Longitudinal transcriptional immune profiles and persistent wheezing in moderate-to-late preterm infants.

BACKGROUND: Prematurity is associated with an increased risk of persistent wheezing but the underlying mechanisms are not well defined. The aim of this study was to identify blood transcriptional profiles associated with the development of wheezing in a cohort of moderate to late preterm infants and to define immune gene expression changes associated with wheezing. MATERIALS AND METHODS: A convenience sample of a multicenter birth cohort (SAREPREM) of moderate-late preterm children followed during the first 3 years of life was analyzed. Children were enrolled in the first 2 weeks of life (Y0) and longitudinally evaluated at 1 (Y1), 2 (Y2), and 3 years (Y3) of age, for the presence of wheezing and to obtain samples for transcriptional profile analysis. Samples were processed on Illumina HT12 chips and genomic expression analyses performed with R programming, modular analysis for biological function, and QuSAGE for quantitative gene expression. RESULTS: Seventy-six children were included in the study; 33 were classified as non-wheezing and 43 (56.6%) in the wheezing group. At Y0, children who developed wheezing had decreased expression of interferon genes and increased expression of B cell genes compared with the non-wheezing group. These changes in IFN and B cell gene expression were especially significant in children with late/persistent wheezing compared with transient wheezers. CONCLUSIONS: Changes in IFN and B lymphocyte gene expression identified in early life suggest the existence of specific immunological mechanisms that play an important role in the development of wheezing in late-preterm infants.

Human milk oligosaccharides are associated with maternal genetics and respiratory health of human milk-fed children.

Breastfeeding provides many health benefits, but its impact on respiratory health remains unclear. This study addresses the complex and dynamic nature of the mother-milk-infant triad by investigating maternal genomic factors regulating human milk oligosaccharides (HMOs), and their associations with respiratory health among human milk-fed infants. Nineteen HMOs are quantified from 980 mothers of the CHILD Cohort Study. Genome-wide association studies identify HMO-associated loci on chromosome 19p13.3 and 19q13.33 (lowest P = 2.4e-118), spanning several fucosyltransferase (FUT) genes. We identify novel associations on chromosome 3q27.3 for 6'-sialyllactose (P = 2.2e-9) in the sialyltransferase (ST6GAL1) gene. These, plus additional associations on chromosomes 7q21.32, 7q31.32 and 13q33.3, are replicated in the independent INSPIRE Cohort. Moreover, gene-environment interaction analyses suggest that fucosylated HMOs may modulate overall risk of recurrent wheeze among preschoolers with variable genetic risk scores (P < 0.01). Thus, we report novel genetic factors associated with HMOs, some of which may protect the respiratory health of children.

Prevalence of Alpha-1 Antitrypsin Deficiency Alleles in a Lithuanian Cohort of Wheezing Small Children.

Severe inherited alpha-1 antitrypsin deficiency (AATD) is an autosomal genetic condition linked to chronic obstructive pulmonary disease (COPD). The significance of heterozygous, milder deficiency variants (PiSZ, PiMZ, PiMS) is less clear. We studied AATD genotypes in 145 children (up to 72 months old) with assessed wheezing severity using the Pediatric Respiratory Assessment Measure (BCCH PRAM score). A control group of 74 children without airway obstruction was included. AAT concentration and Pi phenotype were determined from dry blood spot samples using nephelometry and real-time PCR; PiS and PiZ alleles were identified by isoelectrofocusing. Among the wheezers, the Pi*S allele incidence was 2.07% (3 cases) and the Pi*Z allele was 6.9% (10 cases). The Pi*Z allele frequency was higher in wheezers compared to controls (44.8% vs. 20.27%) and the general Lithuanian population (44.8% vs. 13.6%) and was similar to adult COPD patients in Lithuania: Pi*S 10.3% vs. 15.8% and Pi*Z 44.8% vs. 46.1%. No association was found between AAT genotypes and wheezing severity. Finding that wheezer children exhibit a frequency of Z* and S* alleles like that found in adults with COPD suggests a potential genetic predisposition that links early wheezing in children to the development of COPD in adulthood. Larger cohort studies are needed to confirm this finding.

Genetic predisposition to high BMI increases risk of early life respiratory infections and episodes of severe wheeze and asthma.

