Project description:While unique subsets of Treg cells have been described in some non-lymphoid tissues, their relationship to Treg cells in secondary lymphoid organs and circulation remains unclear. We have identified a recirculating and highly suppressive effector Treg cell subset that expresses the α2 integrin, CD49b, and exhibits a unique tissue distribution. We used TCR sequencing to ask whether CD49b+ activated Treg cells and CD49b− activated Treg cells represent stable states of Treg cell differentiation instructed by distinct TCR repertoires. We did not find a pattern of clonotypes that was specific to either subset, suggesting that essentially all CD49b− Treg cells could give rise to CD49b+ cells. These results shed light on the identity and development of a functionally potent subset of mature effector Treg cells that recirculate through and survey peripheral tissues.
Project description:While unique subsets of Treg cells have been described in some non-lymphoid tissues, their relationship to Treg cells in secondary lymphoid organs and circulation remains unclear. We have identified a recirculating and highly suppressive effector Treg cell subset that expresses the α2 integrin, CD49b, and exhibits a unique tissue distribution. To identify genes and pathways enriched in CD49b+ Treg cells, we performed RNA-seq of splenic CD49b+ and CD49b− Treg cells that were of otherwise similar activation status based on expression of CD44 and CD62L. This revealed that splenic CD49b+ Treg cells express genes related to migration and activation, but are relatively depleted of genes whose expression is TCR-dependent in Treg cells. These results shed light on the identity and development of a functionally potent subset of mature effector Treg cells that recirculates through and surveys peripheral tissues.
Project description:With the aim of understanding how Treg cells in highly vascularized tissues are related to Treg cells in other organs, we performed RNA-seq analysis of bulk Treg and Tconv cells isolated from liver, blood, spleen, and the liver-draining portal lymph node. This revealed a clear separation of cell transcriptomes by both tissue and Treg/Tconv identity, with cells from the liver falling between blood- and spleen-derived cells. Compared to splenic Treg cells, hepatic Treg cells were enriched for genes related to proliferation and activation, and genes encoding chemokine and cytokine receptors.
Project description:Autoimmune diseases, such as rheumatoid arthritis (RA), are believed to be caused, in part, by a defect in the selection of T cells. Developing T cells undergo a process known as “negative selection” where self-derived proteins are presented to the cells; those that react to the self-peptides are either eliminated, or develop into regulatory T cells (Tregs), which suppress immune responses by conventional T cells (Tconvs). In order to study self-reactive T cell receptors (TCRs), and their likely target antigens, we performed single cell repertoire analysis in ZAC mice with impaired TCR signaling. We sequenced TCR and transcriptomes of Treg and Tconv single cells from arthritic ZAC mice spleen (n=2). We discovered a cluster of pro-inflammatory Th17 Tconv cells in ZAC spleens that was absent in WT mice. ZAC Th17 TCRs showed clonal expansion. Different mice had different TCR sequences, which is expected.
Project description:We reported transcriptional characterization of Treg and Tconv cells from thymic, splenic, and visceral adipose tissue (VAT) of vTreg53 TCR transgenic mice and control littermates. We examined the effect of Foxp3 on splenic and VAT CD4+ T cell transcriptome. We profiled gene expression in a novel PPARg+ splenic Treg population. We uncovered that the characteristic phenotype of VAT Treg cells was acquired in two stages.
Project description:miRNA expression profiling in highly purified murine CD4+ Tconv and Treg cells. FoxP3-GFP-hCre1a(high) reporter mice were used to separate both populations based on surface markers and presence or absence of GFP. Two-condition experiment, Tconv vs. Treg. Biological replicates: 1 Tconv, 1 Treg, purified from the same pooled mice. One replicate on 1 array.
Project description:Microarray used to detail the global gene transcription underlying sorted IFNg+ and IFNg- Tregs (CD4+CD25+CD127lo) and Tconv (CD4+CD25-CD127+) for fresh (unexpanded) and 14 day expanded cells from human blood. Treg and Tconv were FACS isolated from five healthy subjects (median age of 26, range 22-30). Sorted cells were separated into two groups: the first group was stimulated for 4 hours with PMA/ionomycin and labeled with the IFNg cytokine capture kit (Miltenyi Biotech) followed by FACS isolation of IFNg- and IFNg+ populations. The second set was expanded to day 14 prior to reactivation and cytokine cell capture. For each IFNg sorted population, cells were pelleted and flash frozen before RNA isolation and processing.
Project description:Thymic-derived natural T regulatory cells (nTregs) are characterized by functional and phenotypic heterogeneity. Recently, a small fraction of peripheral Tregs have been shown to express Klrg1, but it remains unclear the extent Klrg1 defines a unique Treg subset. Here we show that Klrg1+ Tregs represent a terminally differentiated Treg subset derived from Klrg1- Tregs. This subset is a recent antigen-responsive and a highly activated short-lived Treg population that expresses enhanced levels of Treg suppressive molecules and that preferentially resides within mucosal tissues. The development of Klrg1+ Tregs also requires extensive IL-2R signaling. This activity represents a distinct function for IL-2, independent from its contribution to Treg homeostasis and competitive fitness. These and other properties are analogous to terminally differentiated short-lived CD8+ T effector cells. Our findings suggest that an important pathway driving antigen-activated conventional T lymphocytes also operates for Tregs. Gene expression analysis was performed of this and other Treg subsets based on expression of CD62L, CD69, and Klrg1 to define the molecular properties of Klrg1+ Tregs and its relationship to other Treg subsets found in the peripheral immune tissues. Mice were euthanized, spleen cell preparations were made, and each Treg subset was isolated by FACS cell sorting. RNA was immediately prepared for processing.
Project description:We have employed whole genome microarray expression profiling of human Treg and Tconv cells as a discovery platform to identify genes differentially expressed upon signal of pseudostarvation (either mTOR inhibition or leptin neutralizaion) responsible for proliferation of Treg cell on one side and inhibition of Tconv cells on the other. Human Treg and Tconv cells isolated from healthy donors were pretreated or not with rapamycin or leptin mAb and stimulated with anti-CD3 and anti-CD28 beads for 12h.