【摘要】 探讨利用细胞周期特异性药物5-氟尿嘧啶(5-fluorouracil,5-FU)从人白血病细胞系KG-1a中富集肿瘤干细胞样亚群细胞。体外药物敏感试验确定5-FU的最佳作用浓度和作用时间;KG-1a细胞经5-FU药物处理后,流式细胞术检测存活细胞群中CD34+CD38-亚群细胞比例;吖啶橙染色观察细胞内的核酸组成;RT-PCR半定量检测细胞内三磷酸腺苷结合转运蛋白G超家族成员2(ATP-binding cassette superfamily Gmember2,ABCG2)mRNA的表达;半固体培养观察细胞的集落形成能力。结果显示,50μg/ml5-FU作用KG-1a细胞4天后,CD34+CD38-亚群细胞比例提高10倍以上;吖啶橙染色可见大部分细胞核酸以发出绿色荧光的DNA为主,RNA含量低;此类细胞高表达ABCG2 mRNA水平;而药物处理后细胞集落形成数量较未处理细胞明显减少。研究结果表明,利用5-FU能够杀死增殖期细胞的性质,成功建立体外药物筛选富集人白血病细胞系KG-1a中肿瘤干细胞样亚群细胞的方法。更多还原

【Abstract】 To enrich tumor stem cell-like subpopulation in human leukemia cell lines KG-1a with 5-fluorouracil(5-FU), optimal concentration and effect time of 5-FU were determined with in vitro drug sensitivity assay.The CD34+CD38-subpopulation in the enriched subpopulation of KG-1a cells was detected with flow cytometry.The RNA content of enriched subpopulation cells was measured by using acridine orange staining, and their expression of ABCG2 mRNA by RT-PCR.The colony-forming ability of this subpopulation cells was evaluated by semi-solid culturing.It was indicated through our research that after treatment of KG-1a cells with 50 μg/ml 5-FU for 4 days, the proportion of CD34+CD38-cells in enriched subpopulation was elevated by more than 10 folds;The RNA content of enriched subpopulation cells was low, but the ABCG2 mRNA was highly expressed.The colony forming ability of enriched subpopulation cells was lower than of non-enriched KG-1a cells.Our study suggested that utilizing 5-FU for killing most proliferating cells, we successfully established the method in which selecting and enriching tumor stem-like subpopulation cells in human leukemia cell lines KG-1a with 5-FU.更多还原