Expression data of human tonsilar CD4 positive T cell subsets
ABSTRACT: Type 1 regulatory T (Tr1) cells are one of the regulatory T cell subsets that are characterized by the production of high amount of IL-10 and lack of FOXP3 expression. Lymphocyte-activation gene 3 (LAG3) is a CD4 homologue molecule and we have previously reported that LAG3 is expressed on IL-10 producing regulatory T cells. However, naturally occurring Tr1 cells in human secondary lymphoid tissue have not been detected. We identified CD4+CD25-LAG3+ T cells in human tonsil. We compared mRNA expression of five CD4+ T cell subsets in tonsil using microarray analysis (CD4+CD25-LAG3+ T cells, CD4+CD25-CXCR5+PD-1+ follicular helper T cells (TFH), CD4+CD25+ T cells, CD4+CD25-LAG3-CD45RO+ cells and CD4+CD25-LAG3-CD45RO- cells). A human tonsil was obtained from a patient undergoing routine tonsillectomy, and five tonsillar CD4+ T cell subsets were sorted (each 1 x 10^5 cells). There is no biological replication.
Project description:Peripheral Blood Mononuclear Cells (PBMCs) were isolated from a buffy coat (Australian Blood Bank) using Ficoll methodology. CD4+ T cells were isolated using Dynal Beads kit. Pure CD4+ T cells were then stained using a cocktail of monoclonal antobodies (mAbs), including: anti-CD4PE, CD45RO ECD, CD62L APC-Cy7, CD25 APC, CD127 Pacific Blue. After incubation, cells were washed twice in PBS/FCS (0.2%), and sorted into five different cell subsets: CD4+CD25+CD127low CD62L+CD45RO- (naive regulatory T cells), CD4+CD25+CD127low CD62L+/- CD45RO+ (activated regulatory T cells), CD4+CD25+CD127hi CD62L+/- CD45RO+ (memory T cells), CD4+CD25-CD127low CD62L+/- CD45RO+ (effector T cells) and CD4+CD25-CD127hi CD62L+ CD45RO- (naive T cells).
Project description:CD4+CD25+FOXP3+ human regulatory T cells (Treg) are essential for self-tolerance and immune homeostasis. Here, we generated genome-wide maps of poised and active enhancer elements marked by histone H3 lysine 4 monomethylation and histone H3 lysine 27 acetylation for CD4+CD25highCD45RA+ naive and CD4+CD25highCD45RA- memory Treg and their CD25- conventional T cell (Tconv) counterparts after in vitro expansion . In addition we generated genome-wide maps of the transcription factors STAT5, FOXP3, RUNX1 and ETS1 in expanded CD4+CD25highCD45RA+ Treg- and CD4+CD25- Tconv to elucidate their role in cell type-specific gene regulation. ChIP-seq of 2 histone marks and transcription factors ETS1, STAT5, FOXP3 and RUNX1 in expanded T cell subpopulations
Project description:Regulatory T cells (Treg) prevent the emergence of autoimmune disease. Prototypic natural Treg (nTreg) are programmed by Forkhead-box P3 (FOXP3) and can be reliably identified by demethylation at the FOXP3 locus. To explore the nTreg methylation landscape we performed genome-wide methylation studies on human naïve resting nTreg (rTreg) and conventional naïve CD4+ T cells (Naïve). We detected 2,315 differentially methylated CpGs between these two cell types, many of which clustered into 127 regions of differential methylation (RDMs). T cell activation induced changes in 466 individual CpGs and 18 RDMs in naïve CD4+ T cells, but did not alter DNA methylation in rTreg. Expression of mRNA for TIGIT, an immune suppressive receptor demethylated in rTreg, was upregulated in nTreg and reduced in peripheral blood mononuclear cells of individuals at risk for autoimmune (type 1) diabetes. Gene-set testing of the 127 RDMs revealed enrichment of common Treg signature genes, FOXP3 bound genes and genes directly upregulated by FOXP3, which was primarily driven by the subset of demethylated RDMs. A putative Forkhead-binding motif overrepresented in promoter-associated RDMs suggests methylation regulates gene expression by influencing FOXP3 binding. Our findings provide new insights into epigenetic regulation of human nTreg and the potential to exploit differential methylation as an immune biomarker in human diseases. Naïve and rTreg cells were sorted from buffy coats of 3 healthy male donors (M28, 29, 30) and then activated for 6 days with anti-CD3 and anti-CD28 antibodies, supplemented with IL-2 at day 4. DNA was harvested and bisulfite converted for methylation analysis on illumina HM450 array from 2-3 biological replicates of each cell type: rTreg (M28, M30), naïve (M28, M29, M30), Act-naïve (M28, M29, M30) and Act-rTreg (M29, M30).