BACKGROUND: High body mass index (BMI) is an established risk factor for asthma, but the underlying mechanisms remain unclear. OBJECTIVE: To increase understanding of the BMI-asthma relationship by studying the association between genetic predisposition to higher BMI and asthma, infections and other asthma traits during childhood. METHODS: Data were obtained from the two ongoing Copenhagen Prospective Studies on Asthma in Childhood (COPSAC) mother-child cohorts. Polygenic risk scores for adult BMI were calculated for each child. Replication was done in the large-scale register-based Integrative Psychiatric Research (iPSYCH) cohort using data on hospitalisation for asthma and infections. RESULTS: In the COPSAC cohorts (n=974), the adult BMI polygenic risk score was significantly associated with lower respiratory tract infections (incidence rate ratio (IRR) 1.20, 95% CI 1.08-1.33, false discovery rate p-value (pFDR)=0.005) at age 0-3 years and episodes of severe wheeze (IRR 1.30, 95% CI 1.06-1.60, pFDR=0.04) at age 0-6 years. Lower respiratory tract infections partly mediated the association between the adult BMI polygenic risk score and severe wheeze (proportion mediated: 0.59, 95% CI 0.28-2.24, p-value associated with the average causal mediation effect (pACME)=2e-16). In contrast, these associations were not mediated through the child's current BMI and the polygenic risk score was not associated with an asthma diagnosis or reduced lung function up to age 18 years. The associations were replicated in iPSYCH (n=114 283), where the adult BMI polygenic risk score significantly increased the risk of hospitalisations for lower respiratory tract infections and wheeze or asthma throughout childhood to age 18 years. CONCLUSION: Children with genetic predisposition to higher BMI had increased risk of lower respiratory tract infections and severe wheeze, independent of the child's current BMI. These results shed further light on the complex relationship between body mass BMI and asthma.

ST2 and IL-33 polymorphisms and the development of childhood asthma: a prospective birth cohort study in Finnish children.

The ST2/IL-33 signaling pathway has an important role in the host inflammatory response. Here we aimed to study the association of ST2 and IL-33 polymorphisms with serum soluble (s) ST2 and IL-33 concentrations in healthy Finnish children and, in addition, their association with childhood asthma. In total, 146 children were followed from birth to the age 7 years for the development of asthma. Single-nucleotide polymorphisms (SNPs) in ST2 and IL-33 were determined, and associations of the SNP variants with serum levels of sST2 and IL-33 at age of 13 months and with recurrent wheezing and childhood asthma at 7 years of age were analyzed. Children with ST2 rs1041973 AC/AA genotypes had significantly lower level of serum sST2 (2453 pg/mL; IQR 2265) than those with CC genotype (5437 pg/mL; IQR 2575; p = < 0.0001). Similar difference was also observed with ST2 rs13408661. No differences were observed between subjects with studied IL-33 SNPs. Children who carried genetic variants of ST2 rs1041973 or rs13408661 seemed to have a higher risk of asthma. In contrast, children who carried genetic variants of IL-33 rs12551268 were less often diagnosed with asthma. Even though these SNPs seemed to associate with asthma, the differences were not statistically significant.

Association between polymorphisms on chromosome 17q12-q21 and rhinovirus-induced interferon responses.

BACKGROUND: Single nucleotide polymorphisms (SNPs) in genes on chromosome 17q12-q21 are associated with childhood-onset asthma and rhinovirus-induced wheeze. There are few mechanistic data linking chromosome 17q12-q21 to wheezing illness. OBJECTIVE: We investigated whether 17q12-q21 risk alleles were associated with impaired interferon responses to rhinovirus. METHODS: In a population-based birth cohort of European ancestry, we stimulated peripheral blood mononuclear cells with rhinovirus A1 (RV-A1) and rhinovirus A16 (RV-A16) and measured IFN and IFN-induced C-X-C motif chemokine ligand 10 (aka IP10) responses in supernatants. We investigated associations between virus-induced cytokines and 6 SNPs in 17q12-q21. Bayesian profile regression was applied to identify clusters of individuals with different immune response profiles and genetic variants. RESULTS: Five SNPs (in high linkage disequilibrium, r2 ≥ 0.8) were significantly associated with RV-A1-induced IFN-ß (rs9303277, P = .010; rs11557467, P = .012; rs2290400, P = .006; rs7216389, P = .008; rs8079416, P = .005). A reduction in RV-A1-induced IFN-ß was observed among individuals with asthma risk alleles. There were no significant associations for RV-A1-induced IFN-α or CXCL10, or for any RV-A16-induced IFN/CXCL10. Bayesian profile regression analysis identified 3 clusters that differed in IFN-ß induction to RV-A1 (low, medium, high). The typical genetic profile of the cluster associated with low RV-A1-induced IFN-ß responses was characterized by a very high probability of being homozygous for the asthma risk allele for all SNPs. Children with persistent wheeze were almost 3 times more likely to be in clusters with reduced/average RV-A1-induced IFN-ß responses than in the high immune response cluster. CONCLUSIONS: Polymorphisms on chromosome 17q12-q21 are associated with rhinovirus-induced IFN-ß, suggesting a novel mechanism-impaired IFN-ß induction-links 17q12-q21 risk alleles with asthma/wheeze.

