人PC-3细胞系中前列腺癌干细胞的富集与鉴定  

Enrichment and Identification of Prostate Cancer Stem Cells in Human Prostate Cancer PC-3 Cell Line

作　　者：张波[1] 范新兰[2] 林天歆[1] 许可慰[1] 黄健[1] 

机构地区：[1]中山大学附属孙逸仙纪念医院泌尿外科,广东广州510012 [2]中山大学附属孙逸仙纪念医院医学研究中心,广东广州510012

出　　处：《中山大学学报（医学科学版）》2010年第6期878-883,共6页Journal of Sun Yat-Sen University:Medical Sciences

基　　金：国家自然科学基金(30500507;30672092);广东省自然科学基金重点项目(6104605);广东省自然科学基金(5001758;07117366);教育部博士点基金(20060558027)

摘　　要：【目的】从人前列腺癌PC-3细胞系中分离、富集并鉴定前列腺癌干细胞。【方法】采用无血清培养液(SFM)悬浮细胞聚球法体外培养PC-3细胞系,通过传代更新、诱导分化、交替在含血清/无血清培养液中培养等方法鉴定PC-3悬浮成球细胞具有强增殖性、自我更新和分化潜能等肿瘤干细胞特性。以PC-3悬浮成球细胞为实验组,常规在含血清培养液(SSM)中单层贴壁培养的PC-3细胞为对照组,采用流式细胞术检测两种细胞中CD44+CD133+细胞及侧群(SP)细胞比例。【结果】PC-3成球细胞可以在SFM中生存并形成悬浮细胞球,悬浮培养第5天时开始可以观察到典型的悬浮细胞球形成,第10天时细胞球形成效率约为(2.6±1.0)%,较第5天时明显增多(P<0.05)。PC-3悬浮成球细胞具有自我更新和分化能力,在体外可以连续、稳定传代,在SFM中滴加血清后可以分化,交替培养于SSM和SFM中可以实现与贴壁细胞之间的相互转化。PC-3悬浮成球细胞中CD44+CD133+细胞比率为13.94%,SP细胞含量为3.1%,均显著高于常规培养的PC-3贴壁细胞(分别为0.77%和0.0%,P<0.05)。【结论】PC-3悬浮成球细胞具有肿瘤干细胞特性,采用无血清悬浮细胞聚球培养法可以从PC-3细胞系中简便、高效地富集前列腺癌干细胞。【Objective】 To isolate,enrich and identify the prostate cancer stem cells in human prostate cancer PC-3 cell line. 【Methods】 PC-3 cells were suspension cultured in vitro with a serum-free medium （SFM） and reformed into floating spheres which were isolated. Then,the isolated floating PC-3 sphere-forming cells with the properties of cancer stem cells,such as strong proliferative,self-renewal and differentiation potentials,were identified by self-renewal assay,differentiation assay,and alternative cultivation assay with SFM and serum supplemented medium （SSM）. The suspension sphere-forming PC-3 cells were the experimental groups and the single-layer adherent PC-3 cells cultured with SSM conventionally were the control groups. The percentages of CD44＋ CD133＋ cells and side population （SP） cells in the two kinds of cells were detected with flow cytometry （FCM）. 【Results】 The PC-3 sphere-forming cells could survive in SFM and form floating cell spheres successfully. The typical floating cell spheres could be observed after 5 days and the proportion of cell spheres formed was （2.6 ± 1.0）% in day 10,which was significantly higher than that of day 5 （P 0.05）. The PC-3 floating sphere-forming cells possessed the potentials of self-renewal and differentiation and could propagate stably in vitro. Added FBS into the SFM,the sphere-forming cells could differentiate to adherent cells. When cultured alternately in SSM and SFM,the growth modes of the PC-3 sphere-forming cells could transform between adherent mode and sphere-forming mode alternately. The percentages of CD44＋ CD133＋ cells and SP cells in the PC-3 floating sphere-forming cells were higher significantly than that of the PC-3 adherent cells （13.94% vs. 0.77%,3.1%vs. 0.0% ,respectively,P 0.05）. 【Conclusion】 The PC-3 sphere-forming cells which were suspension cultured possess some properties of cancer stem cells. Using our suspension culture method with SFM,the prostate cancer stem cells in PC-3 cell line can be isola

关 键 词：PC-3 前列腺癌 悬浮培养 聚球法 肿瘤干细胞 富集 鉴定 

分 类 号：R737.25[医药卫生—肿瘤]