Obese mice exhibit innate airway hyperresponsiveness (AHR), a feature of asthma. Tumor necrosis factor alpha (TNFα) is implicated in the disease progression and chronic inflammatory status of both obesity and asthma. TNF acts via two TNF receptors, TNFR1 and TNFR2. To examine the role of TNFR2 in the AHR observed in obese mice, we generated obese Cpefat mice that were either sufficient or deficient in TNFR2 (Cpefat and Cpefat/TNFR2−/− mice, respectively) and compared them with their lean controls (WT and TNFR2−/− mice). Compared to WT mice, Cpefat mice exhibited AHR to aerosolized methacholine (measured using the forced oscillation technique) which was ablated in Cpefat/TNFR2−/− mice. Bioplex or ELISA assay indicated significant increases in serum leptin, G-CSF, IL-7, IL-17A, TNFα, and KC in obese versus lean mice, as well as significant obesity-related increases in bronchoalveolar lavage fluid (BALF) G-CSF and IP-10, regardless of TNFR2 status. Importantly, BALF IL-17A was significantly increased over lean controls in Cpefat but not Cpefat/TNFR2−/− mice. Functional annotation clustering of significantly affected genes identified from microarray analysis comparing gene expression in lungs of Cpefat and WT mice, identified blood vessel morphogenesis as the gene ontology category most affected by obesity. This category included several genes associated with AHR, including endothelin and trkB. Obesity increased pulmonary mRNA expression of endothelin and trkB in TNFR2 sufficient but not deficient mice. Our results indicate that TNFR2 signaling is required for the innate AHR that develops in obese mice, and suggest that TNFR2 may act by promoting IL-17A, endothelin, and/or trkB expression.

Background: Acute ozone (O3) exposure results in greater inflammation and airway hyperresponsiveness (AHR) in obese versus lean mice.Objectives: We examined the hypothesis that these augmented responses to O3 are the result of greater signaling through tumor necrosis factor receptor 2 (TNFR2) and/or interleukin (IL)-13.Methods: We exposed lean wild-type (WT) and TNFR2-deficient (TNFR2–/–) mice, and obese Cpefat and TNFR2-deficient Cpefat mice (Cpefat/TNFR2–/–), to O3 (2 ppm for 3 hr) either with or without treatment with anti–IL-13 or left them unexposed.Results: O3-induced increases in baseline pulmonary mechanics, airway responsiveness, and cellular inflammation were greater in Cpefat than in WT mice. In lean mice, TNFR2 deficiency ablated O3-induced AHR without affecting pulmonary inflammation; whereas in obese mice, TNFR2 deficiency augmented O3-induced AHR but reduced inflammatory cell recruitment. O3 increased pulmonary expression of IL-13 in Cpefat but not WT mice. Flow cytometry analysis of lung cells indicated greater IL-13–expressing CD4+ cells in Cpefat versus WT mice after O3 exposure. In Cpefat mice, anti–IL-13 treatment attenuated O3-induced increases in pulmonary mechanics and inflammatory cell recruitment, but did not affect AHR. These effects of anti–IL-13 treatment were not observed in Cpefat/TNFR2–/– mice. There was no effect of anti–IL-13 treatment in WT mice.Conclusions: Pulmonary responses to O3 are not just greater, but qualitatively different, in obese versus lean mice. In particular, in obese mice, O3 induces IL-13 and IL-13 synergizes with TNF via TNFR2 to exacerbate O3-induced changes in pulmonary mechanics and inflammatory cell recruitment but not AHR.

Ozone is an air pollutant that causes pulmonary symptoms. In mice, ozone exposure causes pulmonary injury and increases bronchoalveolar lavage macrophages and neutrophils. We have shown that IL-17A is important in the recruitment of neutrophils after subacute ozone exposure (0.3 ppm for 24–72 h). We hypothesized that γδ T cells are the main producers of IL-17A after subacute ozone. To explore this hypothesis we exposed wildtype mice and mice deficient in γδ T cells (TCRδ−/−) to ozone or room air. Ozone-induced increases in BAL macrophages and neutrophils were attenuated in TCRδ−/− mice. Ozone increased the number of γδ T cells in the lungs and increased pulmonary Il17a mRNA expression and the number of IL-17A+ CD45+ cells in the lungs and these effects were abolished in TCRδ−/− mice. Ozone-induced increases in factors downstream of IL-17A signaling, including G-CSF, IL-6, IP-10 and KC were also decreased in TCRδ−/− versus wildtype mice. Neutralization of IL-17A during ozone exposure in wildtype mice mimicked the effects of γδ T cell deficiency. TNFR2 deficiency and etanercept, a TNFα antagonist, also reduced ozone-induced increases in Il17a mRNA, IL-17A+ CD45+ cells and BAL G-CSF as well as BAL neutrophils. TNFR2 deficient mice also had decreased ozone-induced increases in Ccl20, a chemoattractant for IL-17A+ γδ T cells. Il17a mRNA and IL-17A+ γδ T cells were also lower in obese Cpefat versus lean WT mice exposed to subacute ozone, consistent with the reduced neutrophil recruitment observed in the obese mice. Taken together, our data indicate that pulmonary inflammation induced by subacute ozone requires γδ T cells and TNFα-dependent recruitment of IL-17A+ γδ T cells to the lung.

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