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:Regulatory T cells (Tregs) are engaged in the maintenance of immunological self-tolerance and immune homeostasis. IL-10 has an important role in maintaining the normal immune state. Here, we show that IL-10-secreting Tregs can be delineated in normal mice as CD4(+)CD25(-)Foxp3(-) T cells that express lymphocyte activation gene 3 (LAG-3), an MHC-class-II-binding CD4 homolog. Although approximately 2% of the CD4(+)CD25(-) T cell population consisted of CD4(+)CD25(-)LAG3(+) T cells in the spleen, CD4(+)CD25(-)LAG3(+) T cells are enriched to approximately 8% in the Peyer's patch. They are hypoproliferative upon in vitro antigenic stimulation and suppress in vivo development of colitis. Gene expression analysis reveals that CD4(+)CD25(-)LAG3(+) Tregs characteristically express early growth response gene 2 (Egr-2), a key molecule for anergy induction. Retroviral gene transfer of Egr-2 converts naïve CD4(+) T cells into the IL-10-secreting and LAG-3-expressing phenotype, and Egr-2-transduced CD4(+) T cells exhibit antigen-specific immunosuppressive capacity in vivo. Unlike Foxp3(+) natural Tregs, high-affinity interactions with selecting peptide/MHC ligands expressed in the thymus do not induce the development of CD4(+)CD25(-)LAG3(+) Tregs. In contrast, the number of CD4(+)CD25(-)LAG3(+) Tregs is influenced by the presence of environmental microbiota. Thus, IL-10-secreting Egr-2(+)LAG3(+)CD4(+) Tregs can be exploited for the control of peripheral immunity.
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:Systemic lupus erythematosus (SLE) is a prototypical autoimmune disease characterized by multiorgan inflammation induced by autoantibodies. Early growth response gene 2 (Egr2), a transcription factor essential for T-cell anergy induction, controls systemic autoimmunity in mice and humans. We have previously identified a subpopulation of CD4+ regulatory T cells, CD4+CD25-LAG3+ cells, that characteristically express both Egr2 and LAG3 and control mice model of lupus via TGF-?3 production. However, due to the mild phenotype of lymphocyte-specific Egr2-deficient mice, the presence of an additional regulator has been speculated. Here, we show that Egr2 and Egr3 expressed in T cells cooperatively prevent humoral immune responses by supporting TGF-?3 secretion. T cell-specific Egr2/Egr3 double-deficient (Egr2/3DKO) mice spontaneously developed an early onset lupus-like disease that was more severe than in T cell-specific Egr2-deficient mice. In accordance with the observation that CD4+CD25-LAG3+ cells from Egr2/3DKO mice completely lost the capacity to produce TGF-?3, the excessive germinal center reaction in Egr2/3DKO mice was suppressed by the adoptive transfer of WT CD4+CD25-LAG3+ cells or treatment with a TGF-?3-expressing vector. Intriguingly, latent TGF-? binding protein (Ltbp)3 expression maintained by Egr2 and Egr3 was required for TGF-?3 production from CD4+CD25-LAG3+ cells. Because Egr2 and Egr3 did not demonstrate cell intrinsic suppression of the development of follicular helper T cells, Egr2- and Egr3-dependent TGF-?3 production by CD4+CD25-LAG3+ cells is critical for controlling excessive B-cell responses. The unique attributes of Egr2/Egr3 in T cells may provide an opportunity for developing novel therapeutics for autoantibody-mediated diseases including SLE.
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.