An integrated molecular risk score early in life for subsequent childhood asthma risk.

BACKGROUND: Numerous children present with early wheeze symptoms, yet solely a subgroup develops childhood asthma. Early identification of children at risk is key for clinical monitoring, timely patient-tailored treatment, and preventing chronic, severe sequelae. For early prediction of childhood asthma, we aimed to define an integrated risk score combining established risk factors with genome-wide molecular markers at birth, complemented by subsequent clinical symptoms/diagnoses (wheezing, atopic dermatitis, food allergy). METHODS: Three longitudinal birth cohorts (PAULINA/PAULCHEN, n = 190 + 93 = 283, PASTURE, n = 1133) were used to predict childhood asthma (age 5-11) including epidemiological characteristics and molecular markers: genotype, DNA methylation and mRNA expression (RNASeq/NanoString). Apparent (ap) and optimism-corrected (oc) performance (AUC/R2) was assessed leveraging evidence from independent studies (Naïve-Bayes approach) combined with high-dimensional logistic regression models (LASSO). RESULTS: Asthma prediction with epidemiological characteristics at birth (maternal asthma, sex, farm environment) yielded an ocAUC = 0.65. Inclusion of molecular markers as predictors resulted in an improvement in apparent prediction performance, however, for optimism-corrected performance only a moderate increase was observed (upto ocAUC = 0.68). The greatest discriminate power was reached by adding the first symptoms/diagnosis (up to ocAUC = 0.76; increase of 0.08, p = .002). Longitudinal analysis of selected mRNA expression in PASTURE (cord blood, 1, 4.5, 6 years) showed that expression at age six had the strongest association with asthma and correlation of genes getting larger over time (r = .59, p < .001, 4.5-6 years). CONCLUSION: Applying epidemiological predictors alone showed moderate predictive abilities. Molecular markers from birth modestly improved prediction. Allergic symptoms/diagnoses enhanced the power of prediction, which is important for clinical practice and for the design of future studies with molecular markers.

NetCoMi: network construction and comparison for microbiome data in R.

MOTIVATION: Estimating microbial association networks from high-throughput sequencing data is a common exploratory data analysis approach aiming at understanding the complex interplay of microbial communities in their natural habitat. Statistical network estimation workflows comprise several analysis steps, including methods for zero handling, data normalization and computing microbial associations. Since microbial interactions are likely to change between conditions, e.g. between healthy individuals and patients, identifying network differences between groups is often an integral secondary analysis step. Thus far, however, no unifying computational tool is available that facilitates the whole analysis workflow of constructing, analysing and comparing microbial association networks from high-throughput sequencing data. RESULTS: Here, we introduce NetCoMi (Network Construction and comparison for Microbiome data), an R package that integrates existing methods for each analysis step in a single reproducible computational workflow. The package offers functionality for constructing and analysing single microbial association networks as well as quantifying network differences. This enables insights into whether single taxa, groups of taxa or the overall network structure change between groups. NetCoMi also contains functionality for constructing differential networks, thus allowing to assess whether single pairs of taxa are differentially associated between two groups. Furthermore, NetCoMi facilitates the construction and analysis of dissimilarity networks of microbiome samples, enabling a high-level graphical summary of the heterogeneity of an entire microbiome sample collection. We illustrate NetCoMi's wide applicability using data sets from the GABRIELA study to compare microbial associations in settled dust from children's rooms between samples from two study centers (Ulm and Munich). AVAILABILITY: R scripts used for producing the examples shown in this manuscript are provided as supplementary data. The NetCoMi package, together with a tutorial, is available at https://github.com/stefpeschel/NetCoMi. CONTACT: Tel:+49 89 3187 43258; stefanie.peschel@mail.de. SUPPLEMENTARY INFORMATION: Supplementary data are available at Briefings in Bioinformatics online.

Trajectories of cough without a cold in early childhood and associations with atopic diseases.

