Unknown,Transcriptomics,Genomics,Proteomics

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Gene transcript alterations exhibited in luminal and basal epithelial compartments with impaired Ror2 signaling

ABSTRACT: Microarray analyses revealed distinct gene expression alterations in the luminal and basal epithelial compartments in the absence of Ror2 with marked alterations observed in genes associated with actin filament-based processes and the actin cytoskeletal network. Given that Ror2 expression resides in both basal and luminal epithelial cells, we wanted to identify compartment-specific functions for Ror2 in coordinating development. Accordingly, luminal and basal epithelial cell populations from shLUC and shRor2 outgrowths were isolated by FACS separation on the basis of CD24 and CD29 surface marker expression and gene expression analysis was subsequently performed using Agilent microarrays.

ORGANISM(S): Mus musculus

SUBMITTER: Chad Creighton

PROVIDER: E-GEOD-57985 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Identification and transcriptome analysis of mammary stem cells (MaSC) are important steps toward understanding the molecular and cellular bases of mammary epithelial growth, homeostasis and tissue repair. We identified MaSC based upon their ability to retain bromodeoxyuridine (BrdU) for an extended period. Using laser microdissection, we isolated four categories of cells from the mammary epithelium of female bovine calves. These categories included label retaining epithelial cells (LREC) from the basal layer (LRECb), basal control cells (ECb), LREC from embedded layers (LRECe) and embedded control cells (ECe). Microarray analysis revealed 592 genes that were differentially expressed between LRECb and ECb, and 110 genes between LRECe and ECe. Our analysis supported classification of LRECb as MaSC and LRECe as progenitor cells. Enriched expression of genes in LRECb were associated with stem cell attributes and identified enrichment of Wnt, Notch and MAPK pathways of self renewal and proliferation. Genes expressed in LRECe revealed retention of some stem-like properties along with up-regulation of differentiation factors and the Notch pathway involved in lineage commitment. The basal epithelium provided for the stem cell niche, characterized as a microenvironment containing numerous survival and proliferation factors, growth suppressors and chromatin modifiers in balance state. We conclude that BrdU label retention identifies stem and progenitor cells, wherein MaSC are located in the basal epithelial layer and committed progenitor cells are localized in more apical layers. Results provide novel candidate markers for bovine MaSC with potential applicability to the study of MaSC in other species. Five Holstein heifers were injected with bromodeoxyuridine (BrdU) for five consecutive days and sacrificed 45 days later, at which time mammary samples were obtained and processed. Cryosections were prepared and immunostained to detect epithelial cells that retained BrdU label (LREC). Four classes of cells were excised by laser microdissection, which included LREC and control cells in the basal and in the suprabasal layers of the mammary epithelium. For each cell type, cells were lysed, RNA was reverse transcribed and cDNA amplified for microarray analysis. Twenty microarrays were utilized for sample hybridization and another array for hybridization of amplified cDNA from a water blank.

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Gene transcript alterations exhibited in luminal and basal epithelial compartments with impaired Ror2 signaling

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