63 research outputs found

    Genetic variants associated with increased risk for IA and probable pathogenetic mechanism(s) of susceptibility.

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    <p>SNP – single nucleotide polymorphism; OR – odds ratio; HSCT – hematopoietic stem cell transplantation; D – donor; R – recipient; RAGE – receptor for advanced glycation end products; CXCL – chemokine (C-X-C motif) ligand; MBL – mannose-binding lectin; MASP – MBL-associated serine protease; PLG – plasminogen; TLR – toll-like receptor; TNFR – tumor necrosis factor receptor; n.a. – not available; VNTR – variable number of tandem repeats.</p>1<p>For patients that underwent HSCT, the source of the variant (donor, recipient, or both) associated with susceptibility to IA is indicated.</p>2<p>The controls for the association reported were not hematological patients, but healthy subjects of comparable Dutch ancestry.</p>3<p>Association with increased susceptibility to IA was observed for a haplotype comprising SNPs in the IL-1 gene cluster, namely <i>IL1A</i> rs1800587/<i>IL1B</i> rs16944/<i>IL1RN</i> VNTR 86-bp(n), but not for single loci.</p>4<p>Association with increased susceptibility to IA was observed for the absence of the ACC haplotype in rs1800896, rs1800871, and rs1800872, respectively.</p>5<p>‘MBL-low genotypes’ correspond to a group of genotypes denoted by letters (O/O and LXA/O). LX represents an MBL promoter haplotype comprising SNPs of the MBL2 gene at positions −550 (H/L) and −221 (Y/X), known to influence transcription rates and to result in low concentrations of serum MBL. Nonsynonymous variants are collectively named O (including amino acid replacements at codons 52, 54, or 57) and cause a reduction of the MBL levels due to impaired assembly of MBL monomers into functional oligomers. A indicates the wild type.</p>6<p>Association results regard the comparisons DD <i>vs.</i> DN and DD <i>vs.</i> NN, respectively.</p>7<p>Association results regard R80T, N248S and S249P, and R80T or N248S and S249P, respectively.</p>8<p>Association with increased susceptibility to IA was observed for a <i>TLR4</i> haplotype (termed S4) that included both D299G and T399I.</p

    Effect of melanin content in the immune reactivity of <i>A. fumigatus</i> clinical isolates.

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    <p>Ability of DC to discriminate among different melanin color mutants and clinical isolates of the <i>Aspergillus</i> species was tested as differential cytokine production. DCs were cultured with live conidia of wt CEA10 strain, different knock out (KO) mutants, Aspergillosis isolates or without any stimulus (unstimulated, us) for 24 hours and supernatants used for TNFα, IL-10, IL-1β, IL-6, IL-12p70 and IL-23 measurements. *p≤0.05, **p≤0.01 and ***p≤0.001, ****p≤0.0001, mutants strains <i>vs</i> CEA10 strain, isolates <i>vs</i> CEA10 strain.</p

    Immune response to different WT <i>Aspergillus</i> strains.

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    <p>Different WT strains elicit diverse inflammatory responses and prime peculiar adaptive Th response. (A) DCs were cultured with UV-killed conidia or hyphae of WT strains for 24 hours or without any stimuli (unstimulated, us) and supernatants were used for TNFα, IL-10, IL-1β, IL-6, IL-12p70 and IL-23 measurements. Data are represented as mean+SD (N = 6), *p≤0.05, **p≤0.01, INF<sup>−</sup> conidia <i>vs</i> INF<sup>+</sup>conidia; <sup>§</sup>p≤0.05, <sup>§§</sup>p≤0.01, INF<sup>−</sup> hyphae <i>vs</i> INF<sup>+</sup> hyphae. Complete statistically significant p-values are collected in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056651#pone.0056651.s002" target="_blank">Table S2</a>. (B,C) Healthy PBMCs were cultured with live conidia or without any stimulus, and cytokine protein (A) and transcript (B) levels were measured. Data are represented as mean+SD (N = 6), *p≤0.05, **p≤0.01, INF<sup>− </sup><i>vs</i> INF<sup>+</sup> strains.</p

    Immune response to different <i>Aspergillus</i> mutant strains.