Improvements in lung function following vitamin C supplementation to pregnant smokers are associated with buccal DNA methylation at 5 years of age.

BACKGROUND: We previously reported in the "Vitamin C to Decrease the Effects of Smoking in Pregnancy on Infant Lung Function" randomized clinical trial (RCT) that vitamin C (500 mg/day) supplementation to pregnant smokers is associated with improved respiratory outcomes that persist through 5 years of age. The objective of this study was to assess whether buccal cell DNA methylation (DNAm), as a surrogate for airway epithelium, is associated with vitamin C supplementation, improved lung function, and decreased occurrence of wheeze. METHODS: We conducted epigenome-wide association studies (EWAS) using Infinium MethylationEPIC arrays and buccal DNAm from 158 subjects (80 placebo; 78 vitamin C) with pulmonary function testing (PFT) performed at the 5-year visit. EWAS were performed on (1) vitamin C treatment, (2) forced expiratory flow between 25 and 75% of expired volume (FEF25-75), and (3) offspring wheeze. Models were adjusted for sex, race, study site, gestational age at randomization (≤ OR > 18 weeks), proportion of epithelial cells, and latent covariates in addition to child length at PFT in EWAS for FEF25-75. We considered FDR p < 0.05 as genome-wide significant and nominal p < 0.001 as candidates for downstream analyses. Buccal DNAm measured in a subset of subjects at birth and near 1 year of age was used to determine whether DNAm signatures originated in utero, or emerged with age. RESULTS: Vitamin C treatment was associated with 457 FDR significant (q < 0.05) differentially methylated CpGs (DMCs; 236 hypermethylated; 221 hypomethylated) and 53 differentially methylated regions (DMRs; 26 hyper; 27 hypo) at 5 years of age. FEF25-75 was associated with one FDR significant DMC (cg05814800), 1,468 candidate DMCs (p < 0.001), and 44 DMRs. Current wheeze was associated with 0 FDR-DMCs, 782 candidate DMCs, and 19 DMRs (p < 0.001). In 365/457 vitamin C FDR significant DMCs at 5 years of age, there was no significant interaction between time and treatment. CONCLUSIONS: Vitamin C supplementation to pregnant smokers is associated with buccal DNA methylation in offspring at 5 years of age, and most methylation signatures appear to be persistent from the prenatal period. Buccal methylation at 5 years was also associated with current lung function and occurrence of wheeze, and these functionally associated loci are enriched for vitamin C associated loci. Clinical trial registration ClinicalTrials.gov, NCT01723696 and NCT03203603.

Genetics of preschool wheeze and its progression to childhood asthma.

Wheezing is a common and heterogeneous condition in preschool children. In some countries, the prevalence can be as high as 30% and up to 50% of all children experience wheezing before the age of 6. Asthma often starts with preschool wheeze, but not all wheezing children will develop asthma at school age. At this moment, it is not possible to accurately predict which wheezing children will develop asthma. Recently, studying the genetics of wheeze and the childhood-onset of asthma have grown in interest. Childhood-onset asthma has a stronger heritability in comparison with adult-onset asthma. In early childhood asthma exacerbations, CDHR3, which encodes the receptor for Rhinovirus C, was identified, as well as IL33, and the 17q locus that includes GSDMB and ORMDL3 genes. The 17q locus is the strongest wheeze and childhood-onset asthma locus, and was shown to interact with many environmental factors, including smoking and infections. Finally, ANXA1 was recently associated with early-onset, persistent wheeze. ANXA1 may help resolve eosinophilic inflammation. Overall, despite its complexities, genetic approaches to unravel the early-onset of wheeze and asthma are promising, since these shed more light on mechanisms of childhood asthma-onset. Implicated genes point toward airway epithelium and its response to external factors, such as viral infections. However, the heterogeneity of wheeze phenotypes complicates genetic studies. It is therefore important to define accurate wheezing phenotypes and forge larger international collaborations to gain a better understanding of the pathways underlying early-onset asthma.

Oropharyngeal Microbiota Clusters in Children with Asthma or Wheeze Associate with Allergy, Blood Transcriptomic Immune Pathways, and Exacerbation Risk.

