Homo sapiens
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ABSTRACT:
PROVIDER: PRJNA222314 | ENA |
REPOSITORIES: ENA
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Project description:Research in human immunobiology is mainly based on working with peripheral blood mononuclear cells (PBMC). However, recent investigations have shown that circulating CD4+ T cells are less sensitive to several T-cell activating monoclonal antibodies (mAb) and to recall antigens as compared to tissue-resident cells or cells that were in-vitro cultured at a high cell density of 10^7 cells/mL for 2 days at 37°C and 5% CO2 (RESTORE protocol, Römer et al., Blood 2011, PMID: 21931118). To explain the increase in sensitivity of CD4+ T-cells to mAbs and recall antigens on a molecular level, we performed microarray hybridizations of total RNA from T-cells isolated from PBMC that were cultured at a low or high cell density. To avoid the detection of genes that are up- or down-regulated by the culture process itself, we used low cell density cultured PBMC, instead of freshly prepared PBMC. We used microarrays to detail the global programme of gene expression underlying differences in the cell density of human PBMC and identified genes that are significantly up- or downregulated dependent on the cell density of PBMC. Human PBMC of one healthy blood donor were cultured at a low cell density (10^6 cells/mL) or at a high cell density (10^7 cells/mL) for 2 days at 37°C and 5% CO2 and CD4 or CD8 T-cells of both cultures were isolated by MACSbeads. Expression profiles from total RNA extracts were generated by hybridization to Affymetrix microarrays.
2015-06-09 | E-GEOD-51288 | biostudies-arrayexpress
Project description:Research in human immunobiology is mainly based on working with peripheral blood mononuclear cells (PBMC). However, recent investigations have shown that circulating CD4+ T cells are less sensitive to several T-cell activating monoclonal antibodies (mAb) and to recall antigens as compared to tissue-resident cells or cells that were in-vitro cultured at a high cell density of 10^7 cells/mL for 2 days at 37°C and 5% CO2 (RESTORE protocol, Römer et al., Blood 2011, PMID: 21931118). To explain the increase in sensitivity of CD4+ T-cells to mAbs and recall antigens on a molecular level, we performed microarray hybridizations of total RNA from T-cells isolated from PBMC that were cultured at a low or high cell density. To avoid the detection of genes that are up- or down-regulated by the culture process itself, we used low cell density cultured PBMC, instead of freshly prepared PBMC. We used microarrays to detail the global programme of gene expression underlying differences in the cell density of human PBMC and identified genes that are significantly up- or downregulated dependent on the cell density of PBMC.
2015-06-09 | GSE51288 | GEO
Project description:We analyzed the total proteome of CD4+ and CD8+ T cells isolated from human peripheral blood mononuclear cells (PBMC), and cultured to perform a CRISPR/CAS9 edition of their genome, in order to introduce an OST sequence at the C-terminus of proteins of interest (SLP76 or ZAP70, n=3 biological replicates in each case). Control T cells , isolated and cultured in the same way, but not modified by CRISPR/CAS9, were also analyzed (WT, n=3 or 6 biological replicates).
2022-01-10 | PXD025174 | Pride
Project description:High density cultured CD8+ memory T-cells from peripheral blood mononuclear cells (PBMC) were found to be more sensitive towards antigenic stimulation. We used microarrays to detail the global programme of gene expression underlying differences in the cell density of human PBMC and identified genes that are significantly up- or downregulated dependent on the cell density of PBMC.
2015-06-09 | GSE63430 | GEO
Project description:High density cultured CD8+ memory T-cells from peripheral blood mononuclear cells (PBMC) were found to be more sensitive towards antigenic stimulation. We used microarrays to detail the global programme of gene expression underlying differences in the cell density of human PBMC and identified genes that are significantly up- or downregulated dependent on the cell density of PBMC. Paired analysis of CD8+ memory T-cells from fresh and high density precultured PBMC from three healthy donors.
2015-06-09 | E-GEOD-63430 | biostudies-arrayexpress
Project description:Comprehensive human immune cell proteomic spectral library which contain CD4+T cell, CD8+ T cell, NK cell, B cell, PBMC, Daudi, Jurkat, stimulated Jurkat, Molt-4, Ramos, and THP-1 were generated. This library includes 10,544 proteins groups, and 127,106 peptides.
2023-12-12 | MSV000093644 | MassIVE
Project description:Whole blood was collected from sows and peripheral blood mononuclear cells (PBMC) were isolated. CD3+CD4+, CD3+CD8+ and CD3+CD4+CD8+ were separately sorted by a cell sorter. Transcriptional profile of each T cell subset was obtained.
