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Over the past decade, "omics" approaches have advanced our understanding of the molecular programming of the airways in humans. Several studies have identified potential molecular mechanisms that contribute to early life epigenetic reprogramming, including DNA methylation, histone modifications, microRNAs, and the homeostasis of