Project description:The deletion of the POZ domain of the transcription factor Miz1 leads to a late onset neuropathy with subsequent spontaneous regeneration in mice. In this study, we aim to identify genes that are involved in the development of this neuropathy. We traced first gene regulatory changes in sciatic nerve tissue from Miz1∆POZ mice to 30 days after birth. At this time point, we harvested samples of sciatic nerve tissue from several mice and performed RNAseq analysis to identify differentially expressed genes between Miz1∆POZ and wild type mice.
Project description:The role of FoxP3+ regulatory T (Treg) cells in the maintenance of immunological tolerance is well established. Recently, genome-wide association studies (GWAS) in humans have associated polymorphisms within the BACH2 locus encoding the transcription factor BTB and CNC homology 1, basic leucine zipper transcription factor 2 (Bach2) with diverse allergic and autoimmune diseases including asthma, multiple sclerosis, Crohn's disease, celiac disease, generalized vitiligo and type 1 diabetes. Common to these diseases is a failure to adequately maintain immunological tolerance. However, a role for Bach2 in this process has not been established. Here, by assessing the phenotype of mice in which the Bach2 gene is disrupted, we demonstrate a non-redundant role for Bach2 in the prevention of a spontaneous lethal inflammatory disorder predominantly affecting the lung and gut with excessive T helper 2 (Th2) responses and formation of circulating autoantibodies. Bach2 was necessary for efficient induction of FoxP3 expression both during thymopoesis and upon stimulation of naïve peripheral CD4+ T cells under Treg polarizing conditions in vitro. Consequently, in bone marrow reconstitution experiments, Bach2 expression within the haematopoetic system was necessary for suppression of lethal autoimmunity in a manner that was FoxP3 dependent. These findings demonstrate a requirement for Bach2 in early lineage commitment of both thymic and induced Treg cells and point to shared mechanisms that underlie diverse allergic and autoimmune disorders that may serve as targets in the development of novel therapeutic strategies. Six samples were collected from separate mice: three Ly5.1+ wildtype thymocyte samples (biological replicates) and three Ly5.1- Bach2 knockout thymocyte samples (biological replicates).
Project description:CTLA-4 is thought to inhibit effector T cells both intrinsically, by competing with CD28 for B7 ligands, and extrinsically, through the action of regulatory T cells. We studied in vivo responses of normal and CTLA-4-deficient antigen-specific murine effector CD4+ T cells. In order to do these studies in a physiological model of immunity to foreign antigen, we transferred small numbers of congenically marked RAG2-deficient 5C.C7 T cells with either a normal or knockout allele of CTLA-4 into normal syngeneic B10.A recipient mice. The T cells were then activated by immunization with MCC peptide and LPS. To look for transcriptional signatures of negative regulation of T cell responses by CTLA-4, we used microarray analysis to compare transcripts in wild type and CTLA-4 KO 5C.C7 T cells four days after immunization. This is the first instance in which differences are observed in extent of accumulation of wild type and CTLA-4 KO 5C.C7 T cells. To compare the gene expression profile between wild type and CTLA-4 KO adoptively transferred T cells 4 days after immunization.
