Biophysics Institute, Faculty of Medicine, University of Coimbra, Coimbra, Portugal; CIMAGO, Faculty of Medicine, University of Coimbra, Coimbra, Portugal; CNC.IBILI, University of Coimbra, Coimbra, Portugal.
Biophysics Institute, Faculty of Medicine, University of Coimbra, Coimbra, Portugal; CIMAGO, Faculty of Medicine, University of Coimbra, Coimbra, Portugal; Gynecology A Service, Coimbra Hospital and Universitary Centre, Coimbra, Portugal.
Breast. 2018 Apr;38:22-29. doi: 10.1016/j.breast.2017.11.009. Epub 2017 Nov 26.
This study aimed to characterize mammospheres from hormonal receptor (HR) positive and triple-negative breast cancer (TNBC), hypothesizing a differential profile of CSC and differentiation markers, and a stemness enrichment when successive sphere forming-protocols are performed.
Breast cancer cells MCF-7 and HCC1806 were submitted to sphere-forming protocols. The first sphere generation (MS1) was cultured in adherent conditions (G1). This procedure was repeated and generations of mammospheres (MS1, MS2, and MS3) and sphere-derived cells in adherent conditions (G1, G2, and G3) were obtained. The mammosphere forming capacity, self-renewal, area and doubling time were evaluated. Flow cytometry regarding CD133, CD24, and CD44 and western-blot regarding aldehyde dehydrogenase (ALDH), hormonal receptors and P53 expression was performed.
Breast cancer cell lines harboured the capacity to form spheres, which originated derived adherent populations. The sphere-forming capacity was enhanced in HCC1806-MS3 compared to MS1. Self-renewal was higher in MCF-7 mammospheres, which also had an increased area. The putative CSC markers CD133 showed tendency to be enhanced in mammospheres but the CD44/CD24 phenotype was not identified. The expression of ALDH was greater in mammospheres from MCF-7 and HCC1806 than in the respectively derived adherent cells. The expression of oestrogen receptor (ER)-α, progesterone receptor (PR) and P53 decreased in MCF-7 spheres. ER-β expression was lower in mammospheres from both cell lines compared with parental and derived adherent populations.
Loss of HR and P53 expression in HR-positive mammospheres evidences the minor population of CSC which shares characteristics with the TNBC phenotype.
本研究旨在对激素受体（HR）阳性和三阴性乳腺癌（TNBC）的乳腺球体进行特征描述，假设当进行连续的球体形成方案时，会出现干细胞和分化标志物的差异特征，并且存在干细胞富集。
将乳腺癌细胞 MCF-7 和 HCC1806 进行球体形成方案处理。第一代球体生成（MS1）在贴壁条件下培养（G1）。重复此过程，获得几代乳腺球体（MS1、MS2 和 MS3）和贴壁条件下的球体衍生细胞（G1、G2 和 G3）。评估乳腺球体的形成能力、自我更新、面积和倍增时间。进行 CD133、CD24 和 CD44 的流式细胞术以及醛脱氢酶（ALDH）、激素受体和 P53 表达的 Western blot。
乳腺癌细胞系具有形成球体的能力，这些球体源自贴壁细胞。与 MS1 相比，HCC1806-MS3 的球体形成能力增强。MCF-7 乳腺球体的自我更新能力更高，其面积也更大。假定的干细胞标志物 CD133 在乳腺球体中表现出增强的趋势，但未鉴定出 CD44/CD24 表型。ALDH 的表达在 MCF-7 和 HCC1806 的乳腺球体中高于相应的贴壁细胞。MCF-7 球体中雌激素受体（ER）-α、孕激素受体（PR）和 P53 的表达降低。与亲本和贴壁细胞群体相比，两种细胞系的乳腺球体中 ER-β 的表达均较低。
HR 阳性乳腺球体中 HR 和 P53 表达的丧失表明，具有与 TNBC 表型相似特征的干细胞的亚群较少。
