Development of asthmatic inflammation in mice following early-life exposure to ambient environmental particulates and chronic allergen challenge

Abstract

Childhood exposure to environmental particulates increases the risk of development of asthma. The underlying mechanisms might include oxidant injury to airway epithelial cells (AEC). We investigated the ability of ambient environmental particulates to contribute to sensitization via the airways, and thus to the pathogenesis of childhood asthma. To do so, we devised a novel model in which weanling BALB/c mice were exposed to both ambient particulate pollutants and ovalbumin for sensitization via the respiratory tract, followed by chronic inhalational challenge with a low mass concentration of the antigen. We also examined whether these particulates caused oxidant injury and activation of AEC in vitro. Furthermore, we assessed the potential benefit of minimizing oxidative stress to AEC through the period of sensitization and challenge by dietary intervention. We found that characteristic features of asthmatic inflammation developed only in animals that received particulates at the same time as respiratory sensitization, and were then chronically challenged with allergen. However, these animals did not develop airway hyper-responsiveness. Ambient particulates induced epithelial injury in vitro, with evidence of oxidative stress and production of both pro-inflammatory cytokines and Th2-promoting cytokines such as IL-33. Treatment of AEC with an antioxidant in vitro inhibited the pro-inflammatory cytokine response to these particulates. Ambient particulates also induced pro-inflammatory cytokine expression following administration to weanling mice. However, early-life dietary supplementation with antioxidants did not prevent the development of an asthmatic inflammatory response in animals that were exposed to particulates, sensitized and challenged. We conclude that injury to airway epithelium by ambient environmental particulates in early life is capable of promoting the development of an asthmatic inflammatory response in sensitized and antigen-challenged mice. These findings are likely to be relevant to the induction of childhood asthma.

Fig. 1.

Airway inflammatory response. (A) Number of eosinophils in the airway epithelium, assessed morphologically. (B) Relative accumulation of eosinophils in lung tissue, assessed by peroxidase activity. (C) Percentage of neutrophils in BAL fluid. (D) Number of neutrophils in lung tissue, assessed by immunostaining. Data are mean ± s.e.m. (n=6-8 samples per group). Significant differences are shown as *P<0.05 and **P<0.01 compared with the naïve group and as ^#P<0.05 and ^##P<0.01 compared with the Sham/OVA group.

Fig. 2.

Cytokine concentrations in BAL fluid. Concentrations of (A) CXCL1, (B) CCL3 and (C) G-CSF were measured in BAL fluid. Data are mean ± s.e.m. (n=6-8 samples per group). Significant differences are shown as ***P<0.001 compared with the naïve group and as ^#P<0.05 compared with the Sham/OVA group.

Fig. 3.

Airway responsiveness to β-methacholine. Airway responsiveness was assessed by measurement of (A) transpulmonary resistance (R_L) and (B) dynamic compliance (C_dyn). Data are mean ± s.e.m. (n=6-8 samples per group). Differences are not statistically significant.

Fig. 4.

Expression of mRNA for enzymes involved in the response to oxidative stress by AEC exposed to medium alone, carbon black or ambient environmental particulates. Expression of (A) HO-1, (B) GSTP-1 and (C) GPx-2. Data are mean ± s.e.m. (n=4 samples per group). Significant differences are shown as *P<0.05 compared with medium alone and as ^#P<0.05 and ^##P<0.01 compared with carbon black.

Fig. 5.

Response of 4-week-old mice to intranasal administration of carbon black or ambient particulate matter. (A) Number of neutrophils in lung tissue, assessed by immunostaining. (B) Expression of mRNA for TNF-α. Data are mean ± s.e.m. (n=6-8 samples per group). Significant differences are shown as *P<0.05 compared with the naïve group and as ^#P<0.05 compared with carbon black.

Fig. 6.

Effect of early-life dietary supplementation with lycopene on inflammatory, cytokine and oxidative stress responses. (A) Percentage of neutrophils in BAL fluid. (B) Relative accumulation of eosinophils in lung tissue, as assessed by peroxidase activity. Significant differences compared with the naïve group are shown as *P<0.05 and **P<0.01.

Fig. 7.

Timeline of exposure to particulates, respiratory sensitization and inhalational challenges in the early-life model. Animals were administered ambient particulates on days 14 and 15 of life, then again on days 28 and 29 of life, and were sensitized with intranasal OVA at the latter time point. Commencing at day 49 of life, mice received repeated low-level challenges with aerosolized OVA for 4 weeks, followed by a single moderate-level challenge.