Interaction of bone marrow stroma and monocytes
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE9390 | GEO | 2008/03/01
SECONDARY ACCESSION(S): PRJNA103111
REPOSITORIES: GEO
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Project description:The bone marrow microenvironment is a complex mixture of cells that function in concert to regulate hematopoiesis. Cellular components include fixed nonhematopoietic stromal elements as well as monocytes and resident macrophages, which are derived from the hematopoietic stem cells. Although these monocyte-lineage cells are reported to modify stromal cell function, the reverse also occurs. Given the secretory capability of the monocyte/macrophage and their various potential functions, it is not surprising that stromal cells contained within a particular niche can modify monocyte gene expression and functional maturation. Experiment Overall Design: Monocytes were isolated from peripheral blood mononuclear cells from 2 different normal donors and cultured for 48h in conditioned medium (CM). The CM was collected from each of two functionally distinct human bone marrow stromal cell lines (HS5 and HS27a) representing different compartments of the bone marrow microenvironment (Roecklein BA, Torok-Storb B. Blood. 1995;85:997-1005). Four samples were analyzed, with two biological replicates for each CM.
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Project description:The bone marrow microenvironment is a complex mixture of cells that function in concert to regulate hematopoiesis. Cellular components include fixed nonhematopoietic stromal elements (MSC) as well as monocytes and resident macrophages, which are derived from the hematopoietic stem cells. Clinical studies have shown healing effects, direct or indirect, from the injection of MSC int Keywords: Cell type comparison
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Project description:The bone marrow microenvironment is a complex mixture of cells that function in concert to regulate hematopoiesis. Cellular components include fixed nonhematopoietic stromal elements (MSC) as well as monocytes and resident macrophages, which are derived from the hematopoietic stem cells. Clinical studies have shown healing effects, direct or indirect, from the injection of MSC int Keywords: Cell type comparison Canine bone marrow stromal cells were immortalized by transduction with a replication-defective recombinant retrovirus containing the human papilloma virus E6/E7 genes (pLXSN-16 E6E7), as described for human stromal cell lines in Roecklein and Torok-Storb (Blood 85:997-1005, 1995). Transduced clones were selected with 50 ug/ml G418 and resistant clones were isolated, grown to confluency and characterized for morphology, cytokine production and cell surface molecule expression. These cells are intended for clinical research in the canine transplantation model, as a complement for existing human stromal cell lines, for mitigation of graft-versus-host disease and radiation effects.
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Project description:The hematopoietic microenvironment consists of non-hematopoietic derived stromal elements and hematopoietic derived monocytes and macrophages which interact and function together to control the proliferation and differentiation of early blood-forming cells. Two human stromal cell lines (HS-5 and HS-27a) representing distinct functional components of this microenvironment have been extensively characterized and shown to influence monocyte gene expression. This series of gene expression profiles is intended to extend the previous studies and identify which gene expression changes may require cell-cell contact or occur in the stromal cells as a result of monocyte influence;or in the monocytes as a result of stormal influences. Keywords: cellular response to cell contact and secreted factors
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Project description:Tumor epithelial cells develop within a microenvironment consisting of extracellular matrix, growth factors, and cytokines produced by non-epithelial stromal cells. In response to paracrine signals from tumor epithelia, stromal cells modify the microenvironment to promote tumor growth and metastasis. Here, we identify interleukin (IL)-33 as an epithelial cell-derived regulator of stromal cell activation and mediator of intestinal polyposis. IL-33 expression was elevated in the tumors and serum of colorectal cancer patients and induced in the adenomatous polyps of ApcMin/+ mutant mice. Genetic and antibody suppression of IL-33 signaling in ApcMin/+ mice inhibited proliferation, induced apoptosis, and suppressed angiogenesis in polyps, which reduced both tumor number and size. In ApcMin/+ polyps, IL-33 expression localized to tumor epithelial cells and expression of the IL-33 receptor, IL1RL1, associated with two stromal cell types, namely subepithelial myofibroblasts (SEMFs) and mast cells, whose activation was previously associated with polyposis. In vitro IL-33 stimulation of human SEMFs induced the expression of extracellular matrix components and growth factors associated with intestinal tumor progression. IL-33 deficiency reduced mast cell accumulation in ApcMin/+ polyps and expression of mast cell-derived proteases and cytokines known to promote polyposis. Together, our results suggest that IL-33 is a tumor epithelial cell-derived paracrine signal that promotes polyposis through the coordinated activation of stromal cells and the formation of a reactive stroma microenvironment.
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Project description:Gene expression of adherent cells in a coculture of the human bone marrow stromal cell line HS-27a and peripheral blood monocytes (CD14+ cells) was compared to HS-27a cultured alone. Microarray experiments were conducted using CD14+ cells from 7 healthy donors over a period of >7 months. Monocytes were isolated by ficoll separation of whole blood, followed by labelling cells with CD14 monoclonal antibody Tuek4 and magnetic bead selection (Milteny rat anti mouse IgG2a+2b beads) on an AutoMacs. HS27a cells were plated in T75 flasks at approximately 80% confluence. The next day 1.5x10E6 freshly isloated human CD14+ cells were added to the culture. At 3 days the cultures were washed 3x with Hankâ??s buffer to remove nonadherent monocytes. Co-cultures and HS27a cultured alone were harvested by brief trypsinization followed by pelleting of the cells. All RNA isolation was accomplished with Qiagen RNeasy Mini Kit reagents. The RNA (25 ug) was annealed with 5 ug oligo dT12-18, and reverse-transcribed into cDNA with Superscript II reverse transcriptase for 2h at 42ºC in the presence of 0.5mM dGTP, 0.5mM dCTP, 0.5mM dATP, 0.3mM dTTP, 0.2mM amino-allyl dUTP. After hydrolysis of RNA in 0.2M NaOH, Tris was removed from the reaction with a Microcon-30 concentrator. The cDNA from HS27a and HS27a-monocyte cocultures was covalently coupled separately with Cy5 and Cy3 monoreactive fluors, respectively, in 50mM sodium bicarbonate, pH 9.0, followed by quenching with 2.7M hydroxylamine. The Cy5 and Cy3 labelled cDNAs were combined and purified with a QIAquick PCR purification kit and suspended in 36 ul of 3X SSC and 0.8 mg/ml of poly-A for hybridization to the microarray. Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000A fluorescent scanner and image intensity data were extracted and analyzed with GenePix Pro 3.0 analysis software. After background correction and removal of flagged values, log base 2 expression ratios were mean centered and linear transformed to obtain the log and linear values given in the data tables.
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