BACKGROUND: Although children can frequently experience a cough that affects their quality of life, few epidemiological studies have explored cough without a cold during childhood. OBJECTIVES: The objective of the study was to describe the latent class trajectories of cough from one to 10 years old and analyse their association with wheezing, atopy and allergic diseases. METHODS: Questions about cough, wheeze and allergic diseases were asked at 1, 1.5, 2, 3, 4, 5, 6 and 10 years of age in the European prospective cohort of Protection against Allergy: STUdy in Rural Environment (PASTURE). Specific IgE assays were performed at 10 years of age. Questions regarding a cough without a cold were used to build a latent class model of cough over time. RESULTS: Among the 961 children included in the study, apart from the never/infrequent trajectory (59.9%), eight trajectories of cough without a cold were identified: five grouped acute transient classes (24.1%), moderate transient (6.8%), late persistent (4.8%) and early persistent (4.4%). Compared with the never/infrequent trajectory, the other trajectories were significantly associated with wheezing, asthma and allergic rhinitis. For asthma, the strongest association was with the early persistent trajectory (ORa  = 31.00 [14.03-68.51]), which was inversely associated with farm environment (ORa  = 0.39 [0.19-0.77]) and had a high prevalence of cough triggers and unremitting wheeze. Late and early persistent trajectories were also associated with food allergy. Atopic sensitization was only associated with the late persistent trajectory. CONCLUSION: Late and early persistent coughs without a cold are positively associated with atopic respiratory diseases and food allergy. Children having recurrent cough without a cold with night cough and triggers would benefit from an asthma and allergy assessment. Growing up on a farm is associated with reduced early persistent cough.

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.

Early-life residential exposure to moisture damage is associated with persistent wheezing in a Finnish birth cohort.

BACKGROUND AND AIMS: Moisture damage increases the risk for respiratory disorders in childhood. Our aim was to determine whether early age residential exposure to inspector-observed moisture damage or mold is associated with different wheezing phenotypes later in childhood. METHODS: Building inspections were performed by civil engineers, in a standardized manner, in the children's homes-mostly single family and row houses (N = 344)-in the first year of life. The children were followed up with repeated questionnaires until the age of 6 years and wheezing phenotypes-never/infrequent, transient, intermediate, late onset, and persistent-were defined using latent class analyses. The multinomial logistic regression model was used for statistical analysis. RESULTS: A total of 63% (n = 218) had infrequent or no wheeze, 23% (n = 80) had transient and 9.6% (n = 21) had a persistent wheeze. Due to the low prevalence, results for intermediate (3.8%, n = 13) and late-onset wheeze (3.5%, n = 12) were not further evaluated. Most consistent associations were observed with the persistent wheeze phenotype with an adjusted odds ratio (95% confidence intervals) 2.04 (0.67-6.18) for minor moisture damage with or without mold spots (present in 23.8% of homes) and 3.68 (1.04-13.05) for major damage or any moisture damage with visible mold in a child's main living areas (present in 13.4% of homes). Early-age moisture damage or mold in the kitchen was associated with transient wheezing. CONCLUSION: At an early age, residential exposure to moisture damage or mold, can be dose-dependently associated especially with persistent wheezing phenotype later in childhood.

High levels of butyrate and propionate in early life are associated with protection against atopy.

BACKGROUND: Dietary changes are suggested to play a role in the increasing prevalence of allergic diseases and asthma. Short-chain fatty acids (SCFAs) are metabolites present in certain foods and are produced by microbes in the gut following fermentation of fibers. SCFAs have been shown to have anti-inflammatory properties in animal models. Our objective was to investigate the potential role of SCFAs in the prevention of allergy and asthma. METHODS: We analyzed SCFA levels by high-performance liquid chromatography (HPLC) in fecal samples from 301 one-year-old children from a birth cohort and examined their association with early life exposures, especially diet, and allergy and asthma later in life. Data on exposures and allergic diseases were collected by questionnaires. In addition, we treated mice with SCFAs to examine their effect on allergic airway inflammation. RESULTS: Significant associations between the levels of SCFAs and the infant's diet were identified. Children with the highest levels of butyrate and propionate (≥95th percentile) in feces at the age of one year had significantly less atopic sensitization and were less likely to have asthma between 3 and 6 years. Children with the highest levels of butyrate were also less likely to have a reported diagnosis of food allergy or allergic rhinitis. Oral administration of SCFAs to mice significantly reduced the severity of allergic airway inflammation. CONCLUSION: Our results suggest that strategies to increase SCFA levels could be a new dietary preventive option for allergic diseases in children.

Exposure to nonmicrobial N-glycolylneuraminic acid protects farmers' children against airway inflammation and colitis.

