ATACseq measurements during cord blood-derived human erythropoiesis
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ABSTRACT:
PROVIDER: PRJNA565446 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR10119482_1.fastq.gz | Fastqsanger.gz | |||
| SRR10119482_2.fastq.gz | Fastqsanger.gz | |||
| SRR10119483_1.fastq.gz | Fastqsanger.gz | |||
| SRR10119483_2.fastq.gz | Fastqsanger.gz | |||
| SRR10119484_1.fastq.gz | Fastqsanger.gz |
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Project description:CD34+ cells were subjected to ex vivo erythropoiesis according to the protocol described in Palii et al. (doi: 10.3791/2813. J Vis Exp. 53, 2011). Samples were harvested at different time points and analyzed by ATACseq.
2020-04-23 | GSE137446 | GEO
Project description:The characteristics of global gene expression patterns during umbilical cord blood (UCB)-CD34+ stem cell-derived erythropoiesis are not clearly elucidated. In this study, UCB-stem cells were grown in liquid culture as a model for this process. We observed a high proliferative capacity of UCB-stem cells producing over 10 billion viable cells in culture. Normal erythroid maturation was confirmed by increased expression of CD71 and CD235a biomarkers. Furthermore, the gamma- to beta-globin gene switch was observed around day 45 indicating extended gamma-globin gene expression in fetal erythroid cells. To study global gene expression patterns, microarray analysis was performed on days 21, 42, 49 and 56. Forty-five transcription factors were silenced during the culture period (Profile-1) including CUX1 and HES5 among others. Conversely, 30 transcription factors were activated by day 56 (Profile-2) such as KLF1, GATA1, and MAFB. Both datasets were analyzed further using the MIMI Cytoscape platform which defined transcription factor networks around KLF4 and GATA2 from Profile-1 genes and KLF1 and GATA1 for Profile-2 genes. The characteristics of UCB-CD34+ stem cells including high proliferative capacity and prolonged gamma-globin expression combined with novel transcription factor networks suggest novel mechanisms of fetal UCB-stem cell-derived erythropoiesis. Cd34 cell from umbilical cord blood were grown in three independent cultures using the one-phase protocol. Cells were cultured in aMEM containing 30% fetal bovine serum, 1% deionized BSA with penicillin (100 U/mL) and streptomycin (0.1 mg/mL) at 37°C and 5% CO2. The following growth factors were added on day 0: stem cell factor (50 ng/mL), interleukin-3 (10 ng/mL) and erythropoietin (4 U/mL). Approximately 1.5 million cells were harvested from each culture (triplicate samples) on day 21, 42, 49 and 56 for microarray analysis on the whole-genome Illumina HumanWG-12 V4 Expression BeadChip.
2013-08-02 | E-GEOD-49438 | biostudies-arrayexpress
Project description:Erythropoiesis from pluripotent stem cells is very complex, and mirrors that observed in developmental erythropoiesis, with primitive and definitive waves. For a better understanding of their potential for their use in blood transfusion, they were compared to erythropoiesis from cord derived CD34 haematopoietic stem cells. We used microarrays to detail the global programme of gene expression underlying different stages of erythropoiesis from each source, pluripotent or haematopoietic stem cells.
