NAT1 genetic variation increases asthma risk in children with secondhand smoke exposure.

OBJECTIVE: We previously reported that children exposed to secondhand smoke (SHS) that carried variants in the NAT1 gene had over two-fold higher hair cotinine levels. Our objective was to determine if NAT1 polymorphisms confer increased risk for developing asthma in children exposed to SHS. METHODS: White participants in the Cincinnati Childhood Allergy and Air Pollution Study (n = 359) were genotyped for 10 NAT1 variants. Smoke exposure was defined by hair cotinine and parental report. Asthma was objectively assessed by spirometry and methacholine challenge. Findings were replicated in the Genomic Control Cohort (n = 638). RESULTS: Significant associations between 5 NAT1 variants and asthma were observed in the CCAAPS exposed group compared to none in the unexposed group. There was a significant interaction between NAT1 rs13253389 and rs4921581 with smoke exposure (p = 0.02, p = 0.01) and hair cotinine level (p = 0.048, p = 0.042). Children wildtype for rs4921581 had increasing asthma risk with increasing hair cotinine level, whereas those carrying the NAT1 minor allele had an increased risk of asthma regardless of cotinine level. In the GCC, 13 NAT1 variants were associated with asthma in the smoke-exposed group, compared to 0 in the unexposed group, demonstrating gene-level replication. CONCLUSIONS: Variation in the NAT1 gene modifies asthma risk in children exposed to secondhand-smoke. To our knowledge, this is the first report of a gene-environment interaction between NAT1 variants, smoke exposure, cotinine levels, and pediatric asthma. NAT1 genotype may have clinical utility as a biomarker of increased asthma risk in children exposed to smoke.

Biofilm propensity of Staphylococcus aureus skin isolates is associated with increased atopic dermatitis severity and barrier dysfunction in the MPAACH pediatric cohort.

BACKGROUND: Atopic dermatitis (AD) patients are often colonized with Staphylococcus aureus, and staphylococcal biofilms have been reported on adult AD skin lesions. The commensal S epidermidis can antagonize S aureus, although its role in AD is unclear. We sought to characterize S aureus and S epidermidis colonization and biofilm propensity and determine their associations with AD severity, barrier function, and epidermal gene expression in the first US early-life cohort of children with AD, the Mechanisms of Progression of Atopic Dermatitis to Asthma in Children (MPAACH). METHODS: The biofilm propensity of staphylococcal isolates was assessed by crystal violet assays. Gene expression of filaggrin and antimicrobial alarmins S100A8 and S100A9 was measured in keratinocyte RNA extracted from skin tape strips. Staphylococcal biofilms sampled from MPAACH skin were visualized using scanning electron microscopy. RESULTS: Sixty-two percent of staphylococcal isolates (sampled from 400 subjects) formed moderate/strong biofilms. Sixty-eight percent of subjects co-colonized with both staphylococcal species exhibited strains that formed cooperative mixed-species biofilms. Scanning electron microscopy verified the presence of staphylococcal biofilms on the skin of MPAACH children. Staphylococcus aureus strains showing higher relative biofilm propensity compared with S epidermidis were associated with increased AD severity (P = .03) and increased lesional and nonlesional transepidermal water loss (P = .01, P = .03). CONCLUSIONS: Our data suggest a pathogenic role for S aureus biofilms in AD. We found that strain-level variation in staphylococcal isolates governs the interactions between S epidermidis and S aureus and that the balance between these two species, and their biofilm propensity, has important implications for AD.

Disease-associated KIF3A variants alter gene methylation and expression impacting skin barrier and atopic dermatitis risk.

Single nucleotide polymorphisms (SNPs) in the gene encoding kinesin family member 3A, KIF3A, have been associated with atopic dermatitis (AD), a chronic inflammatory skin disorder. We find that KIF3A SNP rs11740584 and rs2299007 risk alleles create cytosine-phosphate-guanine sites, which are highly methylated and result in lower KIF3A expression, and this methylation is associated with increased transepidermal water loss (TEWL) in risk allele carriers. Kif3aK14∆/∆ mice have increased TEWL, disrupted junctional proteins, and increased susceptibility to develop AD. Thus, KIF3A is required for skin barrier homeostasis whereby decreased KIF3A skin expression causes disrupted skin barrier function and promotes development of AD.

Events in Normal Skin Promote Early-Life Atopic Dermatitis-The MPAACH Cohort.

