Project description:To delineate epithelial subpopulations in human mammary tissue, hematopoietic and endothelial cells were depleted from freshly isolated cell suspensions derived from reduction mammoplasties by fluorescence-activated cell sorting. The resultant Lin- population was fractionated into four distinct subpopulations using CD49f (α6-integrin) and epithelial cell adhesion molecule (EpCAM; also referred to as CD326 and ESA). Based on the immunohistochemical phenotype, and in vivo and in vitro functional assays, these subpopulations were identified as fibroblast-enriched stromal (CD49f -EpCAM-), mammary stem cell (MaSC)-enriched (CD49f hiEpCAM-), luminal progenitor (CD49f +EpCAM+), and mature luminal (CD49f –EpCAM+) cell subpopulations. Microarray profiling was used to derive gene expression signatures representative of these subpopulations using freshly sorted cells (>90% purity) from normal breast tissue. The four mammary cell subpopulations were found to have distinct gene expression profiles.
Project description:To delineate epithelial subpopulations in human mammary tissue, hematopoietic and endothelial cells were depleted from freshly isolated cell suspensions derived from reduction mammoplasties by fluorescence-activated cell sorting. The resultant Lin- population was fractionated into four distinct subpopulations using CD49f (α6-integrin) and epithelial cell adhesion molecule (EpCAM; also referred to as CD326 and ESA). Based on the immunohistochemical phenotype, and in vivo and in vitro functional assays, these subpopulations were identified as fibroblast-enriched stromal (CD49f -EpCAM-), mammary stem cell (MaSC)-enriched (CD49f hiEpCAM-), luminal progenitor (CD49f +EpCAM+), and mature luminal (CD49f âEpCAM+) cell subpopulations. Microarray profiling was used to derive gene expression signatures representative of these subpopulations using freshly sorted cells (>90% purity) from normal breast tissue. The four mammary cell subpopulations were found to have distinct gene expression profiles. Four mammary cell subpopulations from three individual patient samples were analyzed.
Project description:This experiment shows differential expression of genes in the luminal, basal and stromal subpopulations from SNAI2+/+ and SNAI2 LacZ/LacZ mammary epithelial cells. Luminal, basal and stromal populations were sorted from SNAI2+/+ and SNAI2 LacZ/LacZ mammary epithelial cells based on expression of CD49f and Epcam.
Project description:Gene expression profiles of normal human mammary epithelial cell subsets enriched for luminal or myoepithelial characteristics on the basis of EpCAM and CD49f expression. Transcription profiles were compared between luminal- and myoepithelial-enriched human MEC subsets following 3D culture from 4 individuals. Gene expression profiles of normal human mammary epithelial cell subsets enriched for mature, luminal progenitor or bipotent progenitor-enriched characteristics on the bais of CD49f, MUC1, CD10, CD133 and Thy1 expression Transcription profiles were compared between unsorted, mature, luminal progenitor-enriched and bipotent progenitor-enriched human MEC subsets following 3D culture from 2 individuals.
Project description:Genomic instability is a prominent driver of tumorigenesis. However, a significant fraction of human cancers display few genomic aberrations, suggesting alternative roads towards malignancy. Here, we show that the differentiation status of normal human mammary epithelial cells influences the early response to an oncogenic activation and determines the genetic routes towards tumorigenesis. Following an oncogenic insult, luminal progenitors and differentiated luminal cells undergo oxidative and DNA replication stress, initiating genomic instability. In contrast, mammary stem cells exhibit the innate capacity to withstand aberrant mitogenic activation, fostering malignant transformation. This property relies upon a pre-emptive program driven by the ZEB1 transcription factor and the methionine sulfoxide reductase MSRB3. The ZEB1-MSRB3 axis governs cellular pliancy and prevents the continuous formation of oncogene-induced DNA damage, leading to neoplasms with unique pathological features. These gene expression data correspond to the different subpopulations that compose the hierarchy of normal human mammary epithelial cells. These subpopulations were freshly isolated from mammary tissue, originating from reduction mammoplasties and flow sorted using four markers (EpCAM, CD10, CD49f, ALDH). Three subpopulations enriched in mammary stem cells (MaSCs) are designed as MaSC1, 2, 3; the luminal progenitor are designed as LP, and the mature luminal cells as mL1 and mL2.
