在体阿霉素干预联合全克隆培养方法富集肾上腺皮质癌肿瘤干细胞的研究

A novel approach for enriching cancer stem cells from the human SW13 adrenocortical carcinoma cell line by the intervention of doxorubicin in vivo combined holoclone culture

目的通过阿霉素在体干预联合全克隆的富集方法来研究肾上腺皮质癌SW13肿瘤干细胞的特性.方法建立裸鼠SW13移植瘤连续传代3代模型,实验组尾静脉注射阿霉素,对照组注射等量生理盐水.收集移植瘤细胞进行单克隆培养,所获克隆分别进行克隆转移实验、克隆连续传代实验、Hoechst33342/PI双染色检测凋亡率实验,并通过RT-qPCR检测肿瘤标志物CXCR4在两组瘤体细胞中的表达.结果与对照组相比,实验组的瘤体直径小、CXCR4表达更高,实验组全克隆细胞的克隆转移能力、连续传代能力最高、凋亡率最低(P<0.05).结论在体阿霉素干预联合全克隆富集方法可以有效富集肾上腺皮质癌肿瘤干细胞,肾上腺皮质癌肿瘤干细胞高表达CXCR4.

Objective To enriching cancer stem cells(CSC)from the human SW13 adrenocortical carcinoma(ACC)cell line by the intervention of doxorubicin in vivo combined holoclone culture.Methods The xenograft tumor model of experimental group(passaging 3 times consecutively)was induced by doxorubicin,relatively,the control group was induced by physiological saline.The expression of CXCR4 in the 2 groups was determined by RT-qPCR technology.Subsequently,the 3rd generation xenograft tumor cells of the 2 groups were collected for single-cell clone culture.ACC holoclone cells were collected for the experiments as follows:metastatic coloniczation experiment,consecutive passage experiments of colonies and Hoechst33342/PI double staining assay.Results The maximum diameters of xenograft tumors in the control group were larger than those in the experimental group.The expression of CXCR4 in the experimental group was higher than that in the control group.The metastatic coloniczation ability and consecutive passage ability of holoclone cells in the experimental group were higher than others.The apoptosis rate of holoclone in the experimental group was the lowest(P<0.05).Conclusion The novel approach by the intervention of doxorubicin in vivo combined holoclone culture is able to enrich cancer stem cells.CXCR4 may be the tumor specific marker of ACC-CSC.