BACKGROUND: Childhood exposure to a farm environment has been shown to protect against the development of inflammatory diseases, such as allergy, asthma, and inflammatory bowel disease. OBJECTIVE: We sought to investigate whether both exposure to microbes and exposure to structures of nonmicrobial origin, such as the sialic acid N-glycolylneuraminic acid (Neu5Gc), might play a significant role. METHODS: Exposure to Neu5Gc was evaluated by quantifying anti-Neu5Gc antibody levels in sera of children enrolled in 2 farm studies: the Prevention of Allergy Risk factors for Sensitization in Children Related to Farming and Anthroposophic Lifestyle (PARSIFAL) study (n = 299) and the Protection Against Allergy Study in Rural Environments (PASTURE) birth cohort (cord blood [n = 836], 1 year [n = 734], 4.5 years [n = 700], and 6 years [n = 728]), and we associated them with asthma and wheeze. The effect of Neu5Gc was examined in murine airway inflammation and colitis models, and the role of Neu5Gc in regulating immune activation was assessed based on helper T-cell and regulatory T-cell activation in mice. RESULTS: In children anti-Neu5Gc IgG levels correlated positively with living on a farm and increased peripheral blood forkhead box protein 3 expression and correlated inversely with wheezing and asthma in nonatopic subjects. Exposure to Neu5Gc in mice resulted in reduced airway hyperresponsiveness and inflammatory cell recruitment to the lung. Furthermore, Neu5Gc administration to mice reduced the severity of a colitis model. Mechanistically, we found that Neu5Gc exposure reduced IL-17+ T-cell numbers and supported differentiation of regulatory T cells. CONCLUSIONS: In addition to microbial exposure, increased exposure to non-microbial-derived Neu5Gc might contribute to the protective effects associated with the farm environment.

Latent class analysis reveals clinically relevant atopy phenotypes in 2 birth cohorts.

BACKGROUND: Phenotypes of childhood-onset asthma are characterized by distinct trajectories and functional features. For atopy, definition of phenotypes during childhood is less clear. OBJECTIVE: We sought to define phenotypes of atopic sensitization over the first 6 years of life using a latent class analysis (LCA) integrating 3 dimensions of atopy: allergen specificity, time course, and levels of specific IgE (sIgE). METHODS: Phenotypes were defined by means of LCA in 680 children of the Multizentrische Allergiestudie (MAS) and 766 children of the Protection against allergy: Study in Rural Environments (PASTURE) birth cohorts and compared with classical nondisjunctive definitions of seasonal, perennial, and food sensitization with respect to atopic diseases and lung function. Cytokine levels were measured in the PASTURE cohort. RESULTS: The LCA classified predominantly by type and multiplicity of sensitization (food vs inhalant), allergen combinations, and sIgE levels. Latent classes were related to atopic disease manifestations with higher sensitivity and specificity than the classical definitions. LCA detected consistently in both cohorts a distinct group of children with severe atopy characterized by high seasonal sIgE levels and a strong propensity for asthma; hay fever; eczema; and impaired lung function, also in children without an established asthma diagnosis. Severe atopy was associated with an increased IL-5/IFN-γ ratio. A path analysis among sensitized children revealed that among all features of severe atopy, only excessive sIgE production early in life affected asthma risk. CONCLUSIONS: LCA revealed a set of benign, symptomatic, and severe atopy phenotypes. The severe phenotype emerged as a latent condition with signs of a dysbalanced immune response. It determined high asthma risk through excessive sIgE production and directly affected impaired lung function.

Bacterial microbiota of the upper respiratory tract and childhood asthma.

BACKGROUND: Patients with asthma and healthy controls differ in bacterial colonization of the respiratory tract. The upper airways have been shown to reflect colonization of the lower airways, the actual site of inflammation in asthma, which is hardly accessible in population studies. OBJECTIVE: We sought to characterize the bacterial communities at 2 sites of the upper respiratory tract obtained from children from a rural area and to relate these to asthma. METHODS: The microbiota of 327 throat and 68 nasal samples from school-age farm and nonfarm children were analyzed by 454-pyrosequencing of the bacterial 16S ribosomal RNA gene. RESULTS: Alterations in nasal microbiota but not of throat microbiota were associated with asthma. Children with asthma had lower α- and ß-diversity of the nasal microbiota as compared with healthy control children. Furthermore, asthma presence was positively associated with a specific operational taxonomic unit from the genus Moraxella in children not exposed to farming, whereas in farm children Moraxella colonization was unrelated to asthma. In nonfarm children, Moraxella colonization explained the association between bacterial diversity and asthma to a large extent. CONCLUSIONS: Asthma was mainly associated with an altered nasal microbiota characterized by lower diversity and Moraxella abundance. Children living on farms might not be susceptible to the disadvantageous effect of Moraxella. Prospective studies may clarify whether Moraxella outgrowth is a cause or a consequence of loss in diversity.

