Similar Datasets
Project description:CD244 expression represents functional decline of murine hematopoietic stem cells after in vitro culture
| PRJNA681786 | ENA
Project description:Understanding the molecular regulation of hematopoietic stem and progenitor cell (HSPC) engraftment is paramount to improving transplant outcomes. To discover novel regulators of HSPC repopulation, we transplanted >1,300 mice with shRNA-transduced HSPCs within 24 h of isolation and transduction to focus on detecting genes regulating repopulation. We identified 17 regulators of HSPC repopulation: Arhgef5, Armcx1, Cadps2, Crispld1, Emcn, Foxa3, Fstl1, Glis2, Gprasp2, Gpr56, Myct1, Nbea, P2ry14, Smarca2, Sox4, Stat4, and Zfp251. Knockdown of each of these genes yielded a loss of function, except in the cases of Armcx1 and Gprasp2, whose loss enhanced hematopoietic stem cell (HSC) repopulation. The discovery of multiple genes regulating vesicular trafficking, cell surface receptor turnover, and secretion of extracellular matrix components suggests active cross talk between HSCs and the niche and that HSCs may actively condition the niche to promote engraftment. We validated that Foxa3 is required for HSC repopulating activity, as Foxa3(-/-) HSC fails to repopulate ablated hosts efficiently, implicating for the first time Foxa genes as regulators of HSPCs. We further show that Foxa3 likely regulates the HSC response to hematologic stress. Each gene discovered here offers a window into the novel processes that regulate stable HSPC engraftment into an ablated host.
| S-EPMC4813668 | biostudies-literature
Project description:We report a serum-free, 3D murine artificial thymic organoid (M-ATO) system that mimics normal murine thymopoiesis with the production of all T cell stages, from early thymic progenitors to functional single-positive (CD8SP and CD4SP) TCRαβ and TCRγδ cells. RNA sequencing aligns M-ATO-derived populations with phenotypically identical primary thymocytes. M-ATOs initiated with Rag1-/- marrow produce the same differentiation block as seen in the endogenous thymus, and Notch signaling patterns in M-ATOs mirror primary thymopoiesis. M-ATOs initiated with defined hematopoietic stem cells (HSCs) and lymphoid progenitors from marrow and thymus generate each of the downstream differentiation stages, allowing the kinetics of T cell differentiation to be tracked. Remarkably, single HSCs deposited into each M-ATO generate the complete trajectory of T cell differentiation, producing diverse TCR repertoires across clones that largely match endogenous thymus. M-ATOs represent a highly reproducible and efficient experimental platform for the interrogation of clonal thymopoiesis from HSCs.
| S-EPMC7727762 | biostudies-literature
Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
2013-12-03 | E-GEOD-40800 | biostudies-arrayexpress
Project description:Hematopoiesis takes place in the bone marrow and is supported by a complex cellular and molecular network in the bone marrow microenvironment. Commonly used models of the human bone marrow microenvironment include murine models and two-dimensional and three-dimensional tissue cultures. While these model systems have led to critical advances in the field, they fail to recapitulate many aspects of the human bone marrow. This has limited our understanding of human bone marrow pathophysiology and has led to deficiencies in therapy for many bone marrow pathologies such as bone marrow failure syndromes and leukemias. Therefore, we have developed a modular murine bone marrow microenvironment-on-chip using a commercially available microfluidic platform. This model includes a vascular channel separated from the bone marrow channel by a semi-porous membrane and incorporates critical components of the bone marrow microenvironment, including osteoblasts, endothelial cells, mesenchymal stem cells, and hematopoietic stem and progenitor cells. This system is capable of maintaining functional hematopoietic stem cells in vitro for at least 14 days at frequencies similar to what is found in the primary bone marrow. The modular nature of this system and its accessibility will allow for acceleration of our understanding of the bone marrow.
