大细胞肺癌H460细胞系肿瘤干细胞样细胞生物学特性研究.PDFVIP

基础研究 大细胞肺癌大细胞肺癌HH460460 细胞系肿瘤干细胞样细胞生物学特性研究细胞系肿瘤干细胞样细胞生物学特性研究 刘 攀，周向东 第三军医大学西南医院呼吸内科，重庆 400038 提要：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（P0.05），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。 关键词：大细胞肺癌；肿瘤干细胞；无血清条件培养；Nanog Biological behaviors of lung cancer stem-like cells from human large-cell lung cancer cell line H460 LIU Pan, ZHOU Xiangdong Department of Respiratory Disease, Southwest Hospital, Third Military Medical University, Chongqing 400038, China Abstract: 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 mainten