ABSTRACT: Gene expression profiling of CD4+CD25+Foxp3+ regulatory T cells and CD4+CD25-Foxp3- T cells in the course of Plasmodium yoelii infection of BALB/c mice
Project description:The objective of the present study was to characterize the phenotype of CD4+CD25+Foxp3+ regulatory T cells (Tregs) in the course of parasitic Plasmodium yoelii (P .yoelii) infection of BALB/c mice. Therefore we performed microarray expression analysis of CD4+CD25+Foxp3+ Tregs isolated by FACS from spleens of non-infected mice and from spleens of mice infected with P. yoelii 3 days and 5 days post infection. By comparing the gene expression profiles, we were able to identify molecules which were differentially expressed by Tregs during parasitic infection and thereby might be involved in their immune-suppressive function. Moreover, we included CD4+CD25-Foxp3- T cells from spleens of non-infected and P. yoelii-infected mice in our analysis. It was proposed that immune-suppressive CD4+CD25-Foxp3- T cells might be induced during Plasmodium infection of mice. Thus, detailed gene expression data of these cells in comparison to CD4+CD25+Foxp3+ Tregs would contribute a better understanding in the phenotype. FACS sorted CD4+CD25+Foxp3+ Tregs and CD4+CD25-Foxp3- T cells from pooled spleens of non-infected Foxp3/ eGFP mice (served as reference) and from pooled spleens of P. yoelii infected Foxp3/ eGFP mice 3 days and 5 days post infection were analyzed as single probes.
Project description:The objective of the present study was to characterize the phenotype of CD4+CD25+Foxp3+ regulatory T cells (Tregs) in the course of parasitic Plasmodium yoelii (P .yoelii) infection of BALB/c mice. Therefore we performed microarray expression analysis of CD4+CD25+Foxp3+ Tregs isolated by FACS from spleens of non-infected mice and from spleens of mice infected with P. yoelii 3 days and 5 days post infection. By comparing the gene expression profiles, we were able to identify molecules which were differentially expressed by Tregs during parasitic infection and thereby might be involved in their immune-suppressive function. Moreover, we included CD4+CD25-Foxp3- T cells from spleens of non-infected and P. yoelii-infected mice in our analysis. It was proposed that immune-suppressive CD4+CD25-Foxp3- T cells might be induced during Plasmodium infection of mice. Thus, detailed gene expression data of these cells in comparison to CD4+CD25+Foxp3+ Tregs would contribute a better understanding in the phenotype.
Project description:Expression data from antigen-experienced Nfat1+/+ and Nfat1-/- CD4+ T cells following 21 days of Plasmodium yoelii 17XNL infection.
Project description:<p>We use next generation sequencing to investigate the different transcriptomes of closely related CD4+ T-cells from healthy human donors to elucidate the genetic programs that underlie their specialized immune functions. Six cell types were included: Regulatory T-cells (CD25hiCD127low/neg with >95% FOXP3+ purity), regulatory T-cells activated using PMA/ionomycin, CD25-CD45RA+ ('naive' helper T-cells), CD25-CD45RO+ ('memory' helper T-cells), activated Th17 cells (>98% IL17A+ purity) and activated IL17-CD4+ T-cells (called 'ThPI'). Poly-T capture beads were used to isolate mRNA from total RNA, and fragment sizes of ~200 were sequenced from both ends on Illumina's genome analyzer. We confirm many of the canonical signature genes of T-cell populations, but also discover new genes whose expression is limited to specific CD4 T-cell lineages, including long non-coding RNAs. Additionally, we find that genes encoded at loci linked to multiple human autoimmune diseases are enriched for preferential expression upon T-cell activation, suggesting that an aberrant response to T-cell activation is fundamental to pathogenesis.</p>
Project description:Naturally occurring CD25+CD4+ regulatory T cells (T reg cells) are currently intensively characterized because of their major importance in modulating host responses to tumors and infections, in preventing transplant rejection, and in inhibiting the development of autoimmunity and allergy. Originally, CD4+ T reg cells were identified exclusively by the constitutive expression of CD25, and many in vivo experiments have been performed using depleting antibodies directed against CD25. However, both the existence of CD25– T reg cells, especially within peripheral tissues, as well as the expression of CD25 on activated conventional T cells, which precludes discrimination between T reg cells and activated conventional T cells, limits the interpretation of data obtained by the use of anti-CD25 depleting antibodies. The most specific T reg cell marker currently known is the forkhead box transcription factor Foxp3, which has been shown to be expressed specifically in mouse CD4+ T reg cells and acts as a master switch in the regulation of their development and function. To address the question of the in vivo role of T reg cells in immunopathology, we have generated bacterial artificial chromosome (BAC)–transgenic mice termed depletion of regulatory T cell (DEREG) mice, which express a diphtheria toxin receptor (DTR) enhanced GFP (eGFP) fusion protein under the control of the foxp3 locus, allowing both detection and inducible depletion of Foxp3+ T reg cells. The gene expression profile of both CD4+eGFP+FoxP3+ and CD4+eGFPnegFoxP3neg cells isolated from DEREG mice was here analyzed by micro array. Keywords: DEREG, FoxP3, FoxP3-EGFP, mouse, regulatory T cell, CD4 CD4+GFP+FoxP3+ and CD4+GFPnegFoxP3neg cells were isolated from DEREG mice by negative selection of CD4+ T cells (Invitrogen Kit) and subsequent FACS sorting for GFP+ and GFPneg cells. Purity was greater than 99 %. cRNA was prepared according to the Affymetrix Labeling Protocol, fragmented and hybridized to Affymetrix GeneChip Mouse Genome 430.
Project description:Identification of innate immune responses in the livers of mice infected with liver-stage arresting, transgenic, Plasmodium yoelii parasites. Whole liver samples from mock and P. yoelli fabb/f- infected C57BL/6 and BALB/cJ mice. Samples were taken 3 days post infection
Project description:We analyzed the individual transcriptomes of thymic Treg cells (CD4+Foxp3+), their immediate precursors (CD25+CD4+Foxp3-) and mature CD4 single positive thymocytes (CD4+Foxp3-CD25-CD62L+CD24-).
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: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).
Project description:Naturally occurring CD25+CD4+ regulatory T cells (T reg cells) are currently intensively characterized because of their major importance in modulating host responses to tumors and infections, in preventing transplant rejection, and in inhibiting the development of autoimmunity and allergy. Originally, CD4+ T reg cells were identified exclusively by the constitutive expression of CD25, and many in vivo experiments have been performed using depleting antibodies directed against CD25. However, both the existence of CD25– T reg cells, especially within peripheral tissues, as well as the expression of CD25 on activated conventional T cells, which precludes discrimination between T reg cells and activated conventional T cells, limits the interpretation of data obtained by the use of anti-CD25 depleting antibodies. The most specific T reg cell marker currently known is the forkhead box transcription factor Foxp3, which has been shown to be expressed specifically in mouse CD4+ T reg cells and acts as a master switch in the regulation of their development and function. To address the question of the in vivo role of T reg cells in immunopathology, we have generated bacterial artificial chromosome (BAC)–transgenic mice termed depletion of regulatory T cell (DEREG) mice, which express a diphtheria toxin receptor (DTR) enhanced GFP (eGFP) fusion protein under the control of the foxp3 locus, allowing both detection and inducible depletion of Foxp3+ T reg cells. The gene expression profile of both CD4+eGFP+FoxP3+ and CD4+eGFPnegFoxP3neg cells isolated from DEREG mice was here analyzed by micro array. Keywords: DEREG, FoxP3, FoxP3-EGFP, mouse, regulatory T cell, CD4