Rationale: Children with preschool wheezing or school-age asthma are reported to have airway microbial imbalances. Objectives: To identify clusters in children with asthma or wheezing using oropharyngeal microbiota profiles. Methods: Oropharyngeal swabs from the U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes) pediatric asthma or wheezing cohort were characterized using 16S ribosomal RNA gene sequencing, and unsupervised hierarchical clustering was performed on the Bray-Curtis ß-diversity. Enrichment scores of the Molecular Signatures Database hallmark gene sets were computed from the blood transcriptome using gene set variation analysis. Children with severe asthma or severe wheezing were followed up for 12-18 months, with assessment of the frequency of exacerbations. Measurements and Main Results: Oropharyngeal samples from 241 children (age range, 1-17 years; 40% female) revealed four taxa-driven clusters dominated by Streptococcus, Veillonella, Rothia, and Haemophilus. The clusters showed significant differences in atopic dermatitis, grass pollen sensitization, FEV1% predicted after salbutamol, and annual asthma exacerbation frequency during follow-up. The Veillonella cluster was the most allergic and included the highest percentage of children with two or more exacerbations per year during follow-up. The oropharyngeal clusters were different in the enrichment scores of TGF-ß (transforming growth factor-ß) (highest in the Veillonella cluster) and Wnt/ß-catenin signaling (highest in the Haemophilus cluster) transcriptomic pathways in blood (all q values <0.05). Conclusions: Analysis of the oropharyngeal microbiota of children with asthma or wheezing identified four clusters with distinct clinical characteristics (phenotypes) that associate with risk for exacerbation and transcriptomic pathways involved in airway remodeling. This suggests that further exploration of the oropharyngeal microbiota may lead to novel pathophysiologic insights and potentially new treatment approaches.

A meta-analysis of genome-wide association studies of childhood wheezing phenotypes identifies ANXA1 as a susceptibility locus for persistent wheezing.

Background: Many genes associated with asthma explain only a fraction of its heritability. Most genome-wide association studies (GWASs) used a broad definition of 'doctor-diagnosed asthma', thereby diluting genetic signals by not considering asthma heterogeneity. The objective of our study was to identify genetic associates of childhood wheezing phenotypes. Methods: We conducted a novel multivariate GWAS meta-analysis of wheezing phenotypes jointly derived using unbiased analysis of data collected from birth to 18 years in 9568 individuals from five UK birth cohorts. Results: Forty-four independent SNPs were associated with early-onset persistent, 25 with pre-school remitting, 33 with mid-childhood remitting, and 32 with late-onset wheeze. We identified a novel locus on chr9q21.13 (close to annexin 1 [ANXA1], p<6.7 × 10-9), associated exclusively with early-onset persistent wheeze. We identified rs75260654 as the most likely causative single nucleotide polymorphism (SNP) using Promoter Capture Hi-C loops, and then showed that the risk allele (T) confers a reduction in ANXA1 expression. Finally, in a murine model of house dust mite (HDM)-induced allergic airway disease, we demonstrated that anxa1 protein expression increased and anxa1 mRNA was significantly induced in lung tissue following HDM exposure. Using anxa1-/- deficient mice, we showed that loss of anxa1 results in heightened airway hyperreactivity and Th2 inflammation upon allergen challenge. Conclusions: Targeting this pathway in persistent disease may represent an exciting therapeutic prospect. Funding: UK Medical Research Council Programme Grant MR/S025340/1 and the Wellcome Trust Strategic Award (108818/15/Z) provided most of the funding for this study.

Three-quarters of children hospitalized for wheezing or asthma symptoms are preschool-aged. Some will continue to experience breathing difficulties through childhood and adulthood. Others will undergo a complete resolution of their symptoms by the time they reach elementary school. The varied trajectories of young children with wheezing suggest that it is not a single disease. There are likely different genetic or environmental causes. Despite these differences, wheezing treatments for young children are 'one size fits all.' Studying the genetic underpinnings of wheezing may lead to more customized treatment options. Granell et al. studied the genetic architecture of different patterns of wheezing from infancy to adolescence. To do so, they used machine learning technology to analyze the genomes of 9,568 individuals, who participated in five studies in the United Kingdom from birth to age 18. The experiments found a new genetic variation in the ANXA1 gene linked with persistent wheezing starting in early childhood. By comparing mice with and without this gene, Granell et al. showed that the protein encoded by ANXA1 controls inflammation in the lungs in response to allergens. Animals lacking the protein develop worse lung inflammation after exposure to dust mite allergens. Identifying a new gene linked to a specific subtype of wheezing might help scientists develop better strategies to diagnose, treat, and prevent asthma. More studies are needed on the role of the protein encoded by ANXA1 in reducing allergen-triggered lung inflammation to determine if this protein or therapies that boost its production may offer relief for chronic lung inflammation.

