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Inflammatory lung diseases affect men and women disproportionately, suggesting that fluctuations of circulating hormone levels mediate inflammatory responses. Studies have shown that ozone exposure contributes to lung injury and impairment of innate immunity with differential effects in men and women. Here, we hypothesized that 17β-estradiol enhances inflammation and airway hyperresponsiveness (AHR), triggered by ozone exposure, in the female lung. We performed gonadectomy and hormone treatment (17β-estradiol, 2 wk) in C57BL/6J female and male mice and exposed animals to 1 ppm of ozone or filtered air for 3 h. Twenty-four hours later, we tested lung function, inflammatory gene expression, and changes in bronchoalveolar lavage fluid (BALF). We found increased AHR and expression of inflammatory genes after ozone exposure. These changes were higher in females and were affected by gonadectomy and 17β-estradiol treatment in a sex-specific manner. Gonadectomized male mice displayed higher AHR and inflammatory gene expression than controls exposed to ozone; 17β-estradiol treatment did not affect this response. In females, ovariectomy reduced ozone-induced AHR, which was restored by 17β-estradiol treatment. Ozone exposure also increased BALF lipocalin-2, which was reduced in both male and female gonadectomized mice. Treatment with 17β-estradiol increased lipocalin-2 levels in females but lowered them in males. Gonadectomy also reduced ozone-induced expression of lung IL-6 and macrophage inflammatory protein-3 in females, which was restored by treatment with 17β-estradiol. Together, these results indicate that 17β-estradiol increases ozone-induced inflammation and AHR in females but not in males. Future studies examining diseases associated with air pollution exposure should consider the patient’s sex and hormonal status.

Bronchopulmonary dysplasia (BPD) is a form of chronic lung disease that develops in neonates as a consequence of preterm birth, arrested fetal lung development, and inflammation. The incidence of BPD remains on the rise as a result of increasing survival of extremely preterm infants. Severe BPD contributes to significant health care costs and is associated with prolonged hospitalizations, respiratory infections, and neurodevelopmental deficits. In this study, we aimed to detect novel biomarkers of BPD severity. We collected tracheal aspirates (TAs) from preterm babies with mild/moderate (n = 8) and severe (n = 17) BPD, and we profiled the expression of 1048 miRNAs using a PCR array. Associations with biological pathways were determined with the Ingenuity Pathway Analysis (IPA) software. We found 31 miRNAs differentially expressed between the two disease groups (2-fold change, false discovery rate (FDR) < 0.05). Of these, 4 miRNAs displayed significantly higher expression levels, and 27 miRNAs had significantly lower expression levels in the severe BPD group when compared to the mild/moderate BPD group. IPA identified cell signaling and inflammation pathways associated with miRNA signatures. We conclude that TAs of extremely premature infants contain miRNA signatures associated with severe BPD. These may serve as potential biomarkers of disease severity in infants with BPD.

Bronchopulmonary dysplasia (BPD) is a form of chronic lung disease that develops in neonates as a consequence of preterm birth and arrested fetal lung development. The incidence of BPD remains on the rise, as a result of increasing survival of extremely preterm infants. Severe BPD contributes to significant health care costs and is associated with prolonged hospitalizations, respiratory infections, and neurodevelopmental deficits. In this study, we aimed to detect novel biomarkers of severe BPD. We collected tracheal aspirates (TA) from preterm babies with mild/moderate (n = 8) and severe (n = 17) BPD, and we profiled the expression of 1048 miRNAs using a PCR array. Associations with biological pathways were determined with the Ingenuity Pathway Analysis (IPA) software. We found 31 miRNAs differentially expressed between the two disease groups (2-fold change, FDR &amp;lt; 0.05). Of these, 4 miRNAs displayed significantly higher expression levels, and 27 miRNAs had significantly lower expression levels in the severe BPD vs. the mild/moderate BPD group. IPA identified cell signaling and inflammation pathways associated with miRNA signatures. We conclude that TAs of extreme premature infants contain miRNA signatures associated with severe BPD. These signatures may serve as biomarkers of disease severity in infants with BPD.