BACKGROUND: Nonlesional skin in atopic dermatitis (AD) is abnormal, but the pathobiology of lesional and nonlesional skin and the definition of endotypes are poorly understood. OBJECTIVE: To define lesional and nonlesional endotypes of AD by building the first US-based early-life prospective cohort of children with AD, the Mechanisms of Progression from AD to Asthma in Children cohort. METHODS: We assessed lesional and nonlesional skin transepidermal water loss, filaggrin (FLG) and alarmin (S100A8, S100A9) expression, staphylococcal colonization, and patterns of aeroallergen and food sensitization to define nonlesional and lesional phenotypes and endotypes. RESULTS: Pathophysiologic changes were present in lesional and nonlesional skin and were associated with SCORing for Atopic Dermatitis. Nonlesional skin had features characteristic of diseased skin including low FLG and high alarmin expression, and increased colonization with Staphylococcus aureus. In a multivariate model, nonlesional, but not lesional, FLG expression was associated with the development of cosensitization and moderate to severe AD. Lesional skin was characterized by further deficits in FLG expression (P < .001), but alarmin expression was the same as observed in nonlesional skin. CONCLUSIONS: This study reveals that events in the nonlesional, not the lesional, skin promote the subsequent development of AD severity and cosensitization, which is a key risk factor for allergic comorbidities. Collectively, these data suggest the presence of a subclinical eczema endotype that may predispose to the development of allergic disease in the absence of overt eczema. This may represent a new definition of the atopic march that starts with skin barrier dysfunction rather than eczema.

Weighing in on asthma: Insights on BMI, magnesium, and hospitalizations from the Ohio Pediatric Asthma Repository.

Objective: Little is known about weight status and its effects on clinical course during hospitalization for asthma exacerbation. We sought to evaluate associations between weight status, specifically body mass index (BMI), with inpatient clinical course and clinical history.Methods: We retrospectively analyzed data from 2012 to 2013 on children hospitalized for asthma exacerbation in a state-wide longitudinal cohort, the Ohio Pediatric Asthma Repository. We examined BMI continuously (z scores) and categorically, comparing overweight and obese (Ov/Ob) to non-overweight and non-obese (nOv/nOb) children. We used linear mixed models controlling for site effects to determine if BMI was related to length of stay, as determined by physiologic readiness for discharge (PRD), defined as time to albuterol spaced every 4 h, need for nonstandard care or clinical history.Results: Across six hospitals, 874 children were included in analyses. BMI was positively associated with PRD (p=.008) but this increase was unlikely to be clinically significant. Ov/Ob children were more likely than nOv/nOb to require nonstandard care with repeat magnesium dosing in intensive care after dosing in the emergency department (OR = 3.23, 95%CI 1.39-7.78). Hospitalization in the year prior to enrollment was positively associated with BMI percentile (73.3 vs. 66.0, p=.028). Sleep disordered breathing was also associated with higher BMI percentile (78.2 vs. 65.9; p=.0013).Conclusions: Ov/Ob children had similar PRD to nOv/nOb children and were prone to repeat magnesium dosing. Previous hospitalization for exacerbation was positively associated with increasing BMI percentile. Additional research should investigate differential magnesium use by weight status, quantifying risks and benefits.

Identification of two early life eczema and non-eczema phenotypes with high risk for asthma development.

BACKGROUND: The "atopic march" has been considered a linear progression starting with eczema and culminating with development of asthma. Not all asthma cases, however, are preceded by eczema, and not all children with eczema go on to develop asthma. OBJECTIVE: The aim of this study was to explore the impact of allergic sensitization patterns on the association between early eczema and later childhood asthma. Given the numerous reported associations of the ciliary gene KIF3A with the atopic march, we also examined the impact of KIF3A risk allele rs12186803 on our analyses. METHODS: We studied 505 participants in the Cincinnati Childhood Allergy and Air Pollution Study (CCAAPS), a prospective birth cohort, with longitudinal eczema and asthma outcomes as well as prospective data regarding timing of sensitization to foods and aeroallergens. KIF3A genotypes were available on all children. RESULTS: Two high-risk groups were identified: one with and one without early eczema. The high-risk group with early eczema was more likely to be sensitized to food allergens, while the group without early eczema was more likely to be polysensitized to aeroallergens. The KIF3A rs12186803 risk allele interacted with food sensitization to increase asthma risk in children with eczema (P = 0.02). In children without eczema, asthma was associated with the interaction between rs12186803 and aeroallergen sensitization (P = 0.007). CONCLUSIONS & CLINICAL RELEVANCE: KIF3A interacted differentially with sensitization pattern to increase the risk of asthma in two high-risk groups of children with and without early eczema. Given the reported role of KIF3A in epithelial cell functioning, the results add evidence to the hypothesis that an impaired epithelial barrier is a key aspect in the development of allergic disease.