2016-12-13 | GSE92285 | GEO
Project description:Erythropoiesis in mammals replenishes the circulating red blood cell (RBC) pool from hematopoietic stem/progenitor cells (HSPCs). Two distinct erythropoietic programs have been described. In the first trimester, hematopoietic precursors in the fetal yolk sac follow a primitive pattern of erythropoiesis. However, in the second trimester, hematopoietic stem cells (HSCs) from the fetal liver and later from the bone marrow differentiate by a definitive program of erythropoiesis to yield enucleated erythrocytes. RBCs can also be derived from human induced pluripotent stem cells (hiPSCs) and can express many of the red cell proteins required for normal erythrocyte function, presaging in vitro RBC production for clinical use. However, expansion and enucleation from hiPSCs is less efficient than with erythroblasts (EBs) derived from adult or cord blood progenitors. We hypothesized that substantial differential gene expression during erythroid development from hiPSCs compared to that from adult blood or cord blood precursors could account for these hitherto unexplained differences in proliferation and enucleation. We have therefore grown EBs from human adult and cord blood progenitors and from hiPSCs. Gene expression during erythroid culture from each erythroblast source was analyzed using algorithms designed to cluster co-expressed genes in an unsupervised manner and the function of differentially expressed genes explored by gene ontology. Using these methods we identify specific patterns of gene regulation for adult- and cord- derived EBs, regardless of the medium used, that are substantially distinct from those observed during the differentiation of EBs from hiPSC progenitors which largely follows a pattern of primitive erythropoiesis. A total of 74 samples were analyzed. Our primary goal was to compare erythropoiesis from hiPSC-derived EBs with that from adult or cord blood PBMC-derieved EBs. We wanted to provide a high resolution atlas for investigators dissecting the programs of erythroid development and highlight the disparities between erythropoiesis in vitro from diverse stem/progenitor cell origins. Erythroblasts were grown in vitro from CD34+ erythroid progenitors isolated from Human Adult PBMCs (51 AB-EB samples), Human Cord PBMCs (10 CB-EB samples) or human induced pluripotent stem cells (13 hiPSC-EB samples). Three culture media were used for AB-EBs: Standard Erythroid Medium (SEM) containing 2% FBS (SEM-F), SEM in which FBS was replaced with 1% BSA (SEM-B) and SEM optimized for culture of EBs from hiPSCs (SEM-i). After culturing for 4,7,10,12, or 14 days in SEM-F, AB-EBs media were harvested in triplicate and sorted using flow cytometry (FACS) into enriched homogeneous erythroblast populations based on cell surface expression of CD36 (used for days 4-7), CD71 and CD235a. Magnetic beads specific for CD34 were used to isolate CD34+ day 0 progenitors. Magnetic beads were also used to isolate AB-EBs grown in SEM-F and SEM-i after 7 days (CD71+) and 14 days (CD235a beads). AB-EBs were also cultured in SEM-B for 4,7, or 14 days and sorted by FACS. CB-EBs were cultured in SEM-F for 7 or 14 days, and sorted by FACS as for AB-EBs. HiPSC-EBs were cultured in SEM-i for 7 or 14 days, and isolated using magnetic beads as fof AB-EBs. Total RNA was isolated from all these populations in triplicate (with a few exceptions), and also at day zero from CD34+ AB-EB progenitors, CB-EB progenitors and hiPSCs using magnetic beads. Additionally, CD31+ day zero hiPSCs were isolated using magnetic beads. RNA was extracted using MiRVana (Life Technologies) and DNA removed using Turbo DNA-Free (Life Technologies). The microarray target was prepared from 100ng total RNA for hybridization to Affymetrix GeneChip Human Transcriptome 2.0 ST microarrays (HTA2) using the Ambion WT protocol (Life Technologies) and Affymetrix labelling and hybridization kits (Affymetrix). Labelled DNA mean yield was 13.5 μg (minimum: 8.5 μg; maximum: 20.5 μg). HTA2 arrays were hybridized with 5 μg of labelled DNA. The Affymetrix GeneChip Fluidics Station 450 was used to wash and stain the arrays with streptavidin–phycoerythrin, according to the standard protocol for eukaryotic targets (IHC kit, Affymetrix). Arrays were scanned with an Affymetrix GeneChip scanner 3000 at 570nm. Intensity values were determined using GeneChip Operating Software (Affymetrix)and normalized by the Robust Multiarray Average algorithm using Affymetrix Expression Console software. Statistical analysis of differential expression was conducted using the Linear Models for Microarray Data package from the Bioconductor suite in R (www.bioconductor.org). The B values, p-values, and fold changes were used to select differentially expressed (DE) genes reaching a minimum linear expression value of 100 in all replicates of at least one sample group (p ≤ 0.01, fold change (FC) ≥ 2, B > 2.945). Principal component analysis (PCA) was conducted and displayed using Python packages. Hierarchical clustering (HC) analysis was performed using Python SciPy. Heat maps were generated using Python matplotlib. More advanced consensus clustering was conducted using SMART and Bi-CoPaM algorithms in MATLAB, using Biclustering from the biclust package in R. Biopython to fetch 1kb of genomic DNA sequence upstream of the transcriptional start sites (TSS) of each gene in the cluster. MEME suite was used for transcription factor binding site (TFBS) analysis, seeking homology with motifs in the Jolma database. Differentially-expressed genes were examined for enriched functional ontologies using GeneCoDis. Expression profiles of selected genes were validated by QPCR. Gene expression profiles (GEPs) of AB-EBs in SEM-F were as expected for erythroid genes and similar to those observed for CB-EBs in SEM-F. The media used had little effect on AB-EB GEPs. However, substantial robust and specific GEPs were observed in hiPSC-EBs which may, at least in part, underlie reduced proliferation and enucleation of erythroid cells from hiPSCs. These include genes involved in autophagy and cell cycle.