Background Asthma is a chronic airway disease that leads to compromised lung function and affects more than 300 million patients worldwide, leading to >180,000 deaths each year, many of which are avoidable with proper treatment and care. Despite significant advances in diagnostic and treatment of the disease, asthma continues to be a significant health problem and an economic burden. Epidemiological studies in adult populations have revealed sex differences in asthma morbidity, with higher incidence in females than males. The causes of this disparity are still unclear, but studies have shown that sex‐specific signaling pathways regulated by hormones contribute to differences in immune cell activation and airway reactivity in response to different environmental stimuli. Methods With the goal of understanding the mechanisms associated with the observed sex disparities in allergen‐induced asthma, we challenged both male and female adult C57BL/6J mice with a house dust mite (HDM) solution from two species, Dermatophagoides pteronyssinus and Dermatophagoides farinae. The protocol consisted of five consecutive intranasal challenges with 25μg of HDM or control (PBS) followed by two days off, repeated for five weeks. The asthma phenotype was assessed by testing airway hyperresponsiveness (AHR) to methacholine challenge, and airway inflammation was characterized by cell differential assessment of bronchoalveolar lavage fluid obtained from experimental animals. Lung tissue was harvested for inflammatory mRNA expression determinations and histology. Airway remodeling was characterized using the two‐photon imaging technique, second harmonic generation (SHG), on lung tissue sections to determine collagen content in mouse airways (Velocity, imaging quantification software). Results We found that HDM promoted alterations in respiratory mechanics in challenged animals, including alterations in lung function measures that led to airway hyperresponsiveness (AHR) and increased parenchymal elastance in both males and females. While HDM‐challenged male mice showed higher AHR, airway collagen & elastin content, and BALF eosinophilia than females, challenged females displayed higher BALF neutrophilia and expression of type II pro‐inflammatory cytokines than males. Conclusion Our results indicate sex differences in both inflammatory and immune responses, as well as in airway structural changes induced by HDM exposure in mice. Characterization of the mechanisms leading to these sex differences will provide better insight into the causes of sex disparities observed in asthma prevalence and severity. Support or Funding Information This work was supported by NIH grants HL133520 (PS) and HL141618 (PS) and the APS Porter Fellowship (NF)

Ambient ozone, a product of photochemical reactions between volatile organic compounds and nitrogen oxides, is known to be one of the most dangerous air pollutants. Ozone inhalation can aggravate preexisting inflammatory lung diseases such as asthma, which are more frequently diagnosed in females than males. Despite this, the molecular mechanisms underlying the effects of ozone in the male and female lung have yet to be discovered. We hypothesized that exposure to ozone exerts differential inflammatory and immune responses in the lungs of male and females with asthma. To examine this, we treated adult male and female C57BL/6J mice with an allergen (house dust mite extract) intranasally for 5 weeks to trigger asthma. We then exposed mice to 2 ppm of ozone or filtered air (FA) for 3 hours, and collected lung tissue 24 hours later. We extracted lung RNA, and measure the expression of 92 immune response associated genes by PCR with the TaqMan® Array 96‐well Mouse Immune Response Plate (ThermoFisher). We also extracted DNA to determined lung microbiome profiles via 16S rRNA amplicon sequencing using a MiSeq instrument. The resulting paired‐end reads were processed using QIIME 2 2018.11. Amplicon sequencing units from DADA2 were assigned taxonomic identifiers based on the GreenGenes database release 13_08 using the QIIME 2 q2‐featureclassifier. Results were summarized, visualized through Principal Coordinate Analysis, and significance was estimated as implemented in QIIME2. Our preliminary results show that, after exposure to ozone, asthmatic male mice had higher lung expression of immune response genes (cell surface receptors and signaling molecules) whereas asthmatic females had higher expression of pro‐inflammatory cytokines, transcription factors, and regulators of immunity than mice exposed to FA. We also identified differential microbiome profiles between male and female asthmatic mice and in response to ozone, including a higher content of Firmicutes and a higher Firmicutes/Bacteriodetes ratio in males than females. Moreover, asthmatic females displayed a more pronounced lung bacterial diversity and higher relative abundance of Proteobacteria when compared to males. We conclude that ozone exposure triggers differential inflammatory/immune mechanisms in the male and female lungs of asthmatic mice, and that these mechanisms could be mediated by lung resident microbes.