The Early Development of Wheeze. Environmental Determinants and Genetic Susceptibility at 17q21.

RATIONALE: Growing up on a farm protects from childhood asthma and early wheeze. Virus-triggered wheeze in infancy predicts asthma in individuals with a genetic asthma risk associated with chromosome 17q21. OBJECTIVES: To test environmental determinants of infections and wheeze in the first year of life, potential modifications of these associations by 17q21, and the implications for different trajectories of wheeze. METHODS: We followed 983 children in rural areas of Europe from birth until age 6 years. Symptoms of wheeze, rhinitis, fever, and environmental exposures were documented with weekly diaries during year 1. Asthma at age 6 was defined as ever having a reported doctor's diagnosis. Single-nucleotide polymorphisms related to ORMDL3 (rs8076131) and GSDMB (rs7216389, rs2290400) at 17q21 were genotyped. MEASUREMENTS AND MAIN RESULTS: Early wheeze was positively associated with presence of older siblings among carriers of known asthma risk alleles at 17q21 (e.g., rs8076131) (adjusted odds ratio [aOR], 1.53; 95% confidence interval [CI], 1.16-2.01). Exposure to farm animal sheds was inversely related to wheeze (aOR, 0.44; 95% CI, 0.33-0.60). Both effects were similarly observed in children with transient wheeze up to age 3 years without subsequent development of asthma (aOR, 1.71 [95% CI, 1.09-2.67]; and aOR, 0.48 [95% CI, 0.30-0.76], respectively). CONCLUSIONS: These findings suggest that the chromosome 17q21 locus relates to episodes of acute airway obstruction common to both transient wheeze and asthma. The previously identified asthma risk alleles are the ones susceptible to environmental influences. Thus, this gene-environment interaction reveals two faces of 17q21: The same genotype constitutes genetic risk and allows for environmental protection, thereby providing options for prospective prevention strategies.

Comparison of Oropharyngeal Microbiota from Children with Asthma and Cystic Fibrosis.

A genuine microbiota resides in the lungs which emanates from the colonization by the oropharyngeal microbiota. Changes in the oropharyngeal microbiota might be the source of dysbiosis observed in the lower airways in patients suffering from asthma or cystic fibrosis (CF). To examine this hypothesis, we compared the throat microbiota from healthy children (n = 62) and that from children with asthma (n = 27) and CF (n = 57) aged 6 to 12 years using 16S rRNA amplicon sequencing. Our results show high levels of similarities between healthy controls and children with asthma and CF revealing the existence of a core microbiome represented by Prevotella, Streptococcus, Neisseria, Veillonella, and Haemophilus. However, in CF, the global diversity, the bacterial load, and abundances of 53 OTUs were significantly reduced, whereas abundances of 6 OTUs representing opportunistic pathogens such as Pseudomonas, Staphylococcus, and Streptococcus were increased compared to those in healthy controls controls and asthmatics. Our data reveal a core microbiome in the throat of healthy children that persists in asthma and CF indicating shared host regulation favoring growth of commensals. Furthermore, we provide evidence for dysbiosis with a decrease in diversity and biomass associated with the presence of known pathogens consistent with impaired host defense in children with CF.

Polymorphisms related to ORMDL3 are associated with asthma susceptibility, alterations in transcriptional regulation of ORMDL3, and changes in TH2 cytokine levels.

