In vitro multipotent differentiation and barrier function of a human mammary epithelium

Abstract

As demonstrated by a variety of animal studies, barrier function in the mammary epithelium is essential for a fully functioning and differentiated gland. However, there is a paucity of information on barrier function in human mammary epithelium. Here, we have established characteristics of a polarizing differentiating model of human mammary epithelial cells capable of forming a high-resistance/low-conductance barrier in a predictable manner, viz., by using MCF10A cells on permeable membranes. Inulin flux decreased and transepithelial electrical resistance (TEER) increased over the course of several days after seeding MCF10A cells on permeable membranes. MCF10A cells exhibited multipotent phenotypic differentiation into layers expressing basal and lumenal markers when placed on permeable membranes, with at least two distinct cell phenotypes. A clonal subline of MCF10A, generated by culturing stem-like cells under non-adherent conditions, also generated a barrier-forming epithelial membrane with cells expressing markers of both basal and lumenal differentiation (CD10 and MUC1, respectively). Progressive changes associated with differentiation, including wholesale inhibition of cell-cycle genes and stimulation of cell and tissue morphogenic genes, were observed by gene expression profiling. Clustering and gene ontology categorization of significantly altered genes revealed a pattern of lumenal epithelial-cell-specific differentiation.

Growth of lung cancer cells in three-dimensional microenvironments reveals key features of tumor malignancy

“…NextBio meta-analysis revealed that expression data of 2D vs. 3D grown cells negatively correlates with data from differentiated vs. undifferentiated Caco-2 cells, 15 and with polarizing differentiating vs undifferentiated MCF10A cells. 16 This indicates that the 3D lrECM-cultured cells are more similar to the differentiated phenotype of both cell lines, consistent with previous studies showing activation of tissue-relevant phenotypic characteristics for mammary epithelial cells grown in tissue-like context. 23a Among individual transcripts in the 2D vs. 3D grown cells, SERPINB6 was the most consistently upregulated by growth in 3D lrECM (avg.…”

“…2A) 15 and from polarizing differentiating MCF10A cells in a Transwell model (P = 4.5e-45; Supplemental Fig. 2B), 16 consistent with the notion that even cells from long-term established lung cancer cell lines adopt a more differentiated phenotype when grown in 3D lrECM.…”

Multiple Cellular Responses to Serotonin Contribute to Epithelial Homeostasis

“…Through clonogenic assays we have previously shown the presence of mulipotent cells in MCF10A cultures [35]. Here, using BrdU pulse-chase and label retention assays we have been able to visualize the position of these cells, and demonstrate their ability to give rise to the other epithelial cell layers.…”

Loss of breast epithelial marker hCLCA2 promotes epithelial-to-mesenchymal transition and indicates higher risk of metastasis

“…For example, primary HMEC allowed to form acini in Matrigel have fivefold higher levels of hCLCA2 than cells cultured in monolayer (Dontu et al, Table S7 (Dontu, Abdallah et al 2003)). Similarly, MCF10A cultured on permeable membranes that support normal apico-basal polarization and other aspects of differentiation have an eightfold higher level of hCLCA2 than monolayers (Marshall, Pai et al 2009). A gene co-expression analysis using the Oncomine Clusters software revealed that hCLCA2 expression co-varies with epithelial markers such as claudins 1 and 8, E-cadherin, tetraspannins 1 and 7, and gap junctional proteins.…”