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Cord Blood-Derived Mesenchymal Stem Cells with Distinct Growth Kinetics, Differentiation Potentials, Expression Profiles

ABSTRACT: Phenotypic heterogeneity has been observed among mesenchymal stem/stromal cell (MSC) populations, but specific genes associated with this variability have not been defined. To study this question, we analyzed two distinct MSC populations isolated from the same umbilical cord blood (UCB) sample. These populations (UCB1 and UCB2) are from a single donor, minimizing differences contributed by genetic background. We characterized these UCB-MSCs for cell morphology, growth kinetics, immunophenotype and differentiation potential. UCB1 displayed rapid growth kinetics, higher population doublings, and increased adipogenic lineage differentiation compared to UCB2. To identify the MSC-specific and developmental genes associated with these phenotypic differences, we performed expression analysis using Affymetrix HG-U133 microarrays and compared them to bone marrow (BM) MSCs. First, hepatocyte growth factor (HGF) and stromal derived factor 1 (SDF1/CXCL12) were up -regulated in UCB1 cells, potentially contributing to the higher growth kinetics observed in this circulating cell population. Second, we observed that peroxisome proliferation activated receptor gamma (PPARG), a marker for adipogenic differentiation, was significantly increased in undifferentiated UCB1 cells. Moreover, significant expression of gene markers of blastocyst and gatrulation embryonic stages were detected in UCB1 and UCB2 cells, as were selected markers of early hematopoiesis, chondrogenesis, and cardiac differentiation. Comparison of UCB1, UCB2, and BM by microarray analysis clearly demonstrated clusters of developmental genes that displayed significant differences among these cells. Quantitative PCR analysis of selected genes validated the microarray results. Comparison of different UCB-derived adherent cells from a single donor has identified gene profiles potentially useful for therapeutic evaluation of MSC populations. Keywords: cell type comparison

ORGANISM(S): Homo sapiens

PROVIDER: GSE6029 | GEO | 2007/02/01

SECONDARY ACCESSION(S): PRJNA97753

REPOSITORIES: GEO

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Project description:Phenotypic heterogeneity has been observed among mesenchymal stem/stromal cell (MSC) populations, but specific genes associated with this variability have not been defined. To study this question, we analyzed two distinct MSC populations isolated from the same umbilical cord blood (UCB) sample. These populations (UCB1 and UCB2) are from a single donor, minimizing differences contributed by genetic background. We characterized these UCB-MSCs for cell morphology, growth kinetics, immunophenotype and differentiation potential. UCB1 displayed rapid growth kinetics, higher population doublings, and increased adipogenic lineage differentiation compared to UCB2. To identify the MSC-specific and developmental genes associated with these phenotypic differences, we performed expression analysis using Affymetrix HG-U133 microarrays and compared them to bone marrow (BM) MSCs. First, hepatocyte growth factor (HGF) and stromal derived factor 1 (SDF1/CXCL12) were up -regulated in UCB1 cells, potentially contributing to the higher growth kinetics observed in this circulating cell population. Second, we observed that peroxisome proliferation activated receptor gamma (PPARG), a marker for adipogenic differentiation, was significantly increased in undifferentiated UCB1 cells. Moreover, significant expression of gene markers of blastocyst and gatrulation embryonic stages were detected in UCB1 and UCB2 cells, as were selected markers of early hematopoiesis, chondrogenesis, and cardiac differentiation. Comparison of UCB1, UCB2, and BM by microarray analysis clearly demonstrated clusters of developmental genes that displayed significant differences among these cells. Quantitative PCR analysis of selected genes validated the microarray results. Comparison of different UCB-derived adherent cells from a single donor has identified gene profiles potentially useful for therapeutic evaluation of MSC populations. Experiment Overall Design: In order, to identify developmental genes whose expression differences distinguished UCB1 and UCB2 cell MSCs, we carried out microarray analysis using Affymetrix HG-U133 microarrays. For comparison, adult BM-derived MSCs (Cambrex, Lot #4F0312) were analyzed using the same cell culture conditions and confluence as UCB-MSCs. RNA was prepared from three biological replicates for UCB1, UCB2, and BM MSCs, for a total of nine samples analyzed on the HG-U133A and HG-U133B arrays.

Project description:Cyclic regeneration of the endometrium, and its repair after parturition or injury, are crucial for successful reproduction. Mesenchymal stem cells (MSCs) derived from bone marrow (BM-MSC) facilitate tissue repair via their secretome, which contains growth factors and cytokines that promote wound healing. Despite the implication of MSCs in endometrial regeneration and repair, the mechanisms remain unclear. This study tested the hypothesis that the secretome of MSCs from human BM upregulates human endometrial stromal cell (HESC) proliferation, migration and invasion, and activates pathways to increase HESC motility. MSCs were purchased from ATCC (BM-MSC-1) and cultured from the BM aspirate of a healthy female donor (BM-MSC-2). Indirect co-culture of MSCs and hTERT-immortalized HESCs via a transwell system studied the effect of the BM-MSC secretome on HESC proliferation, migration, and invasion. To study the effect of the MSC secretome on HESC gene expression, HESCs were exposed to the BM-MSC secretome via indirect co-culture for 24 h. Total RNA was extracted from HESCs for RNA sequencing (RNA-Seq). Differentially expressed genes (DEG) and significantly altered pathways were identified. Indirect co-culture of HESCs with BM- MSCs resulted in significant increase in HESC migration and invasion regardless of the source of MSCs. Effects on cellular proliferation varied among the BM-MSC source. Exposure of HESCs to the secretome of BM-MSCs changed the expression of 10,141 genes with FDR < 0.05. There was overlap among 4351 genes between HESCs exposed to BM-MSC-1 and BM-MSC-2, including upregulated expression of cell motility genes common to both BM-MSC exposures. Increased HESC motility by the secretome of BM-MSC appears to be mediated by paracrine and autocrine mechanisms, in part by modifying HESC gene expression. These data support the potential for leveraging the MSC secretome as a novel cell-free therapy in the treatment of disorders of endometrial regeneration.

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Cord Blood-Derived Mesenchymal Stem Cells with Distinct Growth Kinetics, Differentiation Potentials, Expression Profiles

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