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    <p>Different mutant strains elicit diverse inflammatory responses and prime peculiar adaptive T<sub>H</sub> response. (A) DCs were cultured with UV-killed conidia or hyphae for 24 hours or without any stimuli (unstimulated, us) and supernatants were used for TNFα, IL-10, IL-1β, IL-6, IL-12p70 and IL-23 measurements. Data are represented as mean+SD (N = 6), *p≤0.05, **p≤0.01, conidial mutant strains <i>vs</i> the conidial WT strain; <sup>§</sup>p≤0.05, <sup>§§</sup>p≤0.01, hyphal mutant strains vs hyphal WT strain. Complete statistically significant p-values are collected in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056651#pone.0056651.s002" target="_blank">Table S2</a>. (B,C) Healthy PBMCs were cultured with live conidia or without any stimulus, and cytokine protein and transcript levels were measured. Data are represented as mean+SD (N = 6), *p≤0.05, **p≤0.01, mutants strain <i>vs</i> WT strain. (D) C57BL/6 mice were infected intranasally with different strains of <i>A. fumigatus</i> (6 mice/group). Survival (%), fungal growth (mean log<sub>10</sub> CFU ± SE, N = 3) in the lungs and brains of infected mice were assessed at different days post-infection (dpi). The CFUs between wild-type and the corresponding mutant strains were statistically significant (p-values ranging from ≤0.01 to ≤0.001). Lung histology (PAS staining) and BAL morphometry [%, mean ± SD, of mononuclear (MNC) or polymorphonuclear (PMN) cells] were done at 4 dpi. Representative images of two independent experiments were depicted; bars indicate magnifications. Total lung RNA was extracted at 4 dpi and the relative expression of <i>Il1β</i>, <i>Cxcl1</i> and <i>Cxcl2</i> genes was assessed by RT-PCR. Lung homogenates at 4 dpi were tested for levels of IL-17A, IFN-γ, and IL-10 by specific ELISA (mean values ± SD, N = 3). *p≤0.05, **p≤0.01 and ***p≤0.001, wild-type strains <i>vs</i> uninfected mice or mutants strains <i>vs</i> the wild-type strain.</p

    Immune response to different <i>Aspergillus</i> color mutants.

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    <p>Different mutant strains elicit diverse inflammatory responses and prime peculiar adaptive T<sub>H</sub> response. (A) DCs were cultured with UV-killed conidia or hyphae for 24 hours or without any stimuli (unstimulated, us) and supernatants were used for TNFα, IL-10, IL-1β, IL-6, IL-12p70 and IL-23 measurements. Data are represented as mean+SD (N = 6), *p≤0.05, **p≤0.01, conidial mutant strains <i>vs</i> conidial WT strain; <sup>§</sup>p≤0.05, <sup>§§</sup>p≤0.01, hyphal mutant strains vs hyphal WT strain. Complete statistically significant p-values are collected in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056651#pone.0056651.s002" target="_blank">Table S2</a>. (B,C) Healthy PBMCs were cultured with live conidia or without any stimulus, and cytokine protein and transcript levels were measured. Data are represented as mean+SD (N = 6), *p≤0.05, **p≤0.01, mutants strain <i>vs</i> WT strain.</p

    <i>In vivo</i> immune response to <i>A. fumigatus</i> mutant strains.

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    <p>Different mutant strains elicit diverse inflammatory responses and prime peculiar adaptive T<sub>H</sub> response. (A) DCs were cultured with UV-killed conidia or hyphae for 24 hours or without any stimuli (unstimulated, us) and supernatants were used for TNFα, IL-10, IL-1β, IL-6, IL-12p70 and IL-23 measurements. Data are represented as mean+SD (N = 6), *p≤0.05, **p≤0.01, conidial mutant strains <i>vs</i> the conidial WT strain; <sup>§</sup>p≤0.05, <sup>§§</sup>p≤0.01, hyphal mutant strains vs hyphal WT strain. Complete statistically significant p-values are collected in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0056651#pone.0056651.s002" target="_blank">Table S2</a>. (B,C) Healthy PBMCs were cultured with live conidia or without any stimulus, and cytokine protein and transcript levels were measured. Data are represented as mean+SD (N = 6), *p≤0.05, **p≤0.01, mutants strain <i>vs</i> WT strain. (D) C57BL/6 mice were infected intranasally with different strains of <i>A. fumigatus</i> (6 mice/group). Survival (%), fungal growth (mean log<sub>10</sub> CFU ± SE, N = 3) in the lungs and brains of infected mice were assessed at different days post-infection (dpi). The CFUs between wild-type and the corresponding mutant strains were statistically significant (p values ranging from ≤0.01 to ≤0.001). Lung histology (PAS staining) and BAL morphometry [%, mean ± SD, of mononuclear (MNC) or polymorphonuclear (PMN) cells] were done at 4 dpi. Representative images of two independent experiments were depicted; bars indicate magnifications. Total lung RNA was extracted at 4 dpi and the relative expression of <i>Il1β</i>, <i>Cxcl1</i> and <i>Cxcl2</i> genes was assessed by RT-PCR. Lung homogenates at 4 dpi were tested for levels of IL-17A, IFN-γ, and IL-10 by specific ELISA (mean values ± SD, N = 3). *p≤0.05, **≤0.01 and ***p≤0.001, wild-type strains <i>vs</i> uninfected mice or mutants strains <i>vs</i> the wild-type strain.</p

    <i>In vivo</i> immune response to <i>A. fumigatus</i> color mutants.

    No full text
    <p>C57BL/6 mice were infected intranasally with different strains of <i>A. fumigatus</i> (6 mice/group). Survival (%), fungal growth (mean log<sub>10</sub> CFU ± SE, N = 3) in the lungs and brains of infected mice were assessed at different days post-infection (dpi). The CFUs between wild-type and the corresponding mutant strains were statistically significant (p values ranging from ≤0.01 to ≤0.001). Lung histology (PAS staining) and BAL morphometry [%, mean ± SD, of mononuclear (MNC) or polymorphonuclear (PMN) cells] were done at 4 dpi. Representative images of two independent experiments were depicted; bars indicate magnifications. Total lung RNA was extracted at 4 dpi and the relative expression of <i>Il1β</i>, <i>Cxcl1</i> and <i>Cxcl2</i> genes was assessed by RT-PCR. Lung homogenates at 4 dpi were tested for levels of IL-17A, IFN-γ, and IL-10 by specific ELISA (mean values ± SD, N = 3). *p≤0.05, **p≤0.01 and ***p≤0.001, wild-type strains <i>vs</i> uninfected mice or mutants strains <i>vs</i> the corresponding wild-type strain.</p

    <i>In vivo</i> immune reactivity to <i>A. fumigatus</i> WT strains.

    No full text
    <p>C57BL/6 mice were infected intranasally with different strains of <i>A. fumigatus</i> (6 mice/group). Survival (%), fungal growth (mean log<sub>10</sub> CFU ± SE, N = 3) in the lungs and brains of infected mice were assessed at different days post-infection (dpi). The CFUs between wild-type and the corresponding mutant strains were statistically significant (p values ranging from ≤0.01 to ≤0.001). Lung histology (PAS staining) and BAL morphometry [%, mean ± SD, of mononuclear (MNC) or polymorphonuclear (PMN) cells] were done at 4 dpi. Representative images of two independent experiments were depicted; bars indicate magnifications. Total lung RNA was extracted at 4 dpi and the relative expression of <i>Il1β</i>, <i>Cxcl1</i> and <i>Cxcl2</i> genes was assessed by RT-PCR. Lung homogenates at 4 dpi were tested for levels of IL-17A, IFN-γ, and IL-10 by specific ELISA (mean values ± SD, N = 3). *p≤0.05, **p≤0.01 and ***p≤0.001, wild-type strains <i>vs</i> uninfected mice.</p

    Binding of ficolin-A to <i>E. coli</i> using confocal laser scanning fluorescence microscopy.

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    <p>Ficolin-A (5 µg/ml) was incubated with <i>E. coli</i> EC2 (A) or <i>E. coli</i> EC3 (B). The bacteria (4×10<sup>6</sup> cells) were incubated with or without ficolin-A. Ficolin-A binding was only observed to <i>E. coli</i> EC2 and this binding was found on aggregated bacteria. The binding was detected with a monoclonal rat anti-mouse ficolin-A antibody.</p

    Binding of ficolin-A to bacterial components.

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    <p>The binding of ficolin-A (0.2 µg) to acBSA was inhibited with sterile filtrated overnight bacterial growth medium from some of the bacteria (A). No differences were observed when the growth medium was heat inactivated at 96°C (B). Purified LPS was coated on polystyrene plates and serum from ficolin-B knock-out serum diluted 1∶10 in Barb-T buffer was added (C). Ficolin-A binding to LPS from <i>E. coli</i> (EC2) was detected in a dose-dependent manner. Error bars indicate standard deviations of duplicates. Shown is one representative experiment of three.</p
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