人肺腺癌细胞株SPC-A1诱导球体形成及肺癌干细胞致瘤能力评估  

Spheroid formation induced by human lung adenocarcinoma cell line SPC-A1 and the tumorigenic ability of lung cancer stem cells

作　　者：郝光军[1] 刘青[1] 王娟[2] 马彦娥[1] 丁延慧[1] 

机构地区：[1]榆林市第一医院,陕西省榆林市719000 [2]西安交通大学第一附院医院,陕西省西安市710061

出　　处：《中国组织工程研究》2015年第23期3681-3685,共5页Chinese Journal of Tissue Engineering Research

摘　　要：背景:目前关于肺癌干细胞是否可以形成球体及其致瘤能力尚缺乏明确的定论。目的:观察人肺腺癌细胞株SPC-A1诱导球体形成及肺癌干细胞的致瘤能力。方法:利用无血清-DF12培养液培养增殖期肺癌细胞株SPC-A1,加入重组人胰岛素样生长因子1和重组人表皮生长因子及重组人成纤维细胞生长因子10诱导球体形成,获得球体细胞沉淀物,进行免疫荧光检测和PCR扩增,了解干细胞相关标志的表达情况。将肺球体细胞植入NOD-SCID免疫缺陷小鼠皮下,观察肿瘤生长情况,并利用活体荧光成像仪进行摄片。结果与结论:培养肺腺癌细胞SPC-A1 5-10 d获得肺球体。RT-PCR检测发现,肺球体细胞表达肺癌干细胞及细支气管肺泡干细胞标志物,CD24、CD221、CCSP和SP-C;另外,肺球体细胞与纯化的CD24+、CD221+肺癌干细胞同样表达肺干细胞的主干基因TTF-1、胚胎干细胞主干基因OCT4、Nanog和Bmi-1肺干细胞的主干基因TTF-1。荧光检测显示,80%以上的肺球体细胞中表达CCSP和OCT4;SPC-A1细胞具有肺泡Ⅱ型细胞特征,并表达SP-C蛋白,但仅约5%的细胞表达CCSP和OCT4。肺球体细胞皮下植入50 d,活体荧光成像可清晰显示小鼠体内肿瘤直径达1 cm。表明人肺腺癌细胞株SPC-A1可诱导球体形成,球体中富含肺癌干细胞并具有致瘤能力。BACKGROUND： There is no clear conclusion on whether the lung cancer stem cells can induce to spheroid formation and have tumorigenicity.OBJECTIVE： To observe the spheroid formation induced by human lung adenocarcinoma cell line SPC-A1 and the tumorigenic ability of lung cancer stem cells.METHODS： SPC-A1 at proliferating phase was cultured in serum-free DF12 culture medium, and then recombinant human insulin-like growth factor-1, recombinant human epidermal growth factor, and recombinant human fibroblast growth factor-10 were added to induce spheroid cells. Immune fluorescence detection and PCR amplification were done to understand the expression of stem cell associated markers. NOD-SCID immunodeficient mice were subcutaneously implanted with lung spheroid cells to observe the tumor growth. In vivo fluorescence imager was used for radiography.RESULTS AND CONCLUSION： After 5-10 days, lung spheroid cells were harvested. RT-PCR results showed that lung spheroid cells were positive for CD24, CD221, CCSP and SP-C. In addition, the lung spheroid cells and purified CD24＋, CD221＋ lung cancer stem cells were both positive for TTF-1 of lung stem cells, OCT4 and Nanog of embryonic stem cells and TTF-1 of Bmi-1 lung stem cells. The fluorescence detection showed that over 80%lung spheroid cells expressed CCSP and OCT4; SPC-A1 cells had the characteristics of alveolar type II cells, and also expressed SP-C protein, but only about 5% of the cells expressed CCSP and OCT4. At 50 days after subcutaneous implantation of lung spheroid cells, in vivo fluorescence imaging showed that the diameter of tumor in mice was 1 cm, indicating human lung adenocarcinoma cell line SPC-A1 can induce the spheroid formation, and lung cancer stem cells rich in the cell spheres have the tumorigenic ability.

关 键 词：干细胞 肿瘤干细胞 肺肿瘤 肺腺癌 肺癌 球体 细胞培养 

分 类 号：R394.2[医药卫生—医学遗传学]