Project description:Investigation of the SHIP1 (Inpp5d)-dependent and Rag2/Il2rg-dependent transcriptional changes in bone and osteoprogenitor cells cultures from murine strains with SHIP1 deficiency (Inpp5dm1Btlr/ m1Btlr, http://www.informatics.jax.org/allele/key/65037) and controls. Specifically, transcriptomic analysis was performed from femoral diaphysis from SHIPstyx/styx, SHIP1+/styx, wild-type (C57BL6/J), Rag2-/-/Il2rg-/-/SHIP1styx/styx, Rag2-/-/Il2rg-/-/SHIP1+/styx, and Rag2-/-/Il2rg-/-/SHIP1+/+ mice (n=3 for each genotype). Alternatively, osteoprogenitor cells isolated from bone marrow from the same genotypes as above were differentiated in vitro into primary osteoclasts in the presence or absence of RANKL. After 5 days, RNA was extracted from these cultures for transcriptomic analysis (n=3 for each genotype). Experimental Designs: Transcripts were analyzed in femoral diaphysis of SHIPstyx/styx, SHIP1+/styx, wild-type (C57BL6/J), Rag2-/-/Il2rg-/-/SHIP1styx/styx, Rag2-/-/Il2rg-/-/SHIP1+styx, and Rag2-/-/Il2rg-/-/SHIP1+/+ mice (n=3 for each genotype). Alternatively, transcripts were analyzed in primary osteoclast cultures from bone marrow cells isolated from SHIPstyx/styx, SHIP1+/styx, wild-type (C57BL6/J), Rag2-/-/Il2rg-/-/SHIP1styx/styx, Rag2-/-/Il2rg-/-/SHIP1+styx, and Rag2-/-/Il2rg-/-/SHIP1+/+ mice (n=3 for each genotype), which were supplemented or not with RANKL.
Project description:Eight weeks old male Adrb1tm1BkkAdrb2tm1Bkk/J , stock number 003810, were purchased from The Jackson Laboratory. These mice are homozygous null for the Adrb1 and Adrb2 genes, and are viable, fertile, normal in size and do not display any gross physical or behavioral abnormalities. Mice were euthanized, and the whole bone marrow was extracted using established methods.1 Whole bone marrow cells from Adrb1tm1BkkAdrb2tm1Bkk/J mice were reconstituted into lethally- irradiated (950 Rad) C57BL/6J mice using a single retro-orbital injection at a ratio of 1:4 (Adrb1tm1BkkAdrb2tm1Bkk/J to C57BL/6J). All reconstituted mice were recovered for 2-3 months prior to tissue collection for mRNA arrays (1) The FASEB Journal 2015:29(1),652.13. Seven control animals and seven animals that received new bone marrow from double adrenergic receptor knockout (treated KO chimera). Following recovery, tissues that were harvested included bone marrow, brain, and distal colon.
Project description:Eight weeks old male Adrb1tm1BkkAdrb2tm1Bkk/J , stock number 003810, were purchased from The Jackson Laboratory. These mice are homozygous null for the Adrb1 and Adrb2 genes, and are viable, fertile, normal in size and do not display any gross physical or behavioral abnormalities. Mice were euthanized, and the whole bone marrow was extracted using established methods.1 Whole bone marrow cells from Adrb1tm1BkkAdrb2tm1Bkk/J mice were reconstituted into lethally- irradiated (950 Rad) C57BL/6J mice using a single retro-orbital injection at a ratio of 1:4 (Adrb1tm1BkkAdrb2tm1Bkk/J to C57BL/6J). All reconstituted mice were recovered for 2-3 months prior to tissue collection for mRNA arrays (1) The FASEB Journal 2015:29(1),652.13. Eight control animals and eight animals that received new bone marrow from double adrenergic receptor knockout (treated KO chimera). Following recovery, tissues that were harvested included bone marrow, brain, and distal colon.
Project description:Eight weeks old male Adrb1tm1BkkAdrb2tm1Bkk/J , stock number 003810, were purchased from The Jackson Laboratory. These mice are homozygous null for the Adrb1 and Adrb2 genes, and are viable, fertile, normal in size and do not display any gross physical or behavioral abnormalities. Mice were euthanized, and the whole bone marrow was extracted using established methods.1 Whole bone marrow cells from Adrb1tm1BkkAdrb2tm1Bkk/J mice were reconstituted into lethally- irradiated (950 Rad) C57BL/6J mice using a single retro-orbital injection at a ratio of 1:4 (Adrb1tm1BkkAdrb2tm1Bkk/J to C57BL/6J). All reconstituted mice were recovered for 2-3 months prior to tissue collection for mRNA arrays (1) The FASEB Journal 2015:29(1),652.13. Eight control animals and eight animals that received new bone marrow from double adrenergic receptor knockout (treated KO chimera). Following recovery, tissues that were harvested included bone marrow, brain, and distal colon.
Project description:CD4 T cell help is critical for both the generation and maintenance of germinal centers, and T follicular helper (TFH) cells are the CD4 T cell subset required for this process. SAP (SH2D1A) expression in CD4 T cells is essential for germinal center development. However, SAP-deficient mice have only a moderate defect in TFH differentiation as defined by common TFH surface markers. CXCR5+ TFH cells are found within the germinal center as well as along the boundary regions of T/B cell zones. Here we show that germinal center associated T cells (GC TFH) can be identified by their co-expression of CXCR5 and the GL7 epitope, allowing for phenotypic and functional analysis of TFH and GC TFH populations. Here we show GC TFH are a functionally discrete subset of further polarized TFH cells, with enhanced B cell help capacity and a specialized ability to produce IL-4 in a TH2-independent manner. Strikingly, SAP-deficient mice have an absence of the GC TFH subset and SAP- TFH are defective in IL-4 and IL-21 production. We further demonstrate that SLAM (Slamf1, CD150), a surface receptor that utilizes SAP signaling, is specifically required for IL-4 production by GC TFH. GC TFH cells require IL-4 and IL-21 production for optimal help to B cells. These data illustrate complexities of SAP-dependent SLAM family receptor signaling, revealing a prominent role for SLAM receptor ligation in IL-4 production by germinal center CD4 T cells but not in TFH and GC TFH differentiation. This SuperSeries is composed of the following subset Series: GSE21379: Expression Data from WT and Sh2d1a-/- in vivo follicular helper CD4 T cells (TFH) versus non follicular helper CD4 T cells (non-TFH) GSE21380: Expression Data from in vivo Tfh vs GC Tfh vs non-Tfh Refer to individual Series
Project description:RNA microarray profiling analysis was performed on splenic B cell subsets: “IgD+CD27-” (naive B cells) and “IgD+CD27+” (MZB B cells) isolated from splenic samples of 6 adults
Project description:CD4 T cell help is critical for both the generation and maintenance of germinal centers, and T follicular helper (TFH) cells are the CD4 T cell subset required for this process. SAP (SH2D1A) expression in CD4 T cells is essential for germinal center development. However, SAP-deficient mice have only a moderate defect in TFH differentiation as defined by common TFH surface markers. CXCR5+ TFH cells are found within the germinal center as well as along the boundary regions of T/B cell zones. Here we show that germinal center associated T cells (GC TFH) can be identified by their co-expression of CXCR5 and the GL7 epitope, allowing for phenotypic and functional analysis of TFH and GC TFH populations. Here we show GC TFH are a functionally discrete subset of further polarized TFH cells, with enhanced B cell help capacity and a specialized ability to produce IL-4 in a TH2-independent manner. Strikingly, SAP-deficient mice have an absence of the GC TFH subset and SAP- TFH are defective in IL-4 and IL-21 production. We further demonstrate that SLAM (Slamf1, CD150), a surface receptor that utilizes SAP signaling, is specifically required for IL-4 production by GC TFH. GC TFH cells require IL-4 and IL-21 production for optimal help to B cells. These data illustrate complexities of SAP-dependent SLAM family receptor signaling, revealing a prominent role for SLAM receptor ligation in IL-4 production by germinal center CD4 T cells but not in TFH and GC TFH differentiation. Analysis of in vivo polyclonal GC Tfh vs Tfh vs Non-Tfh eight days after LCMV viral infection. Analysis of in vivo follicular helper CD4 T cells (CXCR5high GL7low), versus germinal center follicular helper CD4 T cells (CXCR5hi GL7hi), versus non-follicular helper CD4 T cells (CXCR5low) eight days after viral infection.