Project description:This study uses microarray technology to examine the erythroid progenitor mRNA of patients with transfusion dependent β-thalassaemia and compare it to erythroid progenitor mRNA from healthy controls. We observed no statistical difference in gene expression between the groups following 7 days in culture. However, following 14 days in culture we observed differential expression of 161 genes. Haematopoietic cells from the peripheral blood of 6 β-thalassaemia patients and 6 healthy controls was grown in semi-solid media. After 7 and 14 days in culture cells of erythroid origin were isolated. Total RNA was isolated from these for microarray gene expression analysis
Project description:This study uses microarray technology to examine the erythroid progenitor mRNA of patients with transfusion dependent ?-thalassaemia and compare it to erythroid progenitor mRNA from healthy controls. We observed no statistical difference in gene expression between the groups following 7 days in culture. However, following 14 days in culture we observed differential expression of 277 genes. Haematopoietic cells from the peripheral blood of 6 ?-thalassaemia patients and 6 healthy controls was grown in semi-solid media. After 7 and 14 days in culture cells of erythroid origin were isolated. Total RNA was isolated from these for microarray gene expression analysis
Project description:This study uses microarray technology to examine the erythroid progenitor mRNA of patients with transfusion dependent β-thalassaemia and compare it to erythroid progenitor mRNA from healthy controls. We observed no statistical difference in gene expression between the groups following 7 days in culture. However, following 14 days in culture we observed differential expression of 161 genes.
Project description:This study uses microarray technology to examine the erythroid progenitor mRNA of patients with transfusion dependent β-thalassaemia and compare it to erythroid progenitor mRNA from healthy controls. We observed no statistical difference in gene expression between the groups following 7 days in culture. However, following 14 days in culture we observed differential expression of 277 genes.
Project description:Differential gene expression in erythroid progenitor cells from β-thalassaemia patients and healthy controls [Bioconductor/limma R analysis]
Project description:Reactivation of gamma-globin is considered a promising approach for the treatment of beta-thalassaemia and sickle cell disease. Therapeutic induction of gamma-globin expression is fraught with lack of suitable therapeutic targets. In order to identify new potential targets we analysed the changes in the proteome of human primary erythroid progenitor cells by treatment with decitabine, a known, yet not clinically safe, gamma-globin inducer. Significant differentially expressed proteins were identified which were involved in various biological pathways and functional categories.
Project description:SF3B1 knockdown in human CD34+ cells leads to increased apoptosis and cell cycle arrest of early-stage erythroid cells and generation of abnormally nucleated late-stage erythroblasts. RNA-seq analysis of SF3B1-knockdown erythroid progenitor CFU-E cells revealed altered splicing of an E3 ligase Makorin Ring Finger Protein 1 (MKRN1) and subsequent activation of p53 pathway.Decreased expression of genes involved in mitosis/cytokinesis pathway including polo-like kinase 1 (PLK1) was noted in SF3B1-knockdown polychromatic and orthochromatic erythroblasts comparing to control cells.
Project description:EKLF is a Krüppel-like transcription factor identified as a transcriptional activator and chromatin modifier in erythroid cells. EKLF-deficient (Eklf -/-) mice die at day 14.5 of gestation from severe anemia. In this study, we demonstrate that early progenitor cells fail to undergo terminal erythroid differention in Eklf -/- embryos. To discover potential EKLF target genes responsible for the failure of erythropoiesis, transcriptional profiling was performed with RNA from wild type and Eklf -/- early erythroid progenitor cells. These analyses identified significant perturbation of a network of genes involved in cell cycle regulation, with the critical regulator of the cell cycle, E2f2, at a hub. E2f2 mRNA and protein levels were markedly decreased in Eklf -/- early erythroid progenitor cells, which showed a delay in the G1-to-S-phase transition. Chromatin immunoprecipitation analysis demonstrated EKLF occupancy at the proximal E2f2 promoter in vivo. Consistent with the role of EKLF as a chromatin modifier, EKLF binding-sites in the E2f2 promoter were located in a region of EKLF-dependent DNase I sensitivity in early erythroid progenitor cells. We propose a model in which EKLF-dependent activation and modification of the E2f2 locus is required for cell cycle progression preceding terminal erythroid differentiation. RNA was isolated from flow-sorted early erythroid progenitors in 13.5 day old fetal livers from EKLF knock out mice (n=3 fetal livers) and wild-type control mice (n=3 fetal livers) for gene expression analysis
Project description:Erythroid progenitor BFU-Es are so-named based on their ability to generate in methylcellulose culture large colonies of erythroid cells that consist of “bursts” of smaller erythroid colonies derived from the later CFU-E Epo- dependent progenitors. “Early” BFU-E cells forming large BFU-E colonies presumably have higher capacities for self-renewal than do those forming small BFU-E colonies. In order to understand the mechanism underlying this heterogeneity, we conducted single cell transcriptome analysis on BFU-E cells purified from mouse embryos. Our analyses showed that there are two principal subgroups of mouse BFU-E cells and that the type III TGFβ receptor (TβRIII) is a potential marker that distinguishes “early” and “late” BFU-Es. Expression of TβRIII is correlated with that of GATA1, a gene encoding an erythroid transcription factor induced during the BFU-E to CFU-E transition. The mouse and human BFU-E sub populations (TßRIII10%lo) expressing the 10% lowest amount of surface TβRIII are indeed enriched for early BFU-Es, and are significantly more responsive to glucocorticoid stimulation, which promotes BFU-E self-renewal, as compared to the total BFU-E population. The TßRIII10%lo BFU-E subpopulation presumably represents earlier BFU-Es with maximal capacity for self-renewal. Consistent with this notion, signaling by the TGFβ receptor kinases RI and RII increases during the transition from early (TßRIII10%lo) to late (TßRIII10%hi) BFU-Es and then decreases in CFU-E cells. Blocking TGF-β signaling by receptor kinase inhibitors increase TßRIII10%lo BFU-E cell self-renewal and increases total erythroblast production, suggesting the use of this type of drug in treating Epo unresponsive anemias.