Vitamin D supplementation attenuates asthma development following traffic-related particulate matter exposure.

BACKGROUND: Recent literature suggests that children who are vitamin D deficient are uniquely susceptible to the effects of traffic-related air pollution (TRAP) exposure. This is highly significant because large segments of the population reside in zones of high TRAP exposure. OBJECTIVE: We sought to determine whether vitamin D supplementation mitigates the effect of TRAP exposure on asthma development, asthma exacerbation, and/or airway inflammation and to determine the timing of vitamin D supplementation that confers maximal health benefit. METHODS: Using established mouse models of asthma, we examined the effect of prenatal and postnatal vitamin D supplementation on asthma development, as well as the utility of vitamin D as a treatment for established asthma in the context of diesel exhaust particle (DEP) exposure. RESULTS: DEP and allergen coexposure resulted in increased airway hyperresponsiveness (AHR) and accumulation of pathogenic TH2/TH17 cells in the lungs of vitamin D-deficient mice compared with control mice. Prenatal and postnatal vitamin D supplementation significantly attenuated the development of AHR and decreased pulmonary accumulation of TH2/TH17 cells after coexposure to TRAP and allergen but not to allergen alone. Restoration of normal vitamin D status had no effect on AHR once asthma was already established. CONCLUSIONS: Our data establish that vitamin D confers protection against asthma development specifically in the context of TRAP exposure. Although vitamin D replacement did not reverse established asthma, restoration of normal vitamin D status in early life significantly attenuated the development of AHR in the setting of DEP-exacerbated allergic asthma and reduced numbers of lung TH2/TH17 cells, which portend the development of severe asthma.

A Pediatric Asthma Risk Score to better predict asthma development in young children.

BACKGROUND: Asthma phenotypes are currently not amenable to primary prevention or early intervention because their natural history cannot be reliably predicted. Clinicians remain reliant on poorly predictive asthma outcome tools because of a lack of better alternatives. OBJECTIVE: We sought to develop a quantitative personalized tool to predict asthma development in young children. METHODS: Data from the Cincinnati Childhood Allergy and Air Pollution Study (n = 762) birth cohort were used to identify factors that predicted asthma development. The Pediatric Asthma Risk Score (PARS) was constructed by integrating demographic and clinical data. The sensitivity and specificity of PARS were compared with those of the Asthma Predictive Index (API) and replicated in the Isle of Wight birth cohort. RESULTS: PARS reliably predicted asthma development in the Cincinnati Childhood Allergy and Air Pollution Study (sensitivity = 0.68, specificity = 0.77). Although both the PARS and API predicted asthma in high-risk children, the PARS had improved ability to predict asthma in children with mild-to-moderate asthma risk. In addition to parental asthma, eczema, and wheezing apart from colds, variables that predicted asthma in the PARS included early wheezing (odds ratio [OR], 2.88; 95% CI, 1.52-5.37), sensitization to 2 or more food allergens and/or aeroallergens (OR, 2.44; 95% CI, 1.49-4.05), and African American race (OR, 2.04; 95% CI, 1.19-3.47). The PARS was replicated in the Isle of Wight birth cohort (sensitivity = 0.67, specificity = 0.79), demonstrating that it is a robust, valid, and generalizable asthma predictive tool. CONCLUSIONS: The PARS performed better than the API in children with mild-to-moderate asthma. This is significant because these children are the most common and most difficult to predict and might be the most amenable to prevention strategies.

The antiprotease SPINK7 serves as an inhibitory checkpoint for esophageal epithelial inflammatory responses.

