，而化疗剂可以破坏大多数快速分裂的癌细胞并保留相对静止的BrCSC。在无血清培养基中培养TM 40 D鼠乳腺癌细胞。流式细胞术检测CD 44 + CD 24-的表达。将第10代细胞与不同峰值血浆浓度的抗癌剂紫杉醇和表阿霉素组合处理24小时，然后保持悬浮培养。流式细胞仪Annexin-V异硫氰酸荧光素（FITC）/碘化丙啶（PI）双染法检测细胞凋亡率。将所选细胞以不同量皮下注射到BALB/C小鼠中以观察肿瘤形成。悬浮培养第10代细胞中CD 44 + CD 24-的比例最高（约77%）。0.35PPC中的单个肿瘤细胞可在20只BALB/C小鼠中的3只中形成肿瘤。悬浮培养结合抗癌方案提供了分离、培养和纯化BrCSC的有效手段。
Cancer stem cell (CSC) hypothesis has not been well demonstrated by the lack of the most convincing evidence concerning a single cell capable of giving rise to a tumor. The scarcity in quantity and improper approaches for isolation and purification of CSCs have become the major obstacles for great development in CSCs. Here we adopted suspension culture combined with anticancer regimens as a strategy for screening breast cancer stem cells (BrCSCs). BrCSCs could survive and be highly enriched in non-adherent suspension culture while chemotherapeutic agents could destroy most rapidly dividing cancer cells and spare relatively quiescent BrCSCs. TM40D murine breast cancer cells were cultured in serum-free medium. The expression of CD44+CD24- was measured by flow cytometry. Cells of passage 10 were treated in combination with anticancer agents pacilitaxel and epirubicin at different peak plasma concentrations for 24 hours, and then maintained under suspension culture. The rate of apoptosis was examined by flow cytometry with Annexin-V fluorescein isothiocyanate (FITC)/propidium iodide (PI) double staining method. Selected cells in different amounts were injected subcutaneously into BALB/C mice to observe tumor formation. Cells of passage 10 in suspension culture had the highest percentage of CD44+CD24- (about 77 percent). A single tumor cell in 0.35 PPC could generate tumors in 3 of 20 BALB/C mice. Suspension culture combined with anticancer regimens provides an effective means of isolating, culturing and purifying BrCSCs.
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