Association of Asthma Risk Alleles With Acute Respiratory Tract Infections and Wheezing Illnesses in Young Children.

BACKGROUND: Genome-wide association studies have identified several risk alleles for early childhood asthma, particularly in the 17q21 locus and in the cadherin-related family member 3 (CDHR3) gene. Contribution of these alleles to the risk of acute respiratory tract infections (ARI) in early childhood is unclear. METHODS: We analyzed data from the STEPS birth-cohort study of unselected children and the VINKU and VINKU2 studies on children with severe wheezing illness. Genome-wide genotyping was performed on 1011 children. We analyzed the association between 11 preselected asthma risk alleles and the risk of ARIs and wheezing illnesses of various viral etiologies. RESULTS: The asthma risk alleles in CDHR3, GSDMA, and GSDMB were associated with an increased rate of ARIs (for CDHR3, incidence rate ratio [IRR], 1.06; 95% confidence interval [CI], 1.01-1.12; P = .02), and risk allele in CDHR3 gene with rhinovirus infections (IRR, 1.10; 95% CI, 1.01-1.20, P = .03). Asthma risk alleles in GSDMA, GSDMB, IKZF3, ZPBP2, and ORMDL3 genes were associated with wheezing illnesses in early childhood, especially rhinovirus-positive wheezing illnesses. CONCLUSIONS: Asthma risk alleles were associated with an increased rate of ARIs and an increased risk of viral wheezing illnesses. Nonwheezing and wheezing ARIs and asthma may have shared genetic risk factors. Clinical Trials Registration. NCT00494624 and NCT00731575.

Network analyses of upper and lower airway transcriptomes identify shared mechanisms among children with recurrent wheezing and school-age asthma.

Background: Predicting which preschool children with recurrent wheezing (RW) will develop school-age asthma (SA) is difficult, highlighting the critical need to clarify the pathogenesis of RW and the mechanistic relationship between RW and SA. Despite shared environmental exposures and genetic determinants, RW and SA are usually studied in isolation. Based on network analysis of nasal and tracheal transcriptomes, we aimed to identify convergent transcriptomic mechanisms in RW and SA. Methods: RNA-sequencing data from nasal and tracheal brushing samples were acquired from the Gene Expression Omnibus. Combined with single-cell transcriptome data, cell deconvolution was used to infer the composition of 18 cellular components within the airway. Consensus weighted gene co-expression network analysis was performed to identify consensus modules closely related to both RW and SA. Shared pathways underlying consensus modules between RW and SA were explored by enrichment analysis. Hub genes between RW and SA were identified using machine learning strategies and validated using external datasets and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Finally, the potential value of hub genes in defining RW subsets was determined using nasal and tracheal transcriptome data. Results: Co-expression network analysis revealed similarities in the transcriptional networks of RW and SA in the upper and lower airways. Cell deconvolution analysis revealed an increase in mast cell fraction but decrease in club cell fraction in both RW and SA airways compared to controls. Consensus network analysis identified two consensus modules highly associated with both RW and SA. Enrichment analysis of the two consensus modules indicated that fatty acid metabolism-related pathways were shared key signals between RW and SA. Furthermore, machine learning strategies identified five hub genes, i.e., CST1, CST2, CST4, POSTN, and NRTK2, with the up-regulated hub genes in RW and SA validated using three independent external datasets and qRT-PCR. The gene signatures of the five hub genes could potentially be used to determine type 2 (T2)-high and T2-low subsets in preschoolers with RW. Conclusions: These findings improve our understanding of the molecular pathogenesis of RW and provide a rationale for future exploration of the mechanistic relationship between RW and SA.

Early childhood wheezing phenotypes and determinants in a South African birth cohort: longitudinal analysis of the Drakenstein Child Health Study.

