Co-culture of functionally enriched cancer stem-like cells and cancer-associated fibroblasts for single-cell whole transcriptome analysis
Co-culture of functionally enriched cancer stem-like cells and cancer-associated fibroblasts for single-cell whole transcriptome analysis
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DOI：
10.1093/intbio/zyz029
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发表时间：
2019-09-01
影响因子：
2.5
通讯作者：
Yoon, Euisik
中科院分区：
文献类型：
--
作者：
Chen, Yu-Chih;Jung, Seungwon;Yoon, Euisik
大量证据表明，乳腺癌的发展和转移是由癌症干细胞样细胞（CSC）驱动的。由于其在肿瘤发生中的独特作用，CSC和基质细胞之间的相互作用尤其关键。在这项工作中，我们开发了一个平台，可以可靠地分离悬浮液中的单细胞，并生长单细胞衍生的球体，用于CSC的功能富集。该平台还允许贴壁培养基质细胞用于癌症-基质相互作用。作为概念验证，我们培养了SUM 149乳腺癌细胞，并成功形成了单细胞衍生的球体。发现作为基质细胞的癌症相关成纤维细胞（CAF）显著增强癌症球体的形成和生长，表明肿瘤起始潜力升高。在芯片上培养14天后，我们回收了有和没有CAF共培养的单细胞衍生的球体用于单细胞转录组测序。全转录组分析强调CAF共培养可以促进癌症干细胞，特别是ALDH（高）CSC，并改变上皮/间充质状态。单细胞分辨率允许识别个体CSC和研究癌症细胞异质性。利用公共患者数据库的全转录组测序数据，我们发现了与癌症-CAF相互作用相关且对患者生存至关重要的新基因。初步工作证明了一个可靠的平台，富集CSC和癌症间质相互作用的研究。
Considerable evidence suggests that breast cancer development and metastasis are driven by cancer stem-like cells (CSCs). Due to their unique role in tumor initiation, the interaction between CSCs and stromal cells is especially critical. In this work, we developed a platform to reliably isolate single cells in suspension and grow single-cell-derived spheres for functional enrichment of CSCs. The platform also allows adherent culture of stromal cells for cancer-stromal interaction. As a proof of concept, we grew SUM149 breast cancer cells and successfully formed single-cell-derived spheres. Cancer-associated fibroblasts (CAFs) as stromal cells were found to significantly enhance the formation and growth of cancer spheres, indicating elevated tumor-initiation potential. After on-chip culture for 14 days, we retrieved single-cell derived spheres with and without CAF co-culture for single-cell transcriptome sequencing. Whole transcriptome analysis highlights that CAF co-culture can boost cancer stemness especially ALDH(high) CSCs and alter epithelial/mesenchymal status. Single-cell resolution allows identification of individual CSCs and investigation of cancer cellular heterogeneity. Incorporating whole transcriptome sequencing data with public patient database, we discovered novel genes associated with cancer-CAF interaction and critical to patient survival. The preliminary works demonstrated a reliable platform for enrichment of CSCs and studies of cancer-stromal interaction.