成纤维细胞对乳腺癌细胞球体形成的影响及对干细胞相关基因表达的调控。

Effect of fibroblasts on breast cancer cell mammosphere formation and regulation of stem cell-related gene expression.

机构信息

Department of Oncology, Suzhou Kowloon Hospital, Shanghai Jiaotong University School of Medicine, Suzhou 215021, PR China.

出版信息

Int J Mol Med. 2011 Sep;28(3):365-71. doi: 10.3892/ijmm.2011.700. Epub 2011 May 13.

DOI:10.3892/ijmm.2011.700

PMID:21573501

Abstract

The purpose of this study was to investigate the regulatory effects of breast cancer fibroblasts (BCFs) vs. normal mammary fibroblasts (NMFs) on mammosphere formation and stem cell-related gene expression in breast cancer cells. Breast cancer cells (MCF-7) were cultured in suspension to generate primary and secondary mammospheres. The proportion of CD44+/CD24low/- cells was assessed by flow cytometry (FCM), and Wnt1, Notch1, β-catenin, CXCR4, SOX2 and ALDH3A1 gene expression was detected by quantitative real-time PCR. The fibroblasts from either breast cancer tissue or normal mammary tissue were purified from tissue specimens and co-cultured with breast cancer cells. The mammosphere formation efficacy was approximately 180/10,000 MCF-7 cells. FCM analysis showed that, compared to the 2.1% positive expression in the MCF-7 monolayer culture cells, the expression of CD44+/CD24low/- in MCF-7 mammosphere cells was significantly elevated to 10.4% (P<0.01). The proportion of the CD44+/CD24low/- subpopulation of the cells in mammospheres was nearly 5-fold higher than that of general MCF-7 cells. Compared with MCF-7 monolayer culture cells, mammosphere cells showed significantly (P<0.01) enhanced expression of Wnt1 [fold-change (FC), 2.25], Notch1 (FC, 2.45), β-catenin (FC, 1.72), CXCR4 (FC, 4.68), SOX2 (FC, 4.25) and ALDH3A1 (FC, 5.38). When BCFs were co-cultured with MCF-7 cells under mammosphere culture conditions, the length of time of mammosphere formation decreased, the volume of the mammo-spheres increased and the mammosphere-forming efficiency (MFE) was higher than that of NMFs and the control group. Both the BCF and NMF groups showed enhanced gene expression for the following genes: Wnt1 (FC, 3.18 and 1.27, respectively), β-catenin (FC, 1.75 and 1.22, respectively), Notch1 (FC, 2.09 and 1.31, respectively), CXCR4 (FC, 2.77 and 1.33, respectively), SOX2 (FC, 2.77 and 1.80, respectively) and ALDH3A1 (FC, 5.23 and 1.85, respectively). Cancer fibroblast cells can promote the MFE and up-regulate stem cell-related gene expression in breast cancer cells.

摘要

本研究旨在探讨乳腺癌成纤维细胞（BCFs）与正常乳腺成纤维细胞（NMFs）对乳腺癌细胞形成乳腺球和干细胞相关基因表达的调控作用。将乳腺癌细胞（MCF-7）在悬浮培养中培养以生成原发性和次级乳腺球。通过流式细胞术（FCM）评估 CD44+/CD24low/-细胞的比例，并用实时定量 PCR 检测 Wnt1、Notch1、β-catenin、CXCR4、SOX2 和 ALDH3A1 基因的表达。从组织标本中纯化来自乳腺癌组织或正常乳腺组织的成纤维细胞，并与乳腺癌细胞共培养。MCF-7 细胞的乳腺球形成效率约为 180/10000。FCM 分析表明，与 MCF-7 单层培养细胞中 2.1%的阳性表达相比，MCF-7 乳腺球细胞中 CD44+/CD24low/-的表达显著升高至 10.4%（P<0.01）。乳腺球中 CD44+/CD24low/-亚群细胞的比例几乎是普通 MCF-7 细胞的 5 倍。与 MCF-7 单层培养细胞相比，乳腺球细胞中 Wnt1[倍变化（FC），2.25]、Notch1（FC，2.45）、β-catenin（FC，1.72）、CXCR4（FC，4.68）、SOX2（FC，4.25）和 ALDH3A1（FC，5.38）的表达显著增强（P<0.01）。当 BCFs 在乳腺球培养条件下与 MCF-7 细胞共培养时，乳腺球形成的时间缩短，乳腺球的体积增加，乳腺球形成效率（MFE）高于 NMFs 和对照组。BCF 和 NMF 组的 Wnt1（FC，3.18 和 1.27，分别）、β-catenin（FC，1.75 和 1.22，分别）、Notch1（FC，2.09 和 1.31，分别）、CXCR4（FC，2.77 和 1.33，分别）、SOX2（FC，2.77 和 1.80，分别）和 ALDH3A1（FC，5.23 和 1.85，分别）的基因表达增强。癌症成纤维细胞可促进 MCF-7 细胞的 MFE，并上调乳腺癌细胞的干细胞相关基因表达。