BACKGROUND: Developmental trajectories of childhood wheezing in low-income and middle-income countries (LMICs) have not been well described. We aimed to derive longitudinal wheeze phenotypes from birth to 5 years in a South African birth cohort and compare those with phenotypes derived from a UK cohort. METHODS: We used data from the Drakenstein Child Health Study (DCHS), a longitudinal birth cohort study in a peri-urban area outside Cape Town, South Africa. Pregnant women (aged ≥18 years) were enrolled during their second trimester at two public health clinics. We followed up children from birth to 5 years to derive six multidimensional indicators of wheezing (including duration, temporal sequencing, persistence, and recurrence) and applied Partition Around Medoids clustering to derive wheeze phenotypes. We compared phenotypes with a UK cohort (the Avon Longitudinal Study of Parents and Children [ALSPAC]). We investigated associations of phenotypes with early-life exposures, including all-cause lower respiratory tract infection (LRTI) and virus-specific LRTI (respiratory syncytial virus, rhinovirus, adenovirus, influenza, and parainfluenza virus) up to age 5 years. We investigated the association of phenotypes with lung function at 6 weeks and 5 years. FINDINGS: Between March 5, 2012, and March 31, 2015, we enrolled 1137 mothers and there were 1143 livebirths. Four wheeze phenotypes were identified among 950 children with complete data: never (480 children [50%]), early transient (215 children [23%]), late onset (104 children [11%]), and recurrent (151 children [16%]). Multivariate adjusted analysis indicated that LRTI and respiratory syncytial virus-LRTI, but not other respiratory viruses, were associated with increased risk of recurrent wheeze (odds ratio [OR] 2·79 [95% CI 2·05-3·81] for all LTRIs; OR 2·59 [1·30-5·15] for respiratory syncytial virus-LRTIs). Maternal smoking (1·88 [1·12-3·02]), higher socioeconomic status (2·46 [1·23-4·91]), intimate partner violence (2·01 [1·23-3·29]), and male sex (2·47 [1·50-4·04]) were also associated with recurrent wheeze. LRTI and respiratory syncytial virus-LRTI were also associated with early transient and late onset clusters. Wheezing illness architecture differed between DCHS and ALSPAC; children included in ALSPAC in the early transient cluster wheezed for a longer period before remission and late-onset wheezing started at an older age, and no persistent phenotype was identified in DCHS. At 5 years, airway resistance was higher in children with early or recurrent wheeze compared with children who had never wheezed. Airway resistance increased from 6 weeks to 5 years among children with recurrent wheeze. INTERPRETATION: Effective strategies to reduce maternal smoking and psychosocial stressors and new preventive interventions for respiratory syncytial virus are urgently needed to optimise child health in LMICs. FUNDING: UK Medical Research Council; The Bill & Melinda Gates Foundation; National Institutes of Health Human Heredity and Health in Africa; South African Medical Research Council; Wellcome Trust.

Dog ownership in infancy is protective for persistent wheeze in 17q21 asthma-risk carriers.

BACKGROUND: Asthma-associated single nucleotide polymorphisms from large genome-wide association studies only explain a fraction of genetic heritability. Likely causes of the missing heritability include broad phenotype definitions and gene-environment interactions (GxE). The mechanisms underlying GxE in asthma are poorly understood. Previous GxE studies on pet ownership showed discordant results. OBJECTIVES: We sought to study the GxE between the 17q12-21 locus and pet ownership in infancy in relation to wheeze. METHODS: Wheezing classes derived from 5 UK-based birth cohorts (latent class analysis) were used to study GxE between the 17q12-21 asthma-risk variant rs2305480 and dog and cat ownership in infancy, using multinomial logistic regression. A total of 9149 children had both pet ownership and genotype data available. Summary statistics from individual analyses were meta-analyzed. RESULTS: rs2305480 G allele was associated with increased risk of persistent wheeze (additive model odds ratio, 1.37; 95% CI, 1.25-1.51). There was no evidence of an association between dog or cat ownership and wheeze. We found significant evidence of a GxE interaction between rs2305480 and dog ownership (P = 8.3 × 10-4) on persistent wheeze; among dog owners, the G allele was no longer associated with an increased risk of persistent wheeze (additive model odds ratio, 0.95; 95% CI, 0.73-1.24). For those without pets, G allele was associated with increased risk of persistent wheeze (odds ratio, 1.61; 95% CI, 1.40-1.86). Among cat owners, no such dampening of the genetic effect was observed. CONCLUSIONS: Among dog owners, rs2305480 G was no longer associated with an increased risk of persistent wheeze (or asthma). Early-life environmental exposures may therefore attenuate likelihood of asthma in those carrying 17q12-21 risk alleles.

Evolution of Eczema, Wheeze, and Rhinitis from Infancy to Early Adulthood: Four Birth Cohort Studies.