BACKGROUND: Chromosome 17q21, harboring the orosomucoid 1-like 3 (ORMDL3) gene, has been consistently associated with childhood asthma in genome-wide association studies. OBJECTIVE: We investigated genetic variants in and around ORMDL3 that can change the function of ORMDL3 and thus contribute to asthma susceptibility. METHODS: We performed haplotype analyses and fine mapping of the ORMDL3 locus in a cross-sectional (International Study of Asthma and Allergies in Childhood Phase II, n = 3557 total subjects, n = 281 asthmatic patients) and case-control (Multicenter Asthma Genetics in Childhood Study/International Study of Asthma and Allergies in Childhood Phase II, n = 1446 total subjects, n = 763 asthmatic patients) data set to identify putative causal single nucleotide polymorphisms (SNPs) in the locus. Top asthma-associated polymorphisms were analyzed for allele-specific effects on transcription factor binding and promoter activity in vitro and gene expression in PBMCs after stimulation ex vivo. RESULTS: Two haplotypes (H1 and H2) were significantly associated with asthma in the cross-sectional (P = 9.9 × 10(-5) and P = .0035, respectively) and case-control (P = 3.15 × 10(-8) and P = .0021, respectively) populations. Polymorphisms rs8076131 and rs4065275 were identified to drive these effects. For rs4065275, a quantitative difference in transcription factor binding was found, whereas for rs8076131, changes in upstream stimulatory factor 1 and 2 transcription factor binding were observed in vitro by using different cell lines and PBMCs. This might contribute to detected alterations in luciferase activity paralleled with changes in ORMDL3 gene expression and IL-4 and IL-13 cytokine levels ex vivo in response to innate and adaptive stimuli in an allele-specific manner. Both SNPs were in strong linkage disequilibrium with asthma-associated 17q21 SNPs previously related to altered ORMDL3 gene expression. CONCLUSION: Polymorphisms in a putative promoter region of ORMDL3, which are associated with childhood asthma, alter transcriptional regulation of ORMDL3, correlate with changes in TH2 cytokines levels, and therefore might contribute to the childhood asthma susceptibility signal from 17q21.

Consumption of unprocessed cow's milk protects infants from common respiratory infections.

BACKGROUND: Breast-feeding is protective against respiratory infections in early life. Given the co-evolutionary adaptations of humans and cattle, bovine milk might exert similar anti-infective effects in human infants. OBJECTIVE: To study effects of consumption of raw and processed cow's milk on common infections in infants. METHODS: The PASTURE birth cohort followed 983 infants from rural areas in Austria, Finland, France, Germany, and Switzerland, for the first year of life, covering 37,306 person-weeks. Consumption of different types of cow's milk and occurrence of rhinitis, respiratory tract infections, otitis, and fever were assessed by weekly health diaries. C-reactive protein levels were assessed using blood samples taken at 12 months. RESULTS: When contrasted with ultra-heat treated milk, raw milk consumption was inversely associated with occurrence of rhinitis (adjusted odds ratio from longitudinal models [95% CI]: 0.71 [0.54-0.94]), respiratory tract infections (0.77 [0.59-0.99]), otitis (0.14 [0.05-0.42]), and fever (0.69 [0.47-1.01]). Boiled farm milk showed similar but weaker associations. Industrially processed pasteurized milk was inversely associated with fever. Raw farm milk consumption was inversely associated with C-reactive protein levels at 12 months (geometric means ratio [95% CI]: 0.66 [0.45-0.98]). CONCLUSIONS: Early life consumption of raw cow's milk reduced the risk of manifest respiratory infections and fever by about 30%. If the health hazards of raw milk could be overcome, the public health impact of minimally processed but pathogen-free milk might be enormous, given the high prevalence of respiratory infections in the first year of life and the associated direct and indirect costs.

Clinical and epidemiologic phenotypes of childhood asthma.

RATIONALE: Clinical and epidemiologic approaches have identified two distinct sets of classifications for asthma and wheeze phenotypes. OBJECTIVES: To compare epidemiologic phenotype definitions identified by latent class analysis (LCA) with clinical phenotypes based on patient histories, diagnostic work-up, and treatment responses. To relate phenotypes to genetic and environmental determinants as well as diagnostic and treatment-related parameters. METHODS: LCA was performed in an international multicenter birth cohort based on yearly questions about current wheeze until age 6 years. Associations of wheeze classes and clinical phenotypes with asthma-related characteristics such as atopy, lung function, fraction of exhaled nitric oxide, and medication use were calculated using regression models. MEASUREMENTS AND MAIN RESULTS: LCA identified five classes, which verified the clinically defined wheeze phenotypes with high sensitivity and specificity; the respective receiver operating characteristics curves displayed an area under the curve ranging from 84% (frequent wheeze) to 85% (asthma diagnosis) and 87% (unremitting wheeze) to 97% (recurrent unremitting wheeze). Recurrent unremitting wheeze was the most specific and unremitting wheeze at least once the most sensitive definition. The latter identified a subgroup of children with decreased lung function, increased genetic risk, and in utero smoke exposure (ODDS RATIO, 2.03; 95% CONFIDENCE INTERVAL, 1.12-3.68; P = 0.0191), but without established asthma diagnosis and treatment. CONCLUSIONS: Clinical phenotypes were well supported by LCA analysis. The hypothesis-free LCA phenotypes were a useful reference for comparing clinical phenotypes. Thereby, we identified children with clinically conspicuous but undiagnosed disease. Because of their high area under the curve values, clinical phenotypes such as (recurrent) unremitting wheeze emerged as promising alternative asthma definitions for epidemiologic studies.