2018-01-10 | GSE105784 | GEO
Project description:Bone marrow Hdc-GFPhi and Hdc-GFPlo HSPC (SLAM-LSK, Lin-c-kit+Sca-1+CD150+CD48-) HSCs were isolated from mouse femur and tibia from histidine decarboxylase (Hdc) green fluorescent protein (Hdc-GFP) mice. Hdc-GFPhi HSC and Hdc-GFPlo HSC cells were sorted by combinations of GFP and the cell surface markers of HSC. total RNA was isolated from sorted 2,500 HSPCs using ARCTURUS PicoPure RNA isolation kit (Life Technologies). cDNA was amplified and libraries were constructed by using SMARTer Ultra Low Input RNA kit (Clontech Laboratories) and Nextera XT DNA Library Preparation kit (Illumina) according to manufacturer’s instructions respectively. Sequencing was performed on the Illumina HiSeq2000 platform.
2019-11-20 | GSE90787 | GEO
Project description:To investigate the role of PPAR-γ in human TH cells, transcriptional response of activated “TH9” clones to treatment with GW9662, a potent PPAR-γ antagonist was assessed. “TH9” cell clones were incubated with GW9662 for 48h and activated with αCD3/2/28 for 12 h. Transcriptomic profiling was performed by bulk RNA-seq. To generate TH9 clones, Peripheral Blood Mononuclear Cells (PBMC) were isolated by Ficoll-Plaque Plus (GE Healthcare, UK) density gradient centrifugation. Human CD4+ T cells were isolated from PBMC by EasySep positive selection kit (Stemcell Technologies) according to manufacturer’s instruction. Positively selected CD4+ T cells were washed with PBS and stained for subsequent TH cell subset sorting. Effector memory “TH9” cells (CXCR3-CCR8+CCR6-CCR4+) were sorted to over 90% purity according to their expression of chemokine receptors from CD45RA-CD25-CD8-CD3+ cells. Single cell “TH9” clones were directly sorted into 96well plate according to their expression of chemokine receptors (see above). Single cell clones were expanded and maintained by periodic restimulation with PHA (1 µg/ml, Sigma-Chemicals) and irradiated allogenic feeder cells (5x104/well) in culture medium.
2022-10-31 | E-MTAB-12197 | biostudies-arrayexpress
Project description:Purpose: We aim to identify transciptional changes of human CD4+/CD8+ T cells due to high fat low carbohydrate ketogenic diet (KD) in vivo. Method: Healthy volunteers conducted a 21 days ketogenic diet, limiting carbohydrate intake to <30g/day. Before the start (T0) and at the end (T1) of the diet, blood samples were taken and PBMC were isolated. PBMCs were obtained by density centrifugation (Histopaque 1077, Sigma-Aldrich, St. Louis, MO, USA). A ViCell analyzer (Beckman Coulter, Fullerton, CA, USA) was used to evaluate the cell count and viability. Only samples exceeding a cell viability of 90% were processed further. PBMCs were subjected to cell cultivation in RPMI 1640 (Invitrogen, Carlsbad, CA, USA) containing 10% heat-inactivated fetal calf serum (Biochrom, Berlin, Germany), 1% HEPES (Sigma-Aldrich, St. Louis, MO) and 1% L-glutamine (Life Technologies, Carlsbad, CA, USA). T cells were stimulated via the addition of CD3/CD28 Dynabeads (Thermo Fisher Scientific, Waltham, MA, USA) with a bead-to-cell ratio of 1:8 for a duration 24 hours. After stimulation, CD3/CD28 Dynabeads were magnetically removed. Pan T cell-, CD4+- and CD8+-cell-isolation was performed by magnetic cell separation (Pan T Cell Isolation Kit, # 130-096-535 | human CD4 MicroBeads, # 130-045-101 | human CD8 MicroBeads, # 130-045-201, Miltenyi Biotec, Bergisch Gladbach, Germany) using an AutoMACS Pro Separator ( # 130-092-545, Miltenyi Biotec, Bergisch-Gladbach, Germany) according to the manufacturer’s instructions. Results: 11.545 expressed genes were identified, for CD4/CD8 T cells, we detected 5.667/ 5.799 up-regulated genes and 5.878/5.746 down-regulated genes. 294/346 genes and 325/252 genes were significantly up/down-regulated for CD4+/CD8+ T cells (p-val. <0.05). Gene set enrichment analysis revealed 117/17 and 22/6 significantly up/down-regulated pathways for CD4+/CD8+ T cells (p-val. <0.05). Genes and Gene sets differentially regulated were relevant for T cell immune response and metabolic function. Conclusion: KD resulted in immunometabolic reprogramming of human CD4+/CD8+ T cells.
2020-12-31 | GSE158407 | GEO