| S-EPMC9252162 | biostudies-literature
Project description:Hematopoietic stem and progenitor cells (HPCs) can be maintained in vitro, but the vast majority of their progeny loses stemness during culture. In this study, we compared DNA-methylation (DNAm) profiles of freshly isolated and culture-expanded HPCs. Culture conditions of CD34(+) cells - either with or without mesenchymal stromal cells (MSCs) - had relatively little impact on DNAm, although proliferation is greatly increased by stromal support. However, all cultured HPCs - even those which remained CD34(+) - acquired significant DNA-hypermethylation. DNA-hypermethylation occurred particularly in up-stream promoter regions, shore-regions of CpG islands, binding sites for PU.1, HOXA5 and RUNX1, and it was reflected in differential gene expression and variant transcripts of DNMT3A. Low concentrations of DNAm inhibitors slightly increased the frequency of colony-forming unit initiating cells. Our results demonstrate that HPCs acquire DNA-hypermethylation at specific sites in the genome which is relevant for the rapid loss of stemness during in vitro manipulation.
| S-EPMC3842544 | biostudies-literature
Project description:Proliferation of HPSCs in vitro can promote its broad clinical therapeutic use. For in vitro co-culture, interaction between the stem cell and feeder cell as well as their spatial position are essential. To imitate the natural microenvironment, a 3D engineered scaffold for CD34+ cells co-culture was established via 3D bioprinting. Herein, the concentration of hydrogel and the ratio of two kinds of cells were optimized. Flow cytometry, real time PCR and RNA-seq technology were applied to analyze the effect of the engineered scaffold on expanded cells. After 10 days co-culture with the engineered scaffold, the expansion of CD34+CD38- cells can reach 33.57-folds and the expansion of CD34+CD184+ cells can reach 16.66-folds. Result of PCR and RNA-seq indicates that the CD34+ cells in 3D group exhibited a tendency of interaction with the engineered scaffold. Compared to 2D co-culture, this customizable 3D engineered scaffold can provide an original and integrated environment for HPSCs growth. Additionally, this scaffold can be modified for different cell co-culture or cell behavior study.
| S-EPMC7359311 | biostudies-literature
Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
2015-11-20 | E-GEOD-66382 | biostudies-arrayexpress
Project description:Isolation of long-term reconstituting hematopoietic stem cell (HSC) has been possible by utilizing flow cytometry with a combination of multiple antibodies against cell surface markers. However those cell surface phenotypes do not represent functional HSC after in vitro culture. We compared gene expression profiling of phenotypic HSC (CD48-KSL cells) before and after in vitro culture, and discovered that cultured HSC express mast cell-related genes including Cd244. After in vitro culture, phenotypic HSC can be divided into CD244- and CD244+ subpopulations, and only CD244- cells that have low mast cell gene expression and maintain HSC-related genes sustain reconstitution potential. Induction of CD244 may partially be triggered by induction of endoplasmic reticulum (ER) stress, as chemically induced ER stress signal increased CD244+ subpopulation while ER stress suppression using a molecular chaperone, TUDCA, decreased CD244+ population, which were correlated to the output of reconstitution assay. These data suggest CD244 positivity is a potent marker to exclude non-functional HSC after in vitro culture thereby useful to elucidate mechanism of functional decline of HSC during ex vivo treatment.
2021-11-01 | GSE162400 | GEO
Project description:Aging of the hematopoietic stem cell (HSC) compartment is characterized by lineage bias and reduced stem cell function, the molecular basis of which is largely unknown. Using single-cell transcriptomics, we identified a distinct subpopulation of old HSCs carrying a p53 signature indicative of stem cell decline alongside pro-proliferative JAK/STAT signaling. To investigate the relationship between JAK/STAT and p53 signaling, we challenged HSCs with a constitutively active form of JAK2 (V617F) and observed an expansion of the p53-positive subpopulation in old mice. Our results reveal cellular heterogeneity in the onset of HSC aging and implicate a role for JAK2V617F-driven proliferation in the p53-mediated functional decline of old HSCs.
| S-EPMC5457484 | biostudies-literature