Project description:Gene expression profiles of normal human mammary epithelial cell subsets enriched for luminal or myoepithelial characteristics on the basis of EpCAM and CD49f expression. Transcription profiles were compared between luminal- and myoepithelial-enriched human MEC subsets following 3D culture from 4 individuals. Gene expression profiles of normal human mammary epithelial cell subsets enriched for mature, luminal progenitor or bipotent progenitor-enriched characteristics on the bais of CD49f, MUC1, CD10, CD133 and Thy1 expression Transcription profiles were compared between unsorted, mature, luminal progenitor-enriched and bipotent progenitor-enriched human MEC subsets following 3D culture from 2 individuals. Four replicates of luminal-enriched human MECs and 3 replicates of myoepithelial-enriched MECs Transcription profiles of two replicates of unsorted, mature, luminal progenitor-enriched and bipotent progenitor-enriched human MEC subsets were analysed following 3D culture
Project description:In this study, we grew primary breast epithelial cells from breast biopsies of healthy donors, five each of African Ancestry, European ancestry and those who identify as Hispanic using epithelial cell reprogramming assay growth method. Cells grown were sorted into luminal progenitors (CD49F+/EpCAM+, labelled as PP cells) and mature luminal (CD49F-/EpCAM+, labelled as NP cells) by flow cytometry. Flow sorted cells were subjected to RNA-seq.
Project description:Many normal adult tissues contain rare stem cells with extensive self-maintaining regenerative potential. During development, the stem cells of the hematopoietic and neural systems undergo intrinsically specified changes in their self-renewal potential. In the mouse, mammary stem cells with transplantable regenerative activity are first detectable a few days before birth. They share some phenotypic properties with their adult counterparts but are enriched in a subpopulation that displays a distinct gene expression profile. Here we show that fetal mammary epithelial cells have a greater direct and inducible growth potential than their adult counterparts. The latter feature is revealed in a novel culture system that enables large numbers of mammary stem cells with serially transplantable activity as well as in vitro clonogenic progenitors to be produced within 7 days from single fetal or adult input cells. We further show that these responses are highly dependent on novel factors produced by fibroblasts. These findings provide new avenues for elucidating mechanisms that regulate normal mammary epithelial stem cell properties at the single-cell level, how these change during development, and how their perturbation may contribute to transformation. We used microarrays to compare the transcriptome of E18.5 fetal and adult MRU-enriched mammary cells. Three biological replicates each of CD31-CD45-Ter119-BP-1-EpCAM+CD49f+ adult basal cells and CD31-CD45-Ter119-EpCAM++CD49f+ fetal cells were sorted. RNA was extracted and hybridized to the Agilent One-Color Gene Expression Arrays .
Project description:Stromal-epithelial interactions play a fundamental role in tissue homeostasis, controlling cell proliferation and differentiation. The goals and objectives of this study were 1.) To generate gene expression profiles and identify signatures of freshly isolated, non-cultured, luminal epithelial or basal epithelial and CD10 positive or negative stromal cell populations from the human breast. 2.) Use the generated signatures to characterize and validate the molecular identity of human primary epithelial and stromal/mesenchymal breast cells maintained long-term in novel ex vivo culture conditions in serum free medium (previous study in our lab) Breast tissue from three different reduction mammoplasties was dissociated and single cells were labeled with antibodies against lineage markers (CD31, CD45 CD235ab), CD10, EpCAM, CD49f and DAPI. Different cell populations were identified, separated and sorted with a fluorescence activated cell sorter. Gene expression profiling of luminal epithelial (EpCAM+, CD49f+/-), basal epithelial (EPCAM-, CD49f+), CD10 negative and CD10 positive stromal cells was performed and distinct gene expression signatures for the four cell populations were identified.