Project description:Molecular mechanism underline immune cell type population shift upon anti-DLL4 treatment C57BL/6 mice were injected with anti-DLL4, or an isotype control antibody as controls. Two weeks later mice were sacrificed, and thymi was harvested from 4 anti-DLL4 and 4 control animals. Total thymocytes, DN cells (CD4-CD8-) and DN1(CD4-CD8- CD44+CD25-) cells were isolated. Samples included in this data set are: 3 Thymocytes-anti-DLL4; 3 Thymocytes-isotype control; 3 DN1-anti-DLL4; 2 DN1-isotype controls; 3 DN-anti-DLL4; 3 DN-isotyoe control.
Project description:Comparison of gene expression profile of CD4+ CD25+, CD4+ CD25- CD45RBlow LAG3+ and CD4+ CD25- CD45RBlow LAG3- T cells. Naive CD4+CD25-CD45RBhigh T cells were used as a reference for pair comparison with values from the three other subsets.
Project description:Human responder T-cells (Tresp) and 2 lineages of regulatory T cells (Tregs), namely G2 and G3, were isolated by flow sorting based on their immunoreacitivities against specific cell surface markers and their gene expression profiles were interrogated using Affymetrix Exon 1.0 ST Arrays Comparison of gene expression profiles between responder T-cells, G2 Tregs and G3 Tregs
Project description:In this study, we used microarrays to investigate the gene expression program in conventional T cells, nTreg and conventional T cells treated with TGFbeta (iTreg) from wild-type mice and and mice having NFAT1/NFAT2 double-deficient (DKO) T cells. CD4+CD25- and CD4+CD25+ T cells were isolated by MACS and stimulated for 24 h with anti-CD3 and anti-CD28 antibodies in the absence or presence of TGFbeta. RNA was extracted and microarray analyses were performed. The data represents two independent biological replicates.
Project description:Foxp3, a transcription factor of the forkhead/winged helix family, plays a unique role in the transcriptional control of regulatory CD4+CD25+ T (Treg) cells. The identification of potential target genes associated with the regulatory phenotype and the characterization of combinatorial interactions of transcription factors (TF) in promoter regions in genes regulated by Foxp3 has become important to understand the transcriptional control of Treg cells.
Project description:Foxp3-mediated gene expression program in resting vs. activated CD4+ T cells The microarray part of this work consists of 12 Affymetrix assays corresponding to 3 biological replicates for each of 4 conditions for CD4+CD25- cells. Cells were either transduced with an empty vector or a vector encoding FOXP3, and in each case cells were either stimulated or not by anti-CD3/anti-CD28.
Project description:Rationale: COPD is characterized by an abnormal regulatory T cell (Treg) response and increases in Th1 and Th17 cell responses. It is unclear if dysregulation of miRNAs within Treg alters the adaptive immunophenotype in COPD. Objectives: To compare the miRNA profile of COPD Treg cells with that of healthy controls and to explore the function of differentially expressed miRNAs. Methods: Treg cells (CD4+CD25+CD49-CD127-) and T effector (CD4+CD25-) cells were obtained from the peripheral blood of 4 nonsmokers, 4 healthy current smokers, and 4 COPD current smokers for analysis of miRNA expression, matching them for age and sex. We assessed expression initially by microarray analysis on the Illumina miRNA platform then conducted real time RT-PCR validation of the microarray results. 24 samples were analyzed. 8 from each patient group of healthy Normal, Healthy Smoker, and COPD Smoker.