Increased food diversity in the first year of life is inversely associated with allergic diseases.

BACKGROUND: The role of dietary factors in the development of allergies is a topic of debate, especially the potential associations between infant feeding practices and allergic diseases. Previously, we reported that increased food diversity introduced during the first year of life reduced the risk of atopic dermatitis. OBJECTIVE: In this study we investigated the association between the introduction of food during the first year of life and the development of asthma, allergic rhinitis, food allergy, or atopic sensitization, taking precautions to address reverse causality. We further analyzed the association between food diversity and gene expression of T-cell markers and of Cε germline transcript, reflecting antibody isotype switching to IgE, measured at 6 years of age. METHODS: Eight hundred fifty-six children who participated in a birth cohort study, Protection Against Allergy Study in Rural Environments/EFRAIM, were included. Feeding practices were reported by parents in monthly diaries during the first year of life. Data on environmental factors and allergic diseases were collected from questionnaires administered from birth up to 6 years of age. RESULTS: An increased diversity of complementary food introduced in the first year of life was inversely associated with asthma with a dose-response effect (adjusted odds ratio with each additional food item introduced, 0.74 [95% CI, 0.61-0.89]). A similar effect was observed for food allergy and food sensitization. Furthermore, increased food diversity was significantly associated with an increased expression of forkhead box protein 3 and a decreased expression of Cε germline transcript. CONCLUSION: An increased diversity of food within the first year of life might have a protective effect on asthma, food allergy, and food sensitization and is associated with increased expression of a marker for regulatory T cells.

Increased regulatory T-cell numbers are associated with farm milk exposure and lower atopic sensitization and asthma in childhood.

BACKGROUND: European cross-sectional studies have suggested that prenatal and postnatal farm exposure decreases the risk of allergic diseases in childhood. Underlying immunologic mechanisms are still not understood but might be modulated by immune-regulatory cells early in life, such as regulatory T (Treg) cells. OBJECTIVE: We sought to assess whether Treg cells from 4.5-year-old children from the Protection against Allergy: Study in Rural Environments birth cohort study are critical in the atopy and asthma-protective effect of farm exposure and which specific exposures might be relevant. METHODS: From 1133 children, 298 children were included in this study (149 farm and 149 reference children). Detailed questionnaires until 4 years of age assessed farming exposures over time. Treg cells were characterized as upper 20% CD4(+)CD25(+) forkhead box protein 3 (FOXP3)(+) (intracellular) in PBMCs before and after stimulation (with phorbol 12-myristate 13-acetate/ionomycin or LPS), and FOXP3 demethylation was assessed. Atopic sensitization was defined by specific IgE measurements; asthma was defined by a doctor's diagnosis. RESULTS: Treg cells were significantly increased in farm-exposed children after phorbol 12-myristate 13-acetate/ionomycin and LPS stimulation. Exposure to farm milk was defined as a relevant independent farm-related exposure supported by higher FOXP3 demethylation. Treg cell (upper 20% CD4(+)CD25(+), FOXP3(+) T cells) numbers were significantly negatively associated with doctor-diagnosed asthma (LPS stimulated: adjusted odds ratio, 0.26; 95% CI, 0.08-0.88) and perennial IgE (unstimulated: adjusted odds ratio, 0.21; 95% CI, 0.08-0.59). Protection against asthma by farm milk exposure was partially mediated by Treg cells. CONCLUSIONS: Farm milk exposure was associated with increased Treg cell numbers on stimulation in 4.5-year-old children and might induce a regulatory phenotype early in life, potentially contributing to a protective effect for the development of childhood allergic diseases.

Atopic sensitization in the first year of life.