Rationale: The relationship between eczema, wheeze or asthma, and rhinitis is complex, and epidemiology and mechanisms of their comorbidities is unclear. Objectives: To investigate within-individual patterns of morbidity of eczema, wheeze, and rhinitis from birth to adolescence/early adulthood. Methods: We investigated onset, progression, and resolution of eczema, wheeze, and rhinitis using descriptive statistics, sequence mining, and latent Markov modeling in four population-based birth cohorts. We used logistic regression to ascertain if early-life eczema or wheeze, or genetic factors (filaggrin [FLG] mutations and 17q21 variants), increase the risk of multimorbidity. Measurements and Main Results: Single conditions, although the most prevalent, were observed significantly less frequently than by chance. There was considerable variation in the timing of onset/remission/persistence/intermittence. Multimorbidity of eczema+wheeze+rhinitis was rare but significantly overrepresented (three to six times more often than by chance). Although infantile eczema was associated with subsequent multimorbidity, most children with eczema (75.4%) did not progress to any multimorbidity pattern. FLG mutations and rs7216389 were not associated with persistence of eczema/wheeze as single conditions, but both increased the risk of multimorbidity (FLG by 2- to 3-fold, rs7216389 risk variant by 1.4- to 1.7-fold). Latent Markov modeling revealed five latent states (no disease/low risk, mainly eczema, mainly wheeze, mainly rhinitis, multimorbidity). The most likely transition to multimorbidity was from eczema state (0.21). However, although this was one of the highest transition probabilities, only one-fifth of those with eczema transitioned to multimorbidity. Conclusions: Atopic diseases fit a multimorbidity framework, with no evidence for sequential atopic march progression. The highest transition to multimorbidity was from eczema, but most children with eczema (more than three-quarters) had no comorbidities.

Early-life predictors and risk factors of peanut allergy, and its association with asthma in later-life: Population-based birth cohort study.

BACKGROUND: Understanding risk factors for peanut allergy (PA) is essential to develop effective preventive measures. OBJECTIVE: The objective was to ascertain associates and predictors of PA, and the relationship between PA and asthma severity. METHODS: In a population-based birth cohort, we investigated the association between objectively confirmed PA with early-life environmental exposures, filaggrin (FLG)-loss-of-function mutations and other atopic disease. We then examined the association of PA with longitudinal trajectories of sensitization, wheeze and allergic comorbidities, which were previously derived using machine learning. Finally, we ascertained the relationship between PA and asthma severity. RESULTS: PA was confirmed in 30/959 participants with evaluable data. In the multivariate analysis, eczema in infancy (OR = 4.4, 95% CI 1.5-13.2, p = 0.007), egg sensitization at age 3 years (OR = 9.7, 95% CI 3.3-29.9, p < 0.001) and early-life cat ownership (OR = 3.0, 95% CI 1.1-8.4, p = 0.04) were independent associates of PA. In the stratified analysis among 700 participants with genetic information, in children with early-life eczema there was no difference in FLG mutations between children with and without PA (3/18 [16.7%] vs. 42/220 [19.1%], p = 1.00). In contrast, among children without eczema, those with PA were almost eight times more likely to have FLG mutations (2/6 [33.3%] vs. 27/456 [5.9%], p = 0.049). We observed associations between PA and multiple allergic sensitization profiles derived using machine learning, with ~60-fold increase in risk among individuals assigned to multiple early sensitization. PA was significantly associated with persistent wheeze (but not other wheeze phenotypes), and with trajectories of atopic disease characterized by co-morbid persistent eczema and wheeze (but not with transient phenotypes). Children with PA were more likely to have asthma, but among asthmatics we found no evidence of an association between PA and asthma severity. CONCLUSIONS: Peanut allergy is associated with multiple IgE sensitization and early-onset persistent eczema and wheeze. FLG loss-of-function mutations were associated with peanut allergy in children without eczema.

Modeling Wheezing Spells Identifies Phenotypes with Different Outcomes and Genetic Associates.

Rationale: Longitudinal modeling of current wheezing identified similar phenotypes, but their characteristics often differ between studies. Objectives: We propose that a more comprehensive description of wheeze may better describe trajectories than binary information on the presence/absence of wheezing. Methods: We derived six multidimensional variables of wheezing spells from birth to adolescence (including duration, temporal sequencing, and the extent of persistence/recurrence). We applied partition-around-medoids clustering on these variables to derive phenotypes in five birth cohorts. We investigated within- and between-phenotype differences compared with binary latent class analysis models and ascertained associations of these phenotypes with asthma and lung function and with polymorphisms in asthma loci 17q12-21 and CDHR3 (cadherin-related family member 3). Measurements and Main Results: Analysis among 7,719 participants with complete data identified five spell-based wheeze phenotypes with a high degree of certainty: never (54.1%), early-transient (ETW) (23.7%), late-onset (LOW) (6.9%), persistent (PEW) (8.3%), and a novel phenotype, intermittent wheeze (INT) (6.9%). FEV1/FVC was lower in PEW and INT compared with ETW and LOW and declined from age 8 years to adulthood in INT. 17q12-21 and CDHR3 polymorphisms were associated with higher odds of PEW and INT, but not ETW or LOW. Latent class analysis- and spell-based phenotypes appeared similar, but within-phenotype individual trajectories and phenotype allocation differed substantially. The spell-based approach was much more robust in dealing with missing data, and the derived clusters were more stable and internally homogeneous. Conclusions: Modeling of spell variables identified a novel intermittent wheeze phenotype associated with lung function decline to early adulthood. Using multidimensional spell variables may better capture wheeze development and provide a more robust input for phenotype derivation.

