【摘要】 目的通过比较观察激素敏感的MCF-7细胞在体外干细胞培养和常规培养条件下雌激素受体(ER)变化及对内分泌治疗药物的敏感性变化,初步探讨肿瘤干细胞与内分泌耐药的关系。方法分别于常规培养及干细胞培养条件下(悬浮球培养)培养激素敏感的MCF-7细胞株,流式细胞仪检测分子表型CD44+CD24-/low与CD44+CD24+亚群细胞比例变化,免疫细胞化学法测定ERα和ERβ的表达变化,MTT法检测细胞对他莫西芬的敏感程度。分别以t检验、卡方检验、方差分析进行统计分析。结果干细胞培养条件下CD44+CD24-/low亚群细胞的比例为(1.60±0.08)%,比常规培养条件下的(0.27±0.08)%显著增加(t=-12.10,P=0.00),而CD44+CD24+亚群细胞比例由(5.59±0.88)%增至(30.63±4.40)%(t=-5.58,P=0.00)。干细胞培养条件培养下ERα和ERβ表达率较常规培养下调,分别由85.27%和90.53%降至69.43%和73.20%,差异有统计学意义(χ2=214.64,P=0.00;χ2=303.58,P=0.00),且对他莫西芬的敏感性降低,IC50值由(9.82±0.31)μmol/L升至(16.46±0.50)μmol/L,再次诱导分化后ER并未出现上调,对他莫西芬的敏感性仍旧降低(F=113.63,P=0.00)。结论与常规培养相比较,体外干细胞培养条件下可以培养出含高比例具有干细胞特性的CD44+CD24-/low与CD44+CD24+亚群细胞的微球囊,其ER仍为阳性表达,但对他莫昔芬治疗敏感性减低,推测其可能是乳腺癌内分泌耐药的原因。更多还原

【Abstract】 Objective To explore estrogen receptor(ER) expression and endocrine resistance of hormone-sensitive MCF-7 cells in normal culture or in stem cell culture in vitro.Methods The MCF-7 cells were cultured in normal culture or in stem cell culture in vitro(suspension sphere).The proportion of CD44+CD24-/low phenotype and CD44+CD24+ phenotype cells were determined using flow cytometry(FCM).The ERa and ERβ expression was detected using immunohistochemical method.The susceptibility to tamoxifen was determined using MTT essay.The data were processed with t-test,Chi-square test and analysis of variance,respectively.Results In suspension culture as stem cells,the proportions of the CD44+CD24-/low and CD44+CD24+ phenotype cells were(1.60±0.08)% and(30.63±4.40)% respectively,significantly higher than(0.27±0.08)% and(5.59±0.88)% in normal medium(t=-12.10,P=0.00;t=-5.58,P=0.00).The positive expressions of ERα and ERβ were 85.27% and 90.53% in normal medium,69.43% and 73.20% in suspension culture,respectively,and the differences were statistically significant(χ2=214.64,P=0.00;χ2=303.58,P=0.00).With regards to tamoxifen sensitivity,IC50 was(9.82±0.31)μmol/L in normal medium,and(16.46±0.50) μmol/L in suspension culture.After the second induced differentiation,the positive expression of ER was not up-regulated and the susceptibility to tamoxifen remained low(F=113.63,P=0.00).Conclusion Suspension sphere with a high ratio of CD44+CD24-/low cells and CD44+CD24+ cells can be incubated in stem cell culture in vitro,with higher positive expression of ER and lower sensitivity to tamoxifen,which may be attributed to endocrine resistance.更多还原