BACKGROUND: There is conflicting evidence on whether allergen-specific memory is primed prenatally, whether this priming affects persistent immunologic effects, and whether it is modulated by the first environmental exposures in infancy. OBJECTIVE: We sought to explore the course of atopic sensitization between birth and 12 months of age. METHODS: Specific IgE levels for 6 food and 13 common inhalant allergens were assessed in cord blood and 1-year blood samples in the Protection against Allergy-Study in Rural Environments (PASTURE) birth cohort including 793 children from rural regions of 5 European countries. Detailed information on children's health, nutrition, and farm-related exposures was gathered by using a pregnancy questionnaire, 2 questionnaires at 2 and 12 months of age, and a diary covering the time in between. RESULTS: Sensitization was more common at 12 months of age than at birth for almost all specificities. On an individual level, persistent sensitization to the same allergens was rare (1%), whereas transient (only at birth, 11%) and incident (only at 12 months, 34%) sensitization was seen in substantial proportions of children. Associations of transient sensitization with maternal sensitization differed with the allergen specificities, with the strongest associations for food allergens (odds ratio [OR], 10.6; 95% CI, 6.0-18.6) and the weakest associations for seasonal allergens (OR, 1.64; 95% CI, 0.94-2.86). Associations of maternal sensitization with incident sensitization were also seen. Incident sensitization was related to distinct prenatal and postnatal environmental exposures of mother and child, such as consumption of cereals for incident sensitization to seasonal allergens (OR, 0.66; 95% CI, 0.50-0.88). CONCLUSION: IgE sensitization patterns change between birth and 12 months and are related to maternal and environmental influences.

Genetic variants regulating ORMDL3 expression contribute to the risk of childhood asthma.

Asthma is caused by a combination of poorly understood genetic and environmental factors. We have systematically mapped the effects of single nucleotide polymorphisms (SNPs) on the presence of childhood onset asthma by genome-wide association. We characterized more than 317,000 SNPs in DNA from 994 patients with childhood onset asthma and 1,243 non-asthmatics, using family and case-referent panels. Here we show multiple markers on chromosome 17q21 to be strongly and reproducibly associated with childhood onset asthma in family and case-referent panels with a combined P value of P < 10(-12). In independent replication studies the 17q21 locus showed strong association with diagnosis of childhood asthma in 2,320 subjects from a cohort of German children (P = 0.0003) and in 3,301 subjects from the British 1958 Birth Cohort (P = 0.0005). We systematically evaluated the relationships between markers of the 17q21 locus and transcript levels of genes in Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines from children in the asthma family panel used in our association study. The SNPs associated with childhood asthma were consistently and strongly associated (P < 10(-22)) in cis with transcript levels of ORMDL3, a member of a gene family that encodes transmembrane proteins anchored in the endoplasmic reticulum. The results indicate that genetic variants regulating ORMDL3 expression are determinants of susceptibility to childhood asthma.

Development of atopic dermatitis according to age of onset and association with early-life exposures.

BACKGROUND: Environmental factors can affect the development of atopic dermatitis, and this was described to be already effective during pregnancy and in early life. An important early postnatal exposure is nutrition, although its association with allergic disease remains unclear. OBJECTIVE: We sought to determine prospectively whether early postnatal exposures, such as the introduction to complementary food in the first year of life, are associated with the development of atopic dermatitis, taking into account the reverse causality. METHODS: One thousand forty-one children who participated in the Protection Against Allergy-Study in Rural Environments birth cohort study were included in the current study. Atopic dermatitis was defined by a doctor's diagnosis reported by the parents of children up to 4 years of age, by questionnaires, and/or by positive SCORAD scores from 1 year of age and according to the age of onset within or after the first year of life. Feeding practices were reported by parents in monthly diaries between the 3rd and 12th months of life. RESULTS: The diversity of introduction of complementary food in the first year of life was associated with a reduction in the risk of having atopic dermatitis with onset after the first year of life (adjusted odds ratio for atopic dermatitis with each additional major food item introduced, 0.76; 95% CI, 0.65-0.88). The introduction of yogurt in the first year of life also reduced the risk for atopic dermatitis (adjusted odds ratio, 0.41; 95% CI, 0.23-0.73). CONCLUSION: As early-life exposure, the introduction of yogurt and the diversity of food introduced in the first year of life might have a protective effect against atopic dermatitis.