Project description:Global gene expression analysis of human embryoid body (hEB) differentiations during differentiation in serum free conditions in the presence of BMP4, VEGF, and FGF2 growth factors A total of 5 samples are analyzed. Each sample is taken at each time point. There is no replicate. H1_hESC_day0 sample is used as control to compare to other samples Keywords: Cell differentiation Cell differentiation compares to undifferentiated status in a time-course of 3-, 5-, 7- and 10-day without replicates.
Project description:The derivation of functional, transplantable HSCs from an pluripotent stem cells in vitro holds great promise for clinical therapies, but is unachieved. In order to achieve full functionality of HSCs, it is vital to determine the extent to which PSCs can currently be differentiated to the HSC program in vitro and identify the remaining dysregulated genetic pathways. Microarrays were used to compare the transcritomes of ESC-derived immunophenotypic HSPCs to endogenous HSPCs from various stages of development to determine the programs important for human HSC development and function, and which programs were lacking in ESC-derived hematopoietic cells. CD34+CD38-CD43+CD90+ HSPCs were sorted from human placenta and embryoid bodies, and CD34+CD38-CD45+CD90+ HSPCs sorted from fetal liver and embryoid bodies co-cultured on OP9-M2 stroma, the RNA was extracted, library created and hybridized to the Affymetrix microarray
Project description:Global gene expression analysis of (a) human embryonic stem cells, (b) adult fibroblasts with and without nucleofection of SOKM, and ( c ) CD34+ cord blood cells at various time points during induction of pluripotency with SOKM, with or without co-culture with bone marrow stromal cells (BMSC). Total RNA was harvested from adult fibroblasts, adult fibroblasts three days after nucleofection with 4F (SOKM), Day -3 unstimulated cord blood (CB) cells, Day 0 growth factor stimulated CB cells, Day +3 CB cells with or without nucleofection at Day zero and with or without Day zero to Day +3 co-culture with bone marrow stromal cells (BMSC), Day 23 CB cells nucleofected on day zero and co-cultured on BMSC from Day Zero to Day +3, and undifferentiated H9 human embryonic stem cells. All experimental conditions repeated in three independent trials, and each replicate at each condition analyzed on an individual microarray.
Project description:This SuperSeries is composed of the following subset Series: GSE35027: Global gene expression analysis of human embryonic stem cells, adult fibroblasts , and CD34+ cord blood (CB) cells before, during, and afer their episomal induction of pluripotency GSE35028: Global gene expression analysis of pluripotent cell lines and corresponding starting donor source cells Refer to individual Series
Project description:We used microarrays to examine the impact of AF1q/MLLT11 on the gene expression profile of CD34+CD45RA-Lin- and CD34+CD45RA+Lin- HPCs isolated from umbilical cord blood CD34+CD45RA-Lin- and CD34+CD45RA+Lin- HPCs correspond respectively to early multipotent and lympho-granulo-macrophagic precursors. A2M is a nuclear mutant-derivative of AF1q/MLLT11 CD34+CD45RA-Lin- and CD34+CD45RA+Lin- HPCs were FACS-sorted, exposed to A2M or control vectors, sorted based on GFP expression, and subjected to global gene expression analysis 72 hrs later.
Project description:In order to identify cells expressing RUNX1c during hematopoietic differentiation of human embryonic stem cells (hESCs), we targeted GFP downstream of the RUNX1 distal promoter. GFP was observed from 10-25 days of embryoid body differentiation and accurately mirrored expression of the endogenous RUNX1c isoform. GFP was restricted to CD45+ hematopoietic cells and GFP+CD34+ cells were highly enriched for progenitor cells. Appearance of the first wave of hematopoietic blast colony forming cells (Bl-CFC) in d3-4 embryoid bodies, antedated the expression of RUNX1c. These results confirm a role for RUNX1c in marking a second wave of hematopoietic progenitor cells in differentiating hESCs. The purpose of the cord blood samples was to perform comparison of them to the gene expression profiles of the Runx fractions - as the cord blood represents a heterogeneous mix of hematopoietic stem cells and multipotent progenitors cells.
Project description:RNA-seq expression analysis of transcripts encoding proteasome subunits in human CD34+ cord blood cell-derived megakaryocytes and mouse bone marrow-derived megakaryocytes. Analysis of transcript expression in human CD34+ cord blood cell-derived megakaryocytes and mouse bone marrow-derived megakaryocytes.
Project description:We investigated the role of amino acid to maintain HSC function. To identify essential amino acids for HSCs, CD34-KSL cells were cultured in single amino acids deficient medium. And cultured cells were transplanted into lethally irradiated mice. Then, the donor chimerism and lineage contribution was estimated. Surprisingly, HSC proliferation was prevented in valine and cysteine deficient medium in vitro. Donor cells cultured in these medium were also not engrafted. To elucidate the effects and influences of cysteine and valine in HSCs, we performed global gene expression profiling experiments by RNA-sequencing analysis. Gene sets categorized with cell cycle, mitosis, cell division or DNA replication were strongly down-regulated in both valine- or cysteine-depleted conditions These results imply distinctive amino-acid metabolism involved in HSC division. Gene expression profiles of ten thousand HSCs cultured in cysteine or valine deficient medium for 24 hours were compared with that of HSCs cultured in complete medium by using RNA-sequencing analysis
Project description:Epigenetic mechanisms including histone modifications have emerged as important factors influencing cell fate determination. The functional role of H3K4 methylation, however, remains largely unclear in the maintenance and differentiation of hematopoietic stem/progenitor cells (HSC/HPCs). Here we show that DPY30, a shared core subunit of the SET1/MLL family methyltransferase complexes and a facilitator of their H3K4 methylation activity, is important for ex vivo proliferation and differentiation of human CD34+ HPCs. DPY30 promotes HPC proliferation by directly regulating the expression of genes critical for cell proliferation. Interestingly, while DPY30 knockdown (KD) in HPCs impaired their differentiation into the myelomonocytic lineage, it potently promoted hemoglobin production and affected the kinetics of their differentiation into the erythroid lineage. In an in vivo model, we show that morpholino-mediated dpy30 KD resulted in severe defects in the development of the zebrafish hematopoietic system, which could be partially rescued by co-injection of dpy30 mRNA. Taken together, our results establish a critical role of DPY30 in the proliferation and appropriate differentiation of hematopoietic progenitor cells as well as in animal hematopoiesis. Finally, we also demonstrate a crucial role of DPY30 in the growth of several MLL1-fusion-mediated leukemia cell lines. Total RNAs from control (scr) or knockdown (hD2, hD5) cells before and after culturing under condition promoting myelomonocytic differentiation were subjected to Illumina microarray analyses.