32 research outputs found

    Effects of neonatal α-GC sensitization with RSV primary infection on airway hyperresponsiveness and viral clearance following re-infection in adulthood.

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    <p>(A) Airway hyperresponsiveness measured by methacholine test (Penh). Wild-type (wt) or CD1d knockout neonatal mice were infected with vehicle (Veh), α-GC, RSV, or α-GC with RSV and at age 8 weeks they were challenged with RSV. Four days after challenge, mice were given nebulized methacholine; 0 or 10 mg/mL. Data are expressed as mean ± SEM of 6 mice given 10 mg/mL of methacholine. **p<0.01. (B) Lungs were harvested and viral titers assessed by plaque assay on day 4 after challenge. Results are representative of two independent experiments. Data are expressed as mean ± SEM of 4–6 mice per group. **p<0.01; ***p<0.001; ****p<0.0001.</p

    Natural killer T cell sensitization during neonatal respiratory syncytial virus infection induces eosinophilic lung disease in re-infected adult mice

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    <div><p>Respiratory syncytial virus (RSV) is a major viral pathogen that causes severe lower respiratory tract infections in infants and the elderly worldwide. Infants with severe RSV bronchiolitis tend to experience more wheezing and asthma in later childhood. Because invariant natural killer T (iNKT) cells are associated with the asthma pathology, we investigated whether neonatal iNKT cells are involved in the aggravation of pulmonary diseases following RSV infection in mice. Intranasal exposure to the iNKT cell ligand α-galactosylceramide (α-GC) with RSV primary infection in neonatal mice elicited neither cytokine production (except for a slight increase of IL-5) nor pulmonary eosinophilia, despite the presence of both CD1d<sup>+</sup> cells and NKT cells. Interestingly, in adult mice re-infected with RSV, neonatal iNKT cell sensitization by α-GC during RSV primary infection resulted in much higher levels of pulmonary Th2 cytokines and elevated eosinophilia with airway hyperresponsiveness, whereas this was not observed in <i>cd1d</i> knockout mice. In contrast, α-GC priming of adults during RSV re-infection did not induce more severe airway symptoms than RSV re-infection in the absence of α-GC. α-GC co-administration during RSV primary infection facilitated RSV clearance regardless of age, but viral clearance following re-infection was not iNKT cell-dependent. This study clearly demonstrates that RSV-induced immune responses can be altered by iNKT cells, suggesting that neonatal iNKT cell sensitization during RSV primary infection is associated with exacerbation of pulmonary diseases following RSV re-infection in adulthood.</p></div

    Effects of neonatal α-GC sensitization with RSV primary infection on pulmonary immune responses following re-infection in adulthood.

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    <p>Wild-type or CD1d KO neonatal mice were i.n. administered PBS, α-GC, RSV, or α-GC+ RSV and challenged at 8 weeks of age with RSV. Lung tissue was formalin-fixed, paraffin-embedded, and stained with hematoxylin-eosin (H&E) (A) or periodic acid–Schiff (PAS) (B) on day4 after RSVA2 challenge. (C) Cytokine levels in BAL fluid were determined by luminex assay. Data are expressed as mean ± SEM of 5 or 6 mice per group. *p<0.05.</p

    Effects of adult NKT cell priming by α-GC during RSV re-infection.

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    <p>Neonate mice (7 days old) were infected with RSV and at age 6 weeks i.n. administered vehicle (Veh), α-GC, RSV, or α-GC with RSV. (A) Four days after infection, cells were isolated from BAL fluid and the ratio of eosinophils per granulocyte determined as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0176940#pone.0176940.g001" target="_blank">Fig 1</a>. Data are mean ± SEM with 5 or 6 mice per group. **p<0.01; ***p<0.001. (B) Histologic examination of lungs on day 4 after infection. Lung tissue was formalin-fixed, paraffin-embedded, and stained with hematoxylin-eosin (H&E) or periodic acid–Schiff (PAS). (C) Cytokine levels in BAL fluid were determined by luminex assay. Data are mean ± SEM of 5 or 6 mice per group. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001. (D) Airway hyperresponsiveness was measured by methacholine test (Penh) 4 days after infection when mice were given nebulized methacholine; 0 and 10 mg/mL. Data are expressed as mean ± SEM of 5 or 6 mice at 10 mg/mL of methacholine in each group.</p

    RSV clearance by intranasal (i.n.) α-GC co-administration at neonate or adult ages.

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    <p>BALB/c or CD1d KO mice were infected i.n. with RSV or α-GC+RSV at ages 7 days (neonates; A) or 6 weeks (adults; B). On day 4 or 7 after administration, lungs were harvested and lung homogenates were assessed by plaque assay. Data are shown as mean ± SEM with 4–8 mice per group (A) and 3–5 mice per group (B). **p<0.01; ***p<0.001; PFU, plaque forming units.</p

    Cytokine production in BAL fluid in response to RSV infection with or without α-GC.

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    <p>(A) Mice were i.n. administered vehicle (Veh), α-GC, RSV, or α-GC+RSV at ages 7 days and 6 weeks. BAL fluid was collected from lungs 4 days after infection. Cytokine levels in BAL fluid were determined by luminex and cytometric bead assay. Data are expressed as mean ± SEM of 9–12 mice in each group of neonates and of 3 or 4 mice in each group of adults. *p<0.05</p

    Effects of neonatal α-GC sensitization with RSV primary infection on eosinophils following re-infection in adulthood.

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    <p>Wild-type or CD1d KO neonatal mice were i.n. administered PBS, α-GC, RSV, or α-GC+ RSV and challenged at 8 weeks of age with RSV. Four days after challenge, cells were isolated from the BAL fluid, and the ratio of eosinophils to granulocytes was determined as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0176940#pone.0176940.g001" target="_blank">Fig 1</a> (A). The total cell count and absolute number of eosinophils are shown (B). Data are mean ± SEM with 2–4 mice per group and representative of three independent experiments. **p<0.01.</p

    Pulmonary eosinophil infiltration and mucus production after NKT cell ligand administration in neonatal and adult mice.

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    <p>Mice were i.n. administered vehicle (Veh), α-GC, RSV, or α-GC+RSV at ages 7 days (neonates) (A) and 6 weeks (adults) (B). At 4 days after injection, cells were isolated from BAL fluid and stained with fluorescence-labeled anti-CD45, CD11c, and Siglec-F antibodies. Percentages of eosinophils, namely CD11c<sup>-</sup>SiglecF<sup>+</sup> cells, among granulocytes are shown. All data are mean ± SEM. (A) Data are for 6–8 mice per group using pooled samples from 2 neonatal mice and are representative of 4 independent experiments. *p<0.05; **p<0.01 (B) Data are for 4 or 5 mice per group and are representative of 2 independent experiments. Lung tissues in left and right panels, respectively, are stained with hematoxylin-eosin (H&E) and periodic acid–Schiff (PAS).</p

    CD1d expressing and NKT cells in neonatal and adult mice.

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    <p>(A) Mice were i.n. administered vehicle, α-GC, RSV, or α-GC+RSV at ages 7 days (A) and 6 weeks (B). After 4 days, lungs were harvested and stained with fluorescence-labeled anti-CD1d, CD3ε, and CD49b antibodies. Percentages of CD1d-positive or NKT cells (CD3<sup>+</sup>CD49b<sup>+</sup>) are shown. Data are shown as mean ± SEM with 3–7 mice per group (A) and with 3 or 4 mice per group (B). *p<0.05; ***p<0.001.</p

    SL immunization with DelNS1 vaccine activates mucosal and extramucosal CD4+ T cells.

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    <p>CFSE-labeled CD4<sup>+</sup> T cells from HA-TCR transgenic mice were adoptively transferred into naïve mice. One day later, recipients were given SL 2×10<sup>7</sup> pfu of Delta H1N1, 2×10<sup>5</sup> pfu of mouse-adapted wt live or 40 µg of formalin-inactivated A/PR8. Three days after the immunization, proliferating (CFSE stained) HA-TCR CD4<sup>+</sup> T cells were detected in cervical lymph nodes (CLN), lungs, mediastinal lymph nodes (MdLN) and spleens by FACS analysis. The data are representative of two experiments showing similar results.</p
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