2016-10-05 | GSE66260 | GEO
Project description:The characteristics of global gene expression patterns during umbilical cord blood (UCB)-CD34+ stem cell-derived erythropoiesis are not clearly elucidated. In this study, UCB-stem cells were grown in liquid culture as a model for this process. We observed a high proliferative capacity of UCB-stem cells producing over 10 billion viable cells in culture. Normal erythroid maturation was confirmed by increased expression of CD71 and CD235a biomarkers. Furthermore, the gamma- to beta-globin gene switch was observed around day 45 indicating extended gamma-globin gene expression in fetal erythroid cells. To study global gene expression patterns, microarray analysis was performed on days 21, 42, 49 and 56. Forty-five transcription factors were silenced during the culture period (Profile-1) including CUX1 and HES5 among others. Conversely, 30 transcription factors were activated by day 56 (Profile-2) such as KLF1, GATA1, and MAFB. Both datasets were analyzed further using the MIMI Cytoscape platform which defined transcription factor networks around KLF4 and GATA2 from Profile-1 genes and KLF1 and GATA1 for Profile-2 genes. The characteristics of UCB-CD34+ stem cells including high proliferative capacity and prolonged gamma-globin expression combined with novel transcription factor networks suggest novel mechanisms of fetal UCB-stem cell-derived erythropoiesis.
2013-08-02 | GSE49438 | GEO
Project description:In order to revealed the multiple protein functional modules and kinases networks of human early erythropoiesis. We isolated the CD34+ cells drived from Human umbilical cord blood samples and induced to undergo erythropoiesis in vitro. Then, the cultured erythroid cells were sorted with FACS and sampled for proteome and phosphoproteome analysis.
2022-08-03 | PXD035785 |
Project description:Studies of human erythropoiesis have relied, for the most part, on the in vitro differentiation of hematopoietic stem and progenitor cells (HSPC) from different sources. Here, we report that despite the common core erythroid program that exists between cord blood- and peripheral blood-HSPC induced towards erythroid differentiation in vitro, significant functional differences exist. We undertook a comparative analysis of human erythropoiesis using these two different sources of HSPC and differentiated them in vitro. We observed that cells derived from cord blood proliferate 4.5 times more than cells derived from peripheral blood. However, these cells present a delay in their differentiation pattern due to increased quantities of progenitors, notably CFU-E. Using our method of immunophenotyping for the study of erythroid progenitors, we document the presence and maintenance of a specific population in peripheral blood-derived erythroid progenitors. This population, defined as IL3R-GPA-CD34+CD36+, has the ability to form both BFU-E and CFU-E colonies in colony-forming assays, reflecting a higher potential. To further understand the differences between cord blood- and peripheral blood- HSPC, we sorted all stages of erythropoiesis from both sources and compared their transcriptome. We document differences at the CD34, BFU-E, poly- and orthochromatic stages. Among the genes presenting the highest differences in expression, many are involved in the regulation of the cell cycle and autophagy. Altogether, our studies provide a qualitative and quantitative comparative analysis of human erythropoiesis and highlight functional differences, critical to our understanding of the impact of the developmental origin of HSPCs on erythroid differentiation.
2017-12-28 | GSE107218 | GEO
Project description:Cord blood DNA methylation is associated with numerous health outcomes and environmental exposures. Whole cord blood DNA reflects all nucleated blood cell types, while centrifuging whole blood separates red blood cells by generating a white blood cell buffy coat. Both sample types are used in DNA methylation studies. Cell types have unique methylation patterns and processing can impact cell distributions, which may influence comparability. To evaluate differences in cell composition and DNA methylation between buffy coat and whole cord blood samples, cord blood DNA methylation was measured with the Infinium EPIC BeadChip (Illumina) in 8 individuals, each contributing buffy coat and whole blood samples.
2018-01-01 | GSE103189 | GEO
Project description:Erythroblasts from human cord blood
| PRJNA544314 | ENA