单细胞蛋白质组图谱鉴定3,3′-二吲哚甲烷诱导乳腺癌的干细胞特性

Single cell proteomic profiling identified cancer stem cell plasticity of breast cancer cell induced by 3,3′-diindolylmethane

目的3,3’-二吲哚甲烷(DIM)是一种由吲哚-3-甲醇转化而来的二聚体化合物,被认为在乳腺癌化学预防药物具有应用前景。本研究首次从单细胞水平探讨了DIM诱导乳腺癌干细胞(CSC)的可塑性。方法采用MTT法检测细胞增殖。流式细胞仪检测MCF-7细胞周期分布和凋亡情况。采用单细胞质谱流式技术(CyTOF)检测DIM诱导的MCF-7细胞的单细胞蛋白质组学特征及其对CSCs的可塑性。结果DIM呈浓度依赖性抑制MCF-7细胞增殖。当DIM浓度为10、20、30μmol/L时,DIM分别诱导7%、11.3%和15%的细胞凋亡,S期细胞比例分别由16.6%降至15.1%、14%和8.5%,流式细胞仪检测结果显示,DIM处理后MCF-7细胞CD133+Nanog+亚群比例明显增加。SPADE分析显示DNA损伤和凋亡阳性细胞主要发生在干性标志表达相对较低的类群中。通过t-SNE分析,DIM诱导的信号通路蛋白在非CSC样细胞和CSC样细胞中的变化模式有显著差异。结论上述结果提示DIM在抑制MCF-7细胞增殖的诱导肿瘤干细胞的可塑性。

Objective 3,3′-diindolylmethane(DIM)is a dimer compound converted from indoly-3-carbinol that had been studied as promising chemo-preventive agent against breast cancer.In this study,we explored,for the first time,plasticity of breast cancer stem cell(CSC)induced by DIM in a single cell level.Methods Cell proliferation was detected by MTT assay.Flow cytometry was used to test cell cycle distribution and apoptosis in MCF-7 cells.Single cell proteomic profiling cancer stem cell plasticity of MCF-7 cell induced by DIM was performed by using mass cytometry(CyTOF).Results DIM inhibited MCF-7 cell proliferation in a concentration dependent way.When the concentration was 10,20,30μmol/L,DIM induced 7%,11.3%,and 15% apoptotic cell death in MCF-7 cell,respectively.DIM could also decrease S phase distribution from 16.6%to 15.1%,14%,and 8.5% when the concentration was 10,20,30μmol/L,respectively.Results from CyTOF indicated that proportion of CD133+Nanog+subpopulation in MCF-7 cells was significantly increased after treated with DIM.SPADE analysis revealed that DNA damage and apoptotic positive cells were mostly induced in the sebsets with relatively lower expression level of cancer stem cell markers.By using t-SNE analysis method,DIM induced signaling pathway proteins change pattern was significantly different between non-CSC like cells and CSC like cells.Conclusion Taken together,the above observations suggested that DIM could inhibit cell proliferation in MCF-7 cells,meanwhile,cancer stem cell plasticity could also be induced.