Unknown,Transcriptomics,Genomics,Proteomics

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Perivascular Human Endometrial Mesenchymal Stem Cells Express Pathways Relevant to Self-Renewal, Lineage Specification, and Functional Phenotype

ABSTRACT: A population of endometrial cells displaying key properties of mesenchymal stem cells (eMSC) has been identified in human endometrium. eMSC co-express CD146 and PDGFRB surface markers, have a perivascular location, and likely represent the reservoir of progenitors giving rise to the endometrial stromal fibroblast lineage. Endometrial stromal cells isolated from 16 oocyte donors and 3 benign gynecologic surgery subjects were FACS sorted into four populations: CD146+/PDGFRB+ (eMSC); CD146+/PDGFRB- (endothelial cells); CD146-/PDGFRB+ (stromal fibroblasts); CD146-/PDGFRB- (mixed population) then subjected to gene expression analysis on Affymetrix Human Gene 1.0 ST arrays, and differentially expressed genes compared between eMSC, stromal fibroblast, and endothelial cell populations. Ninety-two genes were validated by multiplex quantitative RT-PCR on seventy of these sorted cell populations. Immunohistochemistry was used to verify the perivascular location of eMSCs.Principal component analysis and hierarchical clustering showed eMSC clustering discretely near stromal fibroblasts and separately from endothelial cells. eMSC expressed pericyte markers and genes involved hypoxia response, inflammation, proteolysis, and angiogenesis/vasculogenesis – all relevant to endometrial tissue breakdown and regeneration. Additionally, eMSC displayed distinct gene profiles for cell-cell communication and regulation of gene expression. Overall, the phenotype of the eMSC is that of a multipotent pericyte responsive to hypoxic, proteolytic, and inflammatory stimuli, able to induce angiogenesis, migrate and differentiate into lineage cells, and potentially respond to estradiol and progesterone. Identifying the pathways and gene families described herein in the context of the endometrial niche, will be valuable in understanding normal and abnormal endometrial development in utero and differentiation in adult uterus. The multipotent, perivascular endometrial mesenchymal stem cell has a “niche phenotype” of high Notch, TGFB, IGF, and Hedgehog and low canonical/non-canonical Wnt and EGF signaling. Oocyte donors with no known uterine pathology underwent endometrial biopsy at the time of oocyte retrieval, following comparable GnHR agonist downregulated ovarian stimulation protocols. Tissue was digested and stromal cells isolated and sorted based on expression of CD146 and PDGFRB. RNA was extracted and hybridized on Affymetrix microarrays. Resulting data were compared between sorted isolated cell populations.

ORGANISM(S): Homo sapiens

SUBMITTER: Linda Giudice

PROVIDER: E-GEOD-31152 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Perivascular human endometrial mesenchymal stem cells express pathways relevant to self-renewal, lineage specification, and functional phenotype.

Spitzer Trimble L B TL Rojas Angela A Zelenko Zara Z Aghajanova Lusine L Erikson David W DW Barragan Fatima F Meyer Michelle M Tamaresis John S JS Hamilton Amy E AE Irwin Juan C JC Giudice Linda C LC

Biology of reproduction 20120229 2

Human endometrium regenerates on a cyclic basis from candidate stem/progenitors whose genetic programs are yet to be determined. A subpopulation of endometrial stromal cells, displaying key properties of mesenchymal stem cells (MSCs), has been characterized. The endometrial MSC (eMSC) is likely the precursor of the endometrial stromal fibroblast. The goal of this study was to determine the transcriptome and signaling pathways in the eMSC to understand its functional phenotype. Endometrial stroma ...[more]

PMID: 22075475

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Project description:A population of endometrial cells displaying key properties of mesenchymal stem cells (eMSC) has been identified in human endometrium. eMSC co-express CD146 and PDGFRB surface markers, have a perivascular location, and likely represent the reservoir of progenitors giving rise to the endometrial stromal fibroblast lineage. Endometrial stromal cells isolated from 16 oocyte donors and 3 benign gynecologic surgery subjects were FACS sorted into four populations: CD146+/PDGFRB+ (eMSC); CD146+/PDGFRB- (endothelial cells); CD146-/PDGFRB+ (stromal fibroblasts); CD146-/PDGFRB- (mixed population) then subjected to gene expression analysis on Affymetrix Human Gene 1.0 ST arrays, and differentially expressed genes compared between eMSC, stromal fibroblast, and endothelial cell populations. Ninety-two genes were validated by multiplex quantitative RT-PCR on seventy of these sorted cell populations. Immunohistochemistry was used to verify the perivascular location of eMSCs.Principal component analysis and hierarchical clustering showed eMSC clustering discretely near stromal fibroblasts and separately from endothelial cells. eMSC expressed pericyte markers and genes involved hypoxia response, inflammation, proteolysis, and angiogenesis/vasculogenesis – all relevant to endometrial tissue breakdown and regeneration. Additionally, eMSC displayed distinct gene profiles for cell-cell communication and regulation of gene expression. Overall, the phenotype of the eMSC is that of a multipotent pericyte responsive to hypoxic, proteolytic, and inflammatory stimuli, able to induce angiogenesis, migrate and differentiate into lineage cells, and potentially respond to estradiol and progesterone. Identifying the pathways and gene families described herein in the context of the endometrial niche, will be valuable in understanding normal and abnormal endometrial development in utero and differentiation in adult uterus. The multipotent, perivascular endometrial mesenchymal stem cell has a “niche phenotype” of high Notch, TGFB, IGF, and Hedgehog and low canonical/non-canonical Wnt and EGF signaling.