Immune Responsiveness to LPS Determines Risk of Childhood Wheeze and Asthma in 17q21 Risk Allele Carriers.

Rationale: In murine models, microbial exposures induce protection from experimental allergic asthma through innate immunity. Objectives: Our aim was to assess the association of early life innate immunity with the development of asthma in children at risk. Methods: In the PASTURE farm birth cohort, innate T-helper cell type 2 (Th2), Th1, and Th17 cytokine expression at age 1 year was measured after stimulation of peripheral blood mononuclear cells with LPS in n = 445 children. Children at risk of asthma were defined based on single-nucleotide polymorphisms at the 17q21 asthma gene locus. Specifically, we used the SNP rs7216389 in the GSDMB gene. Wheeze in the first year of life was assessed by weekly diaries and asthma by questionnaire at age 6 years. Measurements and Main Results: Not all cytokines were detectable in all children after LPS stimulation. When classifying detectability of cytokines by latent class analysis, carrying the 17q21 risk allele rs7216389 was associated with risk of wheeze only in the class with the lowest level of LPS-induced activation: odds ratio (OR), 1.89; 95% confidence interval [CI], 1.13-3.16; P = 0.015. In contrast, in children with high cytokine activation after LPS stimulation, no association of the 17q21 risk allele with wheeze (OR, 0.63; 95% CI, 0.29-1.40; P = 0.258, P = 0.034 for interaction) or school-age asthma was observed. In these children, consumption of unprocessed cow's milk was associated with higher cytokine activation (OR, 3.37; 95% CI, 1.56-7.30; P = 0.002), which was in part mediated by the gut microbiome. Conclusions: These findings suggest that within the 17q21 genotype, asthma risk can be mitigated by activated immune responses after innate stimulation, which is partly mediated by a gut-immune axis.

17q12-21 risk-variants influence cord blood immune regulation and multitrigger-wheeze.

BACKGROUND: Childhood wheeze represents a first symptom of asthma. Early identification of children at risk for wheeze related to 17q12-21 variants and their underlying immunological mechanisms remain unknown. We aimed to assess the influence of 17q12-21 variants and mRNA expression at birth on the development of wheeze. METHODS: Children were classified as multitrigger/viral/no wheeze until six years of age. The PAULINA/PAULCHEN birth cohorts were genotyped (n = 216; GSA-chip). mRNA expression of 17q21 and innate/adaptive genes was measured (qRT-PCR) in cord blood mononuclear cells. Expression quantitative trait loci (eQTL) and mediation analyses were performed. Genetic variation of 17q12-21 asthma-single nucleotide polymorphisms (SNPs) was summarized as the first principal component (PC1) and used to classify single SNP effects on gene expression as (locus)-dependent/independent eQTL SNPs. RESULTS: Core region risk variants (IKZF3, ZPBP2, GSDMB, ORMDL3) were associated with multitrigger wheeze (OR: 3.05-5.43) and were locus-dependent eQTL SNPs with higher GSDMA, TLR2, TLR5, and lower TGFB1 expression. Increased risk of multitrigger wheeze with rs9303277 was in part mediated by TLR2 expression. Risk variants distal to the core region were mainly locus-independent eQTL SNPs with decreased CD209, CD86, TRAF6, RORA, and IL-9 expression. Distinct immune signatures in cord blood were associated either with multitrigger wheeze (increased innate genes, e.g., TLR2, IPS1, LY75) or viral wheeze (decreased NF-κB genes, e.g., TNFAIP3 and TNIP2). CONCLUSION: Locus-dependent eQTL SNPs (core region) associated with increased inflammatory genes (primarily TLR2) at birth and subsequent multitrigger wheeze indicate that early priming and imbalance may be crucial for asthma pathophysiology. Locus-independent eQTL SNPs (mainly distal region, rs1007654) may be involved in the initiation of dendritic cell activation/maturation (TRAF6) and interaction with T cells (CD209, CD86). Identifying potential mechanistic pathways at birth may point to critical key points during early immune development predisposing to asthma.