目的 探讨ALDH1高表达对乳腺癌MCF-7细胞干细胞特性的影响,为乳腺癌的治疗提供理论依据.方法 构建慢病毒ALDH1RNA,转染乳腺癌MCF-7细胞,嘌呤霉素筛选稳定高表达ALDH1的亚克隆,命名为MCF-ALDH1细胞株.荧光定量PCR和Western blot法检测MCF-ALDH1细胞株及对照细胞中ALDH1在mRNA和蛋白水平的表达;克隆形成实验、悬浮球形成实验及裸鼠成瘤实验检测MCF-ALDH1及对照组细胞的干细胞特性,MTT法测定细胞的耐药性.结果 与MCF-7组细胞比较,MCF-ALDH1细胞株的ALDH1在mRNA和蛋白表达量明显升高;MCF-ALDH1细胞克隆形成数目为128±22个,高于MCF-ALDH1细胞球克隆形成数(46±15个);悬浮成球实验,MCF-ALDH1细胞悬浮成球数目明显增多(16±2个),高于对照细胞悬浮球数(8.0±1.5个);肿瘤细胞在裸鼠皮下接种后,MCF-7细胞组有3只裸鼠成瘤,MCF-ALDH1有5只成瘤,瘤体积明显大于MCF-7组,差异有统计学意义(P＜0.05).耐药实验显示,MCF-ALDH1对顺铂、紫杉醇的耐药性明显提高.结论 高表达ALDH1的乳腺癌MCF-7细胞干细胞特性增强,ALDH1可能成为乳腺癌治疗的靶点之一.%Objective To investigate the expression and significance of ALDH1 in cancer stem cell-like characteristics in breast cancer cell line MCF-7.Methods RNA lenti-virus particles targeting ALDH1 gene were transfected into MCF-7 cells.Subclones with stable ALDH1 expression were selected by puromycin and named MCF-ALDH1 cells.The mRNA and protein expression of ALDH1 were detected by real-time quantitative PCR and Western blot tests.Cell differentiation,stem cell potential were observed by clone formation assay and suspension sphere-forming assay.The abilities of tumor-formation of cells were detected through inoculating the cells into the nod mouse.To detect the drug-resistance ability of cells by MTT test.Results Compared to the MCF-7 cells,the expression of mRNA and protein of ALDH1 by MCF-ALDH1 cells were significantly increased.Clony forming numbers(128 ±22) of the MCF-AL-DH1 cells was higher than controls(46 ± 15)(P ＜ 0.05),The proportion of sphere-initiating cells in MCF-ALDH1 and controls was (16 ± 2) versus(8.0 ± 1.5,P ＜ 0.05).The average tumor volume of MCF-ALDH1 was greater than controls(P ＜ 0.05).The drug resistance index of MCF-ALDH1 was higher than controls.Conclusion Experimental findings from this study showed that MCF-7 cells highly expressing ALDH1 genes,with higher proliferation and differentiation,in vivo tumorigenicity and potential of stem cells.ALDH1 may be used as an target to cure breast cancer.
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