5 research outputs found

    TRAF6ΔDC mice exhibit disruption of microbiotic homeostasis localized to the small intestine.

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    <p><b>(A)</b> Pie charts represent bacterial composition categorized by phylum in the small bowl of control (WT) or TRAF6ΔDC (ΔDC) mice. <b>(B)</b> Heatmap analysis shows relative abundance of taxa as a percentage of total 16S rRNA, organized according to genus or most specific assigned taxon. Color scales reflect proportion contributed by each taxon. Total bacterial genome was isolated from small intestinal contents of control (WT) or TRAF6ΔDC (ΔDC) mice (n = 4, 20 week old littermate and co-housed group). <b>(C)</b> The copy number of total bacterial 16S rRNA bacteria from fecal or small intestine contents of control (WT) and TRAF6ΔDC (ΔDC) was measured by comparing to reference E. coli 16S rRNA plasmids. Some groups were fed by broad-spectrum antibiotic water (Abx) for last 2 weeks before collecting samples. **p < 0.001; n.s., not significant. Data were analyzed by Anova on Prism software (n = 3).</p

    Increased Tregs in GF TRAF6ΔDC lymphoid organs following antibiotic treatment.

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    <p><b>(A)</b> FACS plots gated on CD4<sup>+</sup> T cells show intracellular staining Foxp3 from each indicated organ of germ-free (GF) TRAF6ΔDC bone marrow chimera (ΔDC-BMC) at 8 weeks post-reconstitution. Broad-spectrum antibiotic water was provided to a group of TRAF6ΔDC bone marrow chimeras (ΔDC) for the last 2 weeks. <b>(B)</b> Percentage of Foxp3<sup>+</sup> CD4 T cells were determined from the indicated organs of TRAF6ΔDC bone marrow chimera (ΔDC-BMC) in germ-free (GF) or germ-free under antibiotics (GF-Abx). <b>(C)</b> Representative FACS plots show Foxp3<sup>+</sup> CD4 T cells from each indicated organ of germ-free (GF) B6 mice. Some of the mice were provided with broad-spectrum antibiotic water (GF-Abx) for last 2 weeks. Percentage of Foxp3<sup>+</sup> CD4 T cells were shown in each organs of germ-free B6 mice with (GF-Abx) or without antibiotics (GF). SP, spleen; MLN, mesenteric lymph node; SI, small intestine; COL, colon. **p < 0.01.</p

    Germ-free conditions exacerbate inflammation of TRAF6ΔDC small intestine.

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    <p>(A) Representative gut images from control (WT) and TRAF6ΔDC (ΔDC) bone marrow chimera established in specific pathogen free (SPF) or germ-free (GF) condition at 8 weeks post-reconstitution. (B) Histological analyses were performed by H&E staining of duodenum from each mice. Scale bars represent 100 μm. (C) Lengths and masses of small intestines and colons were measured in control (WT) and TRAF6ΔDC (ΔDC) bone marrow chimera (n = 6). SPF, specific pathogen free; GF, germ-free. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.</p

    Broad-spectrum antibiotics ameliorate TRAF6ΔDC immune pathology exacerbated by GF conditions.

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    <p><b>(A)</b> FACS plots gated on CD4<sup>+</sup> T cells show intracellular staining for interferon-γ (IFN-γ) and IL-13 from each indicated organ of TRAF6ΔDC bone marrow chimera (ΔDC-BMC) in germ-free (GF) condition at 8 weeks post-reconstitution. Some of TRAF6ΔDC bone marrow chimeras (ΔDC) were provided with broad-spectrum antibiotic water (Abx). The antibiotic water, containing 1 g/L Ampicillin, 1 g/L Neomycin, 0.5 g/L Vancomycin, and 1 g/L Metronidazole, was provided ad libitum in water to TRAF6ΔDC bone marrow chimera (ΔDC) for last 2 weeks. <b>(B)</b> Percentage of IL-13<sup>+</sup> CD4 T cells were shown in each organs of TRAF6ΔDC bone marrow chimera (ΔDC-BMC) in germ-free (GF) or germ-free under antibiotics (GF-Abx). <b>(C)</b> Lengths and masses of small intestines were measured in GF and GF-Abx TRAF6ΔDC bone marrow chimera (ΔDC-BMC). <b>(D)</b> Fibrosis markers (Acta2 and Igf-1) and Th2 cell cytokines (IL-13, IL-5, and IL-4) mRNA expression levels in the ileum region of small intestines from GF and GF-Abx TRAF6ΔDC bone marrow chimera (ΔDC-BMC). Histograms (mean ± SD) are representative of four independent experiments. SP, spleen; MLN, mesenteric lymph node. *p < 0.05; **p < 0.01; ***p < 0.001.</p

    data_sheet_1.docx

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    <p>A relatively high affinity/avidity of T cell receptor (TCR) recognition for self-peptide bound to major histocompatibility complex II (self-pMHC) ligands is a distinctive feature of CD4 T regulatory (Treg) cells, including their development in the thymus and maintenance of their suppressive functions in the periphery. Despite such high self-reactivity, however, all thymic-derived peripheral Treg populations are neither homogenous in their phenotype nor uniformly immune-suppressive in their function under steady state condition. We show here that based on the previously defined heterogeneity in the phenotype of peripheral Treg populations, Ly6C expression on Treg marks a lower degree of activation, proliferation, and differentiation status as well as functional incompetence. We also demonstrate that Ly6C expression on Treg in a steady state is either up- or downregulated depending on relative amounts of tonic TCR signals derived from its contacts with self-ligands. Interestingly, peripheral appearance and maintenance of these Ly6C-expressing Treg cells largely differed in an age-dependent manner, with their proportion being continuously increased from perinatal to young adult period but then being gradually declined with age. The reduction of Ly6C<sup>+</sup> Treg in the aged mice was not due to their augmented cell death but rather resulted from downregulation of Ly6C expression. The Ly6C downregulation was accompanied by proliferation of Ly6C<sup>+</sup> Treg cells and subsequent change into Ly6C<sup>−</sup> effector Treg with concomitant restoration of immune-suppressive activity. Importantly, we found that this phenotypic and functional change of Ly6C<sup>+</sup> Treg is largely driven by conventional effector T cell population. Collectively, these findings suggest a potential cross-talk between peripheral Treg subsets and effector T cells and provides better understanding for Treg homeostasis and function on maintaining self-tolerance.</p
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