Loss of barrier integrity has an important role in eliciting type 2 immune responses, yet the molecular events that initiate and connect this with allergic inflammation remain unclear. We reveal an endogenous, homeostatic mechanism that controls barrier function and inflammatory responses in esophageal allergic inflammation. We show that a serine protease inhibitor, SPINK7 (serine peptidase inhibitor, kazal type 7), is part of the differentiation program of human esophageal epithelium and that SPINK7 depletion occurs in a human allergic, esophageal condition termed eosinophilic esophagitis. Experimental manipulation strategies reducing SPINK7 in an esophageal epithelial progenitor cell line and primary esophageal epithelial cells were sufficient to induce barrier dysfunction and transcriptional changes characterized by loss of cellular differentiation and altered gene expression known to stimulate allergic responses (for example, FLG and SPINK5). Epithelial silencing of SPINK7 promoted production of proinflammatory cytokines including thymic stromal lymphopoietin (TSLP). Loss of SPINK7 increased the activity of urokinase plasminogen-type activator (uPA), which in turn had the capacity to promote uPA receptor-dependent eosinophil activation. Treatment of epithelial cells with the broad-spectrum antiserine protease, α1 antitrypsin, reversed the pathologic features associated with SPINK7 silencing. The relevance of this pathway in vivo was supported by finding genetic epistasis between variants in TSLP and the uPA-encoding gene, PLAU We propose that the endogenous balance between SPINK7 and its target proteases is a key checkpoint in regulating mucosal differentiation, barrier function, and inflammatory responses and that protein replacement with antiproteases may be therapeutic for select allergic diseases.

Ohio Pediatric Asthma Repository: Opportunities to Revise Care Practices to Decrease Time to Physiologic Readiness for Discharge.

BACKGROUND: Large-scale, multisite studies in which researchers evaluate patient- and systems-level factors associated with pediatric asthma exacerbation outcomes are lacking. We sought to investigate patient-level risks and system-level practices related to physiologic readiness for discharge (PRD) in the prospective Ohio Pediatric Asthma Repository. METHODS: Participants were children ages 2 to 17 years admitted to an Ohio Pediatric Asthma Repository hospital for asthma exacerbation. Demographics, disease characteristics, and individual hospital practices were collected. The primary outcome was PRD timing (hours from admission or emergency department [ED] presentation until the first 4-hour albuterol spacing). RESULTS: Data for 1005 participants were available (865 ED presentations). Several nonstandard care practices were associated with time to PRD (P < .001). Continuous pulse oximetry was associated with increased time to PRD (P = .004). ED dexamethasone administration was associated with decreased time to PRD (P < .001) and less ICU admittance and intravenous steroid use (P < .0001). Earlier receipt of chest radiograph, antibiotics, and intravenous steroids was associated with shorter time to PRD (P < .05). Care practices associated with shorter time to PRD varied markedly by hospital. CONCLUSIONS: Substantial variation in care practices for inpatient asthma treatment exists among children's hospital systems in Ohio. We found several modifiable, system-level factors and therapies that contribute to PRD that warrant further investigation to identify the best and safest care practices. We also found that there was no standardized measure of exacerbation severity used across the hospitals. The development of such a tool is a critical gap in current practice and is needed to enable definitive comparative effectiveness studies of the management of acute asthma exacerbation.

Nasal DNA methylation is associated with childhood asthma.

AIM: We aim to study DNA methylation (DNAm) variations associated with childhood asthma. METHODS: Nasal DNAm was compared between sibling pairs discordant for asthma, 29 sib pairs for genome-wide association studies and 54 sib pairs for verification by pyrosequencing. Associations of methylation with asthma symptoms, allergy and environmental exposures were evaluated. In vitro experiments and functional genomic analyses were performed to explore biologic relevance. RESULTS: Three CpGs were associated with asthma. cg14830002 was associated with allergies in nonasthmatics. cg23602092 was associated with asthma symptoms. cg14830002 and cg23602092 were associated with traffic-related air pollution exposure. Nearby genes were transcriptionally regulated by diesel exhaust, house dust mite and 5-aza-2'-deoxycytidine. Active chromatin marks and transcription factor binding were found around these sites. CONCLUSION: We identified novel DNAm variations associated with childhood asthma and suggested new disease-contributing epigenetic mechanisms.

Adjunctive Pharmacotherapies in Children With Asthma Exacerbations Requiring Continuous Albuterol Therapy: Findings From The Ohio Pediatric Asthma Repository.