Project description:During carcinoma progression, mesenchymal stromal cells (MSCs) become recruited to tumors and contribute to the pool of cancer-associated fibroblasts (CAFs). Sub-populations of CAFs, have diverse and incompletely understood effects on disease progression and resistance to therapy. Adipose stromal cells (ASCs), the MSCs from fat tissue increasingly recruited by carcinomas in the context of obesity, have been shown to support cancer progression. Here, the roles of perivascular CAFs expressing platelet-derived growth factor receptor beta (Pdgfrb) and of CAFs expressing non-glycanated decorin (ngDCN), a marker of ASCs, were analyzed. We used mice orthotopically grafted with pancreatic ductal adenocarcinoma (KPC) cells. Ablation of cells expressing thymidine kinase under the control of Pdgfrb promoter with ganciclovir resulted in suppression of primary pancreatic tumor growth. However, ablation Pdgfrb+ cells induced spontaneous metastases to the liver. For depletion of ngDCN+ cells, we used a hunter-killer peptide D-CAN, which has been previously reported to induce apoptosis of ASCs and CAFs in mouse models. Here, D-CAN also had a negative effect on pancreatic tumor growth, however, there was no significant effect on metastatic dissemination. Single cell RNA sequencing of tumors demonstrated that targeting of Pdgfrb+ and ngDCN+ stromal cells had distinct effects on sub-populations of CAFs. Endothelial cell and cancer cell survival was decreased by depletion of either Pdgfrb+ or ngDCN+ stroma. However, pathway analysis revealed that depletion of Pdgfrb+ and ngDCN+ stromal cells has different effects on the markers of cancer cell aggressiveness. D-CAN treatment, and to a less extent Pdgfrb+ cell ablation, suppressed macrophage M2 polarization and increased infiltration of cytotoxic T-lymphocytes. This observation, confirmed in the MyCap graft model of prostate cancer, suggested that ablation of ngDCN+ stroma should synergize with immune checkpoint inhibitors. We tested the effect of D-CAN on the efficacy of anti-PD-L1 antibody in the orthotopic KPC model. Compared to D-CAN and anti-PD-L1 antibody alone, combination treatment had a synergistic effect on tumor growth. Importantly, liver metastases were suppressed by the D-CAN / anti-PD-L1 antibody combination, but not by individual agents. We conclude that improved approaches to target mesenchymal stroma in tumors may be effective in combination with immunotherapy.

Project description:Hematopoietic stem cells (HSCs) primarily reside in the bone marrow where signals generated by stromal cells regulate their self-renewal, proliferation, and trafficking. Endosteal osteoblasts and perivascular stromal cells including endothelial cells3, CXCL12-abundant reticular (CAR) cells, leptin-receptor positive stromal cells, and nestin-GFP positive mesenchymal progenitors have all been implicated in HSC maintenance. However, it is unclear if specific hematopoietic progenitor cell (HPC) subsets reside in distinct niches defined by the surrounding stromal cells and the regulatory molecules they produce. CXCL12 (stromal-derived factor-1, SDF-1) regulates both HSCs and lymphoid progenitors and is expressed by all of these stromal cell populations. Here, we selectively deleted Cxcl12 from candidate niche stromal cell populations and characterized the effect on HPCs. Deletion of Cxcl12 from mineralizing osteoblasts has no effect on HSCs or lymphoid progenitors. Deletion of Cxcl12 from osterix-expressing stromal cells, which includes CAR cells and osteoblasts, results in constitutive HPC mobilization and a loss of B lymphoid progenitors, but HSC function is normal. Cxcl12 deletion in endothelial cells results in a modest loss of long-term repopulating activity. Strikingly, deletion of Cxcl12 in nestin-negative mesenchymal progenitors using Prx1-Cre is associated with a marked loss of HSCs, long-term repopulating activity, HSC quiescence, and common lymphoid progenitors. These data suggest that osterix-expressing stromal cells comprise a distinct niche that supports B lymphoid progenitors and retains HPC in the bone marrow, while expression of CXCL12 from stromal cells in the perivascular region, including endothelial cells and mesenchymal progenitors, support HSCs. Total of three samples of two groups analyzed. Replica samples of CXCL12-abundant reticular (CAR) cells from two CXCL12-GFP knock-in mice and a combined sample of PDGFRa+ Sca+ CD45- lineage- cells from three Prx1-Cre Rosa26Ai9/+ Cxcl12gfp/+ mice were used and analyzed.

Dataset Information

Perivascular Human Endometrial Mesenchymal Stem Cells Express Pathways Relevant to Self-Renewal, Lineage Specification, and Functional Phenotype

Publications

Perivascular human endometrial mesenchymal stem cells express pathways relevant to self-renewal, lineage specification, and functional phenotype.

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