CD34+ treatment with MicroVescicles
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Vittorio Simeon
PROVIDER: E-MTAB-3576 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:Mononuclear cells were isolated from umbilical cord blood (UCB) using Lymphoprep sucrose-gradient centrifugation (1.077 g/ml, Nycomed Pharma, Oslo, Norway). Immunomagnetic cell separation, using magnetic beads coated with CD34 antibodies (Miltenyi Biotec, Gladbach, Germany), was performed to isolate CD34-positive hematopoietic progenitor cells (CB-CD34+ cells). To generate retrovirus, bicistronic retroviral DNA constructs were used expressing the ETV6-RUNX1 or AML1-ETO fusion gene and enhanced Green Fluorescent Protein (eGFP). As a control, a construct expressing only eGFP was used. Both vectors consisted of a pMSCV promoter region, an internal ribosomal entry site and an ampicillin resistance cassette. HEK293T cells were co-transfected with these constructs and second-generation retroviral packaging vectors using XtremeGENE 9 tranfection reagents (Roche, Basel, Switzerland). Viral particles were collected in IMDM 48 hours after transfection. CD34+ haematopoietic progenitors, pre-cultured overnight as described above, upon which cells were divided in two fractions. One fraction was transduced with ETV6-RUNX1-IRES-eGFP or AML1-ETO-IRES-eGFP, while the other fraction was transduced with control EV-IRES-eGFP. Transductions were performed with fresh retrovirus in retronectin (Takara, Otsu, Japan) coated wells. After sorting, DAPI- CD34+ GFP+ CB-CD34+ cells were lysed and RNA was extracted using Nucleospin RNA XS extraction columns according to manufacturer’s protocol (Macherey-Nagel, Düren, Germany). Quality of RNA was determined by on-chip-electrophoresis using a RNA Pico Chip according to manufacturer’s protocol (Agilent Technologies, Santa Clara, CA, USA). RNA Integrity scores (RIN) were higher than 8 for all samples. RNA was subsequently linearly amplified using the Nugen WT-Amplification™ pico system (Nugen, San Carlos, CA, USA). This system is based on RNA-dependent DNA polymerase activity and was previously reported to be most suitable for amplification and gene expression of picograms of input RNA.
2019-01-01 | E-MTAB-3466 | biostudies-arrayexpress
Project description:Although it has been shown that HIF1 and 2 fulfill essential roles within the hematopoietic system and in the regulation of HSC fate, little is currently known about the specific mechanisms that are involved. We identified transcriptome changes induced by hypoxia, constitutively active HIF1(P402/564) and HIF2(P405/531) in human cord blood CD34+ cells. Thus, we were able to identify common hypoxia-HIF1-HIF2 gene signatures, but we also identified specific target genes that were exclusively regulated by HIF1, HIF2 or hypoxia. CB CD34+ cells were isolated by Miltenyi miniMACS column. Cells were prestimulated in HPGM with 100 ng/ml KITL, FLT3L and TPO for 3 days after which cells were placed either at normoxia or hypoxia (1% O2 for an additional 24 hrs). 5 independent CB CD34+ batches were used and isolated RNA was combined and used for Illumina beadhchip arrays HT12 v4
2014-03-01 | E-GEOD-54661 | biostudies-arrayexpress
Project description:Although it has been shown that HIF1 and 2 fulfill essential roles within the hematopoietic system and in the regulation of HSC fate, little is currently known about the specific mechanisms that are involved. We identified transcriptome changes induced by hypoxia, constitutively active HIF1(P402/564) and HIF2(P405/531) in human cord blood CD34+ cells. Thus, we were able to identify common hypoxia-HIF1-HIF2 gene signatures, but we also identified specific target genes that were exclusively regulated by HIF1, HIF2 or hypoxia. CB CD34+ cells were isolated by Miltenyi miniMACS column. Cells were prestimulated in HPGM with 100 ng/ml KITL, FLT3L and TPO for 48 hrs. Cells were transduced with control pRRL-IRS2-EGFP lentiviral vectors or vectors expressing HIF1α(P402A,P564A) or HIF2α(P405A,P531A) in one or two rounds of 12 hrs each. 24 hrs later transduced cells were sorted after which RNA was isolated. 5 independent CB CD34+ batches were isolated, transduced and sorted, and isolated RNA was combined and used for Illumina beadhchip arrays HT12 v4
2014-03-01 | E-GEOD-54662 | biostudies-arrayexpress
Project description:The prognosis of infant B-cell acute lymphoblastic leukemia (iB-ALL) remains dismal, especially in patients harboring the MLL-AF4 (KTM2A-AFF1) rearrangement, which arises prenatally in early hematopoietic stem/progenitor cells (HSPCs). MLL-AF4+ B-ALL shows a bimodal localization of the MLL gene breakpoint within the MLL break cluster region, and two subgroups of patients based on the gene expression pattern of the HOXA/MEIS cluster have been identified. The pathogenic mechanisms in MLL-AF4+ B-ALL are challenging to study functionally due to the absence of faithful human cellular models recapitulating the disease phenotype and latency. Here, we assess the molecular contribution and leukemogenic capacity of MLL breakpoints occurring in either intron 10 (MLLi10, centromeric) or intron 12 (MLLi12, telomeric) in ontogenically-different human HSPCs sourced prenatally (fetal liver) and neonatally (cord blood). CRISPR-Cas9-induced MLL-AF4 (MA) targeting either MLLi10 (Mi10A) or MLLi12 (Mi12A) causes MA-driven in vitro myeloid immortalization in both fetal liver- and cord blood-CD34+ HSPCs. The cellular ontogeny and the location of the MLL breakpoint influenced the capacity of MLL-edited CD34+ HSPCs to initiate pro-B-ALL in vivo, which faithfully recapitulated the molecular, transcriptomic and methylome profiles of patients with primary MA+ iB-ALL. This dataset contains the DNA methylation information (Illumina MethylationEPIC Beadchip platform) of MLL-edited CD34+ cells (i10 or i12). BCPs, used as controls, were obtained from E-MTAB-8505
2023-10-29 | E-MTAB-12806 | biostudies-arrayexpress
Project description:Comparison of gene expression of the adherent and non-adherent fraction of hematopoietic progenitor cells.
2007-03-13 | E-MEXP-586 | biostudies-arrayexpress
Project description:Gene expression profiling of primary cord blood hematopoietic stem cell (day 0, CD34+ cells), enriched control (untreated), Scriptaid and Valproic acid expanded CD34+ cells after a week in culture Individal cord blood CD34+ cells were processed individually and equal number of reisolated CD34+ cells from 3-4 samples were pooled after expansion to avoid the sample variations. Gene expression profiles of primary human cord blood CD34+ cells (day0), primary cells (PC) expanded in the presence or absence of histone deacetylase inhibitors (HDACIs) in serum containing cultures supplemented with a combination of cytokines (SCF, FLT3, IL3 and TPO) for 7 days.
2014-07-29 | E-GEOD-59803 | biostudies-arrayexpress
Project description:LincRNA expression profiles of human embryonic stem cells, CD34+ cells, and CD34+ derived induced pluripotent stem cells Total RNA was extracted from human ESCs, CD34+ cells, and CD34+ derived iPSC lines, then hybridized to Nimblegen tiling array GPL8792; Human lincRNA 391k Tiling Array V2
2010-11-10 | E-GEOD-23970 | biostudies-arrayexpress
Project description:Human CD34 cells isolated from cord blood or peripheral blood transduced with AML1-ETO or empty vector (MIGR1).
2007-08-30 | E-GEOD-2049 | biostudies-arrayexpress
Project description:CB CD34+ cells were were transduced with control (tNGFR) or CITED2 overexpression lentivectors, or control (shSCR) or shRNA CITED2 lentivectors and mRNA was isolated in order to investigate global gene expression changes upon perturbation of CITED2. CB CD34+ cells were isolated by Miltenyi miniMACS column. Cells were prestimulated in HPGM with 100 ng/ml KITL, FLT3L and TPO for 12 hrs. Cells were transduced with control (tNGFR) or CITED2 overexpression lentivectors, or control (shSCR) shRNA CITED2 lentivectors, in three rounds over 48 hrs. Transduced cells were sorted after which RNA was isolated for Illumina beadhchip arrays HT12 v4.
2014-09-05 | E-GEOD-47218 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the following subset Series: GSE23968: Large intergenic non-coding RNAs as novel modulators of reprogramming: ESCs, fibroblast, and fibroblast-derived iPSC (gene expression) GSE23970: Large intergenic non-coding RNAs as novel modulators of reprogramming: human embryonic stem cells, CD34+ cells, and CD34+ derived induced pluripotent stem cells (LincRNA expression) GSE23973: Large intergenic non-coding RNAs as novel modulators of reprogramming: siRNA (gene expression) GSE24181: Large intergenic non-coding RNAs as novel modulators of reprogramming: human embryonic stem cells, fibroblasts, and fibroblast-derived induced pluripotent stem cells (LincRNA expression) Refer to individual Series
2010-11-10 | E-GEOD-24182 | biostudies-arrayexpress