OBJECTIVES: To identify associations between use of ipratropium and/or intravenous magnesium and outcomes of children hospitalized with acute asthma exacerbations and treated with continuous albuterol. METHODS: Secondary analysis of data from children prospectively enrolled in the multicenter Ohio Pediatric Asthma Repository restricted to only children who were treated with continuous albuterol in their initial inpatient location. Children were treated with adjunctive therapies per the clinical team. RESULTS: Among 242 children who received continuous albuterol, 94 (39%) received ipratropium only, 13 (5%) received magnesium alone, 42 (17%) received both, and 93 (38%) received neither. The median duration of continuous albuterol was 7.0 (interquartile range [IQR]: 2.8-12.0) hours. Ipratropium use was associated with a shorter duration of continuous albuterol (4.9 [IQR: 2.0-10.0] hours) compared with dual therapy (11.0 [IQR: 5.6-28.6] hours; P = .001), but magnesium use was not (7.5 [IQR: 2.5-16.0] hours; P = .542). In Cox proportional models (adjusted for hospital, demographics, treatment location, and respiratory failure), magnesium was associated with longer durations of continuous albuterol (hazard ratio, 0.54 [95% confidence interval: 0.37-0.77]; P < .001) and hospitalization (hazard ratio, 0.41 [95% confidence interval: 0.28-0.60]; P < .001), but ipratropium was not. CONCLUSIONS: Ipratropium and magnesium were both often used in children with severe asthma hospitalizations that required continuous albuterol therapy. Magnesium use was associated with unfavorable outcomes, possibly reflecting preferential treatment to patients with more severe cases and differing practices between centers. Given the high prevalence of asthma, wide variations in practice, and the potential to improve outcomes and costs, prospective trials of these adjunctive therapies are needed.

Staphylococcal Biofilms in Atopic Dermatitis.

PURPOSE OF REVIEW: Atopic dermatitis (AD) is a chronic, relapsing inflammatory skin disorder that is a major public health burden worldwide. AD lesions are often colonized by Staphylococcus aureus and Staphylococcus epidermidis. An important aspect of Staphylococcus spp. is their propensity to form biofilms, adhesive surface-attached colonies that become highly resistant to antibiotics and immune responses, and recent studies have found that clinical isolates colonizing AD skin are often biofilm-positive. Biofilm formation results in complex bacterial communities that have unique effects on keratinocytes and host immunity. This review will summarize recent studies exploring the role of staphyloccocal biofilms in atopic dermatitis and the implications for treatment. RECENT FINDINGS: Recent studies suggest an important role for biofilms in the pathogenesis of numerous dermatologic diseases including AD. S. aureus biofilms have been found to colonize the eccrine ducts of AD skin, and these biofilms influence secretion of keratinocyte cytokines and trigger differentiation and apoptosis of keratinocytes. These activities may act to disrupt barrier function and promote disease pathogenesis as well as allergen sensitization. Formation of biofilm is a successful strategy that protects the bacteria from environmental danger, antibiotics, and phagocytosis, enabling chronic persistence in the host. An increasing number of S. aureus skin isolates are resistant to conventional antibiotics, and staphylococcal biofilm communities are prevalent on the skin of individuals with AD. Staphylococcal colonization of the skin impacts skin barrier function and plays multiple important roles in AD pathogenesis.

Nasal DNA methylation differentiates corticosteroid treatment response in pediatric asthma: A pilot study.

BACKGROUND: Treatment response to systemic corticosteroid in asthmatic children is heterogeneous and may be mediated by epigenetic mechanism(s). We aim to identify DNA methylation (DNAm) changes responsive to steroid, and DNAm biomarkers that distinguish treatment response. MATERIALS AND METHODS: We followed 33 children (ages 5-18) presenting to the Emergency Department (ED) for asthma exacerbation. Based on whether they met discharge criteria in ≤24 hours, participants were grouped into good and poor responders to steroid treatment. Nasal samples were collected upon presentation to the ED (T0) and 18-24 hours later (T1). Genome-wide DNAm was measured for both time points in 20 subjects, and compared between T0 and T1 in good and poor responders respectively. DNAm at T1 was also compared between two responder groups. DNAm of selected CpGs was verified in the complete cohort, and expression of associated genes was examined. Interactions between DNAm, common single nucleotide polymorphism (SNP) located at the CpG sites and treatment responses were assessed. RESULTS: Three CpGs located in the OTX2 promoter showed responder-specific DNAm changes from T0 to T1, in which DNAm decreased in good but not in poor responders. Good and poor responders showed differential DNAm at T1 in 127 CpGs without and 182 CpGs with common SNP co-localization. Negative correlations between DNAm and gene expression were observed at CpGs located within the LDHC promoter, suggesting an impact of DNAm on gene regulation. Interactions between SNPs, DNAm and treatment response were detected. CONCLUSION: Acute systemic steroid treatment modifies nasal DNAm in good responders. Nasal DNAm, dependent or independent of SNPs, can differentiate response to treatment in acute asthmatic children.