基底样乳腺癌和管腔A乳腺癌干祖细胞的生物学行为比较

Biological Behaviors of Cancer Stem/Progenitor Cells Derived From Basal-Like and Luminal-A Breast Cancer

目的:从人基底样和管腔A乳腺癌分离肿瘤干/祖细胞,并比较其增殖、自我更新和分化能力等生物学特点。方法:应用乳腺微球无血清悬浮培养法富集乳腺癌干/祖细胞,并通过细胞生长曲线测定和连续克隆形成实验检测了干/祖细胞在体外的增殖能力和自我更新能力,建立裸鼠移植瘤检测在体内的成瘤情况,流式细胞术检测CD44和CD24的表达水平。结果:基底样和管腔A乳腺癌均能在无血清培养基中形成乳腺微球,基底样癌的肿瘤干细胞含量较高,并形成细胞数量多、直径大的微球,球内细胞在体内、外有更强的扩增能力。基底样癌传代过程中,细胞容易贴壁分化,传代多不超过15代;而管腔A癌,虽然丧失克隆形成能力的悬浮单细胞明显增多,但均可传代20次以上;两者CD44和CD24的表达也有明显差异。结论:基底样乳腺癌和管腔A乳腺癌的干/祖细胞表现出差别明显的生物学行为,乳腺癌的肿瘤干细胞可能是异质性的。

Objective： To isolate breast cancer stem/progenitor cells from human basal-like and luminal A breast cancer and to explore their biological behaviors. Methods： Human breast cancer stem/progenitor cells were endched in suspension cultures as nonadherent mammospheres. Cell growth curve determination and serial sphere formation assay were used to determine proliferation and self-renewing ability of breast cancer stem cells in vitro. Cell proliferation in vivo was also inves- tigated by transplanting tumors in nude mice. The expression of CD44 and CD24 in mammosphere-derived cells was evaluated by flow cytometry. Results： Cells from both basal-like and luminal A breast cancer can grow in serum-free conditions as mammosphere cultures. Basal-like tumors had a higher percentage （5.1%±1.08%） of stem cells than luminal A tumors （2.02%±0.30%） and yielded mammospheres with a larger diameter which contained more cells. Mammosphere cells derived from basal-like tumors extensively proliferated in vitro and in vivo, more so than their luminal A counterparts. Mammospheres from both basal-like tumors and luminal A tumors were capable of serial passage. Basal-like tumors showed a pro- gressive decrease in mammosphere forming efficiency, increased tendency to differentiate and a failure to passage beyond 15 generations with most cells adhering and undergoing terminal differentiation. While mammospheres derived from luminal A tumors could serially passage over 20 generations in vitro, increasing numbers of single cells lost their sphere-forming potential. CD44^＋/CD24^- tumor cells were detected in all basal-like tumors, while they were either absent or only a small amount appeared in luminal A breast cancer. Conclusion： Breast cancer stem/progenitor cells of the basal-like and luminal A molecular subtype exhibit different stem cell properties of self-renewal, indefinite proliferation and multi-lineage differentiation. It appears they have transformed separately from breast epithelial cells.