30 research outputs found

    The Role of Peroxisome Proliferator-Activated Receptor γ in Immune Responses to Enteroaggregative <em>Escherichia coli</em> Infection

    Get PDF
    <div><p>Background</p><p>Enteroaggregative <i>Escherichia coli</i> (EAEC) is recognized as an emerging cause of persistent diarrhea and enteric disease worldwide. Mucosal immunity towards EAEC infections is incompletely understood due in part to the lack of appropriate animal models. This study presents a new mouse model and investigates the role of peroxisome proliferator-activated receptor gamma (PPARγ) in the modulation of host responses to EAEC in nourished and malnourished mice.</p> <p>Methods/Principal Findings</p><p>Wild-type and T cell-specific PPARγ null C57BL/6 mice were fed protein-deficient diets at weaning and challenged with 5×10<sup>9</sup>cfu EAEC strain JM221 to measure colonic gene expression and immune responses to EAEC. Antigen-specific responses to <i>E. coli</i> antigens were measured in nourished and malnourished mice following infection and demonstrated the immunosuppressive effects of malnutrition at the cellular level. At the molecular level, both pharmacological blockade and deletion of PPARγ in T cells resulted in upregulation of TGF-β, IL-6, IL-17 and anti-microbial peptides, enhanced Th17 responses, fewer colonic lesions, faster clearance of EAEC, and improved recovery. The beneficial effects of PPARγ blockade on weight loss and EAEC clearance were abrogated by neutralizing IL-17 <i>in vivo</i>.</p> <p>Conclusions</p><p>Our studies provide <i>in vivo</i> evidence supporting the beneficial role of mucosal innate and effector T cell responses on EAEC burden and suggest pharmacological blockade of PPARγ as a novel therapeutic intervention for EAEC infection.</p> </div

    Immune responses during enteroaggregative Escherichia coli (EAEC) infection in peroxisome proliferator-activated receptor γ (PPARγ)-deficient mice associated with bacterial clearance.

    No full text
    <p>Antigen specific recall responses of spleenocytes from mice infected with EAEC were measured ex vivo using the lymphocyte blastogenesis test. EAEC JM221 whole cell and whole cell sonicate were used in parallel to two negative controls, <i>E. coli</i> HS and mutant EAEC Aff/I strains as well as one positive control, concanavalin A (ConA). Lymphocyte proliferation is expressed stimulation indexes which are calculated by dividing the counts per minute (CPM) of antigen-stimulated wells by the CPM of unstimulated wells (A). IL-17 expression was assessed in colonic lamina propria (B) and whole blood (C) CD4+ T cells by flow cytometry and in the colon by quantitative real time RT-PCR (D) 14 days PI. Mice per group: n = 10. Asterisks indicate values where differences are statistically significant (<i>p</i><0.05) while bars connect groups where comparisons are made.</p

    Early beneficial effects of PPARγ deficiency in T cells during enteroaggregative Escherichia coli (EAEC) challenge.

    No full text
    <p>Growth retardation in wild type (A) and T cell specific PPARγ deficient mice (B) is expressed as percent growth from day 0 after challenge. Gene expression for IL-6 and TNF-α in colonic tissue of malnourished C57BL/6 and PPARγ CD4cre+ mice was analyzed using quantitative real-time RT-PCR on day 5 PI (C). Representative photomicrographs of colonic specimens of infected mice at 5 or 14 days PI in infected wild type mice (D,E,I,J), infected mice lacking PPARγ expression in T cells (F,G,K,L), and uninfected controls (H,M). The top panel corresponds to nourished mice whereas the bottom panel corresponds to malnourished mice. Original magnification 200×. Boxes and arrows are areas where an amplified image (400×) is provided to emphasize examples of leukocyte infiltration, mucosal thickening, goblet cell hyperplasia, and vasodilation. Mice per group: n = 8. Asterisks indicate values where differences are statistically significant (<i>p</i><0.05).</p

    Pharmacological blockade of peroxisome proliferator-activated receptor γ (PPARγ) associated with antimicrobial response and bacterial clearance.

    No full text
    <p>Enteroaggregative <i>Escherichia coli</i> (EAEC) burden in colon was assessed by quantitative real time RT-PCR using bacterial DNA isolated from feces of infected mice treated with PPARγ antagonist GW9662 (n = 9) or left untreated (n = 9). Data is presented as CFU/mg of tissue. S100A8 and S100A9 gene expression was analyzed in colonic tissue from C57BL/6 malnourished mice at day 5 days PI (n = 10) using quantitative real-time RT-PCR (B and C). S100 proteins are presented as values normalized to β-actin. Asterisks indicate values where differences are statistically significant (<i>p</i><0.05).</p

    Gene expression suggests a T helper 17 response in mice when peroxisome proliferator-activated receptor γ (PPARγ) is antagonized.

    No full text
    <p>Gene expression data from colonic tissue of malnourished C57BL/6 mice was analyzed using quantitative real-time RT-PCR and reported as values normalized to β-actin. IL-6, IL-1β, MCP-1, CCL20, and CXCL1 were quantified at day 5PI (mice per group: n = 10) (A–E) while IL-6, TGF-β, and IL-17 were quantified 14 days PI (n = 10) (F–H). Asterisks indicate values where differences are statistically significant (<i>p</i><0.05) while bars connect groups where comparisons are made.</p

    Neutralization of IL-17 abrogates the beneficial effects of GW9662 on weight loss and bacterial burden.

    No full text
    <p>Growth retardation in infected wild type mice is expressed as percent growth from day 0 after challenge (A). Enteroaggregative <i>Escherichia coli</i> (EAEC) burden in the colon was assessed by quantitative real time RT-PCR using bacterial DNA isolated from feces of infected mice treated with 1 µM PPARγ antagonist GW9662 (n = 3), 50 µg anti-IL17 and 1 µM GW9662 combined (n = 3) or untreated (n = 3). Asterisks indicate values where differences are statistically significant (<i>p</i><0.05), NS signifies no significant difference, and bars are present to indicate significance between groups.</p

    Modeling the Role of Peroxisome Proliferator-Activated Receptor γ and MicroRNA-146 in Mucosal Immune Responses to <em>Clostridium difficile</em>

    Get PDF
    <div><p><em>Clostridium difficile</em> is an anaerobic bacterium that has re-emerged as a facultative pathogen and can cause nosocomial diarrhea, colitis or even death. Peroxisome proliferator-activated receptor (PPAR) γ has been implicated in the prevention of inflammation in autoimmune and infectious diseases; however, its role in the immunoregulatory mechanisms modulating host responses to <em>C. difficile</em> and its toxins remains largely unknown. To characterize the role of PPARγ in <em>C. difficile</em>-associated disease (CDAD), immunity and gut pathology, we used a mouse model of <em>C. difficile</em> infection in wild-type and T cell-specific PPARγ null mice. The loss of PPARγ in T cells increased disease activity and colonic inflammatory lesions following <em>C. difficile</em> infection. Colonic expression of IL-17 was upregulated and IL-10 downregulated in colons of T cell-specific PPARγ null mice. Also, both the loss of PPARγ in T cells and <em>C. difficile</em> infection favored Th17 responses in spleen and colonic lamina propria of mice with CDAD. MicroRNA (miRNA)-sequencing analysis and RT-PCR validation indicated that miR-146b was significantly overexpressed and nuclear receptor co-activator 4 (NCOA4) suppressed in colons of <em>C. difficile</em>-infected mice. We next developed a computational model that predicts the upregulation of miR-146b, downregulation of the PPARγ co-activator NCOA4, and PPARγ, leading to upregulation of IL-17. Oral treatment of <em>C. difficile</em>-infected mice with the PPARγ agonist pioglitazone ameliorated colitis and suppressed pro-inflammatory gene expression. In conclusion, our data indicates that miRNA-146b and PPARγ activation may be implicated in the regulation of Th17 responses and colitis in <em>C. difficile</em>-infected mice.</p> </div

    The loss of PPARγ in T cells regulates colonic cytokine expression of mice infected with <i>Clostridium difficile</i>.

    No full text
    <p>Colonic expression of interleukin 10 (IL-10) (A), interleukin 17 (IL-17) (B), monocyte chemoattractant protein 1 (MCP-1) (C) and tumor necrosis factor (TNF-α) (D) were assessed by real-time quantitative RT-PCR in wild type and T cell PPARγ null mice infected with <i>C. difficile</i> (n = 8). Data are represented as mean ± standard error. Points with an asterisk are significantly different when compared to the wild type control group (<i>P</i><0.05).</p

    Figure 1

    No full text
    <p>A. Body weight gain (% initial weight) from experimental uninfected and undernourished groups under a low protein diet. APOE 4/4 targeted replacement (APOE 4/4 TR) mice (n = 17) showed 643 a better growth response in comparison with APOE 3/3 targeted replacement (APOE 3/3 TR) mice (n = 8). <b>B.</b> Body weight gain (% initial infection weight) from experimental mice challenged by a compounded malnutrition and <i>Cryptosporidium parvum</i> insult. Undernourished mice were orally inoculated with 10<sup>7</sup>- unexcysted oocysts diluted in 100 µl of PBS. APOE deficient mice show impaired growth following <i>Cryptosporidium parvum</i> infection as compared to the other groups. Results are shown as mean ±SEM.</p

    Computational modeling of mucosal immune responses to <i>Clostridium difficile</i> infection.

    No full text
    <p>CellDesigner-based illustration of the Complex Pathway SImulator model of the model for Clostridium difficile immune response (A). The model represents the interaction between <i>C. difficile</i>, miRNA-146, nuclear receptor coactivator 4 (NCOA4), peroxisome proliferator-activated receptor γ (PPAR γ), interleukin 10 (IL-10) and interleukin 17 (IL-17) in Systems Biology Markup Language format. Inhibition is represented in red and activation in green. COPASI steady state scan showing the variation on the species concentrations with increasing computational concentration of <i>C. difficile</i> (B). In silico simulations show how increasing concentrations of <i>C. difficile</i> increase miRNA-146b levels, thus decreasing NCOA4 and PPAR γ. In line with the experimental data, IL-17 expression also increases with the infection.</p
    corecore