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. 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.

Project description:Brain perivascular cells have been recently identified as new mesodermal cell type of the human brain. These cells reside in the perivascular niche and were shown to have mesodermal and â to a lesser extend â tissue-specific differentiation potential. Mesenchymal stem cells (MSCs) are widely discussed for the use in cell therapy in many neurological disorders. Therefore it is of importance to better understand the âintrinsicâ MSC population of the human brain. Here we systematically characterized adult human brain-derived pericytes during in vitro expansion and differentiation and compared these cells to fetal and adult human brain-derived NSCs and adult human bone marrow derived MSCs. We found that adult human brain pericytes can be isolated from hippocampal as well as cortical white matter, are â in contrast to adult human NSCs â easily expandable in monolayer cultures and show high similarities to human bone marrow-derived MSCs both regarding surface marker expression and whole transcriptome analysis. Human brain pericytes differentiated only in negligible amounts into neuroectodermal cell types using various differentiation conditions but efficiently differentiated into mesodermal progenies. Thus bone marrow-derived MSCs resemble human brain pericytes and might be therefore very interesting for possible autologous NPC-based treatment strategies, cell therapeutic approaches of neurological diseases. For the gene expression microarray analysis we used the Affymetrix U133A chips The whole procedure was performed following the manufacturer's standard protocol (Affymetrix, Santa Clara, CA). For the data processing, normalization was calculated with the GCRMA (GC content corrected Robust Multi-array Analysis) algorithm. Data post-processing and graphics was performed with in-house developed functions in Matlab. 17 samples were analyzed fNSC, Neural Stem Cell, 2 replicates ANPC-hip, adult Neuroprogenitor - Cell Hippocampus, 3 replicates ANPC-wm, adult Neuroprogenitor Cell - White Matter, 3 replicates ABPMC-hip, adult Brain Perivascular Mesodermal Cell - Hippocampus, 3 replicates ABPMC-wm, adult Brain Perivascular Mesodermal Cell - White Matter, 3 replicates MSC, Mesenchymal Stem Cell, 3 replicates

Project description:Despite their key role in immunity our understanding of primary and secondary lymphoid stromal cell heterogeneity and ontogeny remains limited. Here, using genome-wide expression profiling and phenotypic and localization studies, we identify a functionally distinct subset of BP3-PDPN+PDGFRÎ²+/Î±+CD34+ stromal adventitial cells in both lymph nodes and thymus that is located within the perivascular niche surrounding PDPN-PDGFRÎ²+/Î±-Esam-1+ITGA7+ pericytes. In re-aggregate organ grafts adult CD34+ adventitial cells gave rise to multiple thymic and lymph node mesenchymal subsets including pericytes, FRC-, MRC- and FDC-like cells, the development of which was lymphoid environment dependent. During thymic ontogeny pericytes developed from a transient population of BP3-PDPN+PDGFRÎ²+/Î±+CD34-/lo anlage-seeding progenitors that subsequently up-regulated CD34 and we provide evidence suggesting that similar embryonic progenitors give rise to lymph node mesenchymal subsets. These findings extend the current understanding of lymphoid mesenchymal cell heterogeneity and highlight a role of the CD34+ vascular adventitia as a potential ubiquitous source of lymphoid stromal precursors in postnatal tissues. To comprehensively study the differences and similarities between mesenchymal stromal subsets in the thymus and lymph nodes, global gene expression analysis was performed on sorted PDPN-, BP-3-PDPN+ and BP-3+PDPN+ PDGFRb+ lymph node mesenchymal cells (LNMC) as well as PDPN- and BP-3-PDPN+ PDGFRb+ thymic mesenchymal cells (TMC) from 2 w old mice by microarray. Total RNA was prepared from TMC and LNMC (pooled inguinal, brachial and axillary LN) subsets sorted from 3 (TMC) and 10-11 (LNMC) 2 weeks old mice per experiment. Isolated RNA from 3 individual experiments was amplified and prepared for hybridization to the Affymetrix Mouse Gene 1.1 ST Array at a genomics core facility: Center of Excellence for Fluorescent Bioanalytics (KFB, University of Regensburg, Germany)

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