Gene-Environment Interactions with Ozone and Non-atopic Asthma

PROJECT SUMMARY Epidemiologic studies have provided evidence that ozone (O3) exposure is associated with the development of asthma, in particular the non-atopic (non-allergic) sub-type.

These findings are supported by clinical studies showing that ambient O3 exposure is associated with eosinophilic airway inflammation in non-atopic children.

Laboratory studies have shown that O3 exposure causes mucous cell metaplasia/hyperplasia in non-human primates, rats and mice.

This includes recent results from our groups demonstrating that classical inbred strains of mice exposed to 0.8 ppm O3 (4 hours/day) for 9 days develop eosinophilic airway inflammation in addition to mucous cell hyperplasia.

Thus, data in three species and humans provide strong rationale for studies geared towards understanding the underlying mechanisms of the O3 ® non-atopic asthma association. We take a genetic approach to the question of mechanism.

We surveyed strains from the Collaborative Cross (CC) mouse population, a new panel of recombinant inbred strains and identified one strain, CC002, that exhibited much higher levels of eosinophilic inflammation after repeated O3 exposure than any other strain tested to date.

O3 exposed CC002 mice also developed mucous cell metaplasia and airway hyper- responsiveness (AHR), two other hallmark phenotypes of asthma.

Thus, CC002 provides a new genetic model that we can exploit to identify mechanisms underlying the of O3-induced non-atopic asthma association.

In the first Aim, we will test the hypothesis that CC002?s exaggerated response to O3 exposure are due to dysregulated epithelial signaling via alarmins that in turn stimulates type 2 innate lymphoid cells (ILC2s).

We will determine if epithelial-derived innate cytokines (IL-1a/b, IL-25, IL-33, TSLP) are increased in O3-exposed CC002 mice compared to non-responder strains, and if neutralization of these cytokines attenuates O3-induced airway inflammation and AHR.

Next, we will determine if ILC2 numbers and activation are increased in O3- exposed CC002 mice, and if depletion of ILC2s or inhibition of ILC2-derived cytokines (IL-5, IL-13) ameliorates O3-induced airway inflammation and AHR in CC002 mice.

To comprehensively characterize molecular responses to O3 in epithelia, ILC2s, and other leukocytes, we will perform single cell RNA-sequencing.

Using this innovative and powerful approach, we will quantify differences in gene expression and cellular composition +/- O3, both within and between strains.

We will also examine the specificity of CC002?s response by testing whether this strain exhibits exaggerated responses to other exposures, including endotoxin, cigarette smoke, and house dust mite allergen. In the second aim, we will identify the genetic loci that render CC002 sensitive to O3.

We will perform quantitative trait locus mapping to identify chromosomal regions harboring CC002 alleles associated with O3 sensitivity, followed by bioinformatic analyses to identify high priority candidate genes at each locus.

In total, our work will reveal mechanisms underlying the association between O3 exposure and the development of non-atopic asthma phenotypes and identify potential pathways to target therapeutically.