中文摘要：

目的利用无血清条件悬浮培养人大细胞肺癌H460系，从中分离到肺癌干细胞并进行其生物学特性研究。方法无血清条件培养法悬浮培养H460细胞形成悬浮细胞球，通过qPcR、Western blot、流式细胞术、平皿克隆形成等实验，比较H460细胞和H460细胞球中干性相关分子表达水平及细胞增殖能力强弱，并利用裸鼠移植瘤形成实验研究肺癌细胞干细胞球的体内生物学特性。结果利用无血清条件培养法成功从H460细胞中分离出悬浮生长的肿瘤干细胞样细胞，其增殖能力强于H460贴壁细胞（P〈0．05），干细胞相关转录因子Sox2、Oct4和Nanog表达在mRNA及蛋白水平均有增加（尤以Nanog提升水平明显（P〈0．01）），细胞球在裸鼠体内具有较强成瘤能力。结论无血清条件培养法可以有效富集到H460细胞系中的肺癌干细胞，Nanog可能与H460细胞系中CSLCs的干性特征维持相关。

英文摘要：

Objective To isolate lung cancer stem-like cells （LCSCs） from human large-cell lung cancer cell line NCI-H460 （H460） and explore their biological characteristics. Methods H460 cells were cultured in serum-free medium in the presence of specific growth factors. Quantitative PCR （qPCR）, flow cytometry and colony formation assay were performed to characterize the stemness of H460 spheres. Adherent H460 cells and H460 cell spheres were inoculated subcutaneously in nude mice and the tumor growth was assessed. Results The isolated LCSCs from H460 cells in serum-free medium grew as floating cell spheres and exhibited stronger proliferative activity than H460 cells. Compared with H460 cells, H460 cells spheres showed higher expressions of stem cell markers Sox2, Oct4, and especially Nanog, and possessed a stronger tumorgenicity in nude mice. Conclusion The serum-free culture system can effectively enrich lung cancer stem cells from human lung cancer stem cell line H460, and the high expression of Nanog may importantly contribute to the maintenance of cancer stem cell-like properties of the isolated LCSCs.