Project description:Hematopoiesis in adult mammals involves cognate interactions between developing hematopoietic cells and bone marrow stromal cell niches. SCF and CXCL12 play key roles in the maintenance of HSC, while early B cell differentiation requires CXCL12 and IL7. In this study, we characterized mouse BM stromal cells expressing IL7 by performing a transcriptomic analysis. We found that SCF, CXCL12 and IL7 were co-expressed by a unique peri-sinusoidal stromal cell subset.
Project description:Hematopoiesis in adult mammals involves cognate interactions between developing hematopoietic cells and bone marrow stromal cell niches. SCF and CXCL12 play key roles in the maintenance of HSC, while early B cell differentiation requires CXCL12 and IL7. In this study, we characterized mouse BM stromal cells expressing IL7 by performing a transcriptomic analysis. We found that SCF, CXCL12 and IL7 were co-expressed by a unique peri-sinusoidal stromal cell subset.
Project description:In the bone marrow, CXCL12 and IL7 are essential for B cell differentiation whereas hematopoietic stem cells (HSC) maintenance requires SCF and CXCL12. Previous studies, including ours, indicate that peri-sinusoidal stromal (PSS) cells are heterogeneous and that part of them express and are the main source of IL7. Our single cell qPCR and transcriptomic analysis demonstrate that SCF, CXCL12 and IL7 are co-expressed by a unique peri-sinusoidal stromal cell subset.
Project description:Fate decisions of haematopoietic stem cells (HSCs) to self-renew or differentiate in response to various demands are finely tuned by specialized microenvironments called “niches” in the bone marrow. Recent studies suggest that arterioles and sinusoids accompanied with distinct stromal cells marked by nerve/glial antigen 2 (NG2) and leptin receptor (LepR), compose distinct niches regulating quiescence and proliferation of HSCs, respectively. However, it remains unknown how the distinct niche cells differentially regulate the HSC functions. Here we show that effects of cytokines regulating HSC functions are dependent on the producing cell sources. Deletion of chemokine C-X-C motif ligand 12 (CXCL12) in NG2-cre targeted cells, which exclusively overlap with Nestin-GFP (Nes-GFP)+ stromal cells associated with arterioles and sinusoids, resulted in a robust reductions of HSCs in the bone marrow and massive mobilization. Deletion of CXCL12 from arteriolar NG2+ vascular smooth muscle cells caused a significant decrease of HSCs and altered HSC location in the marrow, while CXCL12 depletion from sinusoidal LepR+ cells did not reduce HSC numbers in the bone marrow. By contrast, deletion of stem cell factor (SCF) in LepR+ cells led to significant reductions in HSC numbers whereas SCF deletion in arteriolar NG2+ cells showed no effect on HSC numbers in the marrow. These results uncover the distinct contributions of cytokines derived from perivascular cells in separate vascular niches for HSC maintenance and mobilization. We sought to obtain comprehensive understanding of differences between peri-arteriolar and peri-sinusoidal niche cells by the present RNA-seq analysis.
Project description:Cell cycle quiescence is a critical feature contributing to haematopoietic stem cell (HSC) maintenance. Although various candidate stromal cells have been identified as potential HSC niches, the spatial localization of quiescent HSC in the bone marrow (BM) remains unclear. Here, using a novel approach that combines whole-mount confocal immunofluorescence imaging technique and computational modelling to analyse significant tridimensional associations among vascular structures, stromal cells and HSCs, we show that quiescent HSCs associate specifically with small arterioles that are preferentially found in endosteal BM. These arterioles are ensheathed exclusively by rare Nestin-GFP-peri/NG2+ pericytes, distinct from sinusoid-associated Nestin-GFP-retic/LepR+ cells. The present RNA-seq study sought to obtain a comprehensive understanding of the differences between the two distinct HSC cellular niches. mRNA profiles of sorted Nestin-GFP-peri and -GFP-retic bone marrow stromal cells were generated from pooled mice in triplicate by Illumina HiSeq 2000 sequencing.
Project description:RNA sequencing of two stromal cell subsets of the bone marrow, namely CXCL12-abundant reticular cells (CARc) and sinusoidal endothelial cells (SECs) isolated from untreated mice (day 0) and mice infected with Lymphocytic Choriomeningitis Virus cl-13, 56 days after infection (day 56)
Project description:RNA-sequencing of a longitudinal comparison of four principal mesenchymal and endothelial stromal cell types (CXCL12-abundant reticular cells, PDGFR-α+ Sca-1+, sinusoidal and arterial endothelial cells), isolated from early postnatal, adult and aged mice. Additional transcriptional profiling of the response of CXCL12-abundant reticular cells and sinusoidal endothelial cells to infection-mimicking agents.
Project description:To analyze the role of VEGF-C in the adult bone marrow, we analyzed the bone marrow stromal fraction on LepR+ cells and endothelial cells to further dissect the source of VEGF-C in the adult bone marrow.
Project description:c-kit signaling plays pivotal roles in regulating the self-renewal and/or differentiation of many adult stem cells, such as hematopoietic stem cells. However, it remains controversial whether c-kit is expressed by and contribute to skeletal stem cells (SSCs). To test this, we lineage-traced c-kit+ cells and investigated the physiological importance of c-kit+ cells and c-kit signaling in bone development. We found that c-kit was not expressed by postnatal SSCs, but by fetal SSC precursors at the growth cartilage. Lineage-tracing of fetal c-kit+ cells marked approximately 20% of Lepr+ bone marrow stromal cells, generating nearly half of all osteoblasts in adult bone marrow. Disruption of mTOR signaling in c-kit+ cells impaired bone formation. Conditional deletion of Kitl (c-kit ligand, also known as Scf) from fetal, but not adult bone marrow stromal cells increased bone formation. Together, our work identified c-kit+ SSC precursors as an important source of bones formed during development. The osteogenic differentiation of these cells is temporally inhibited by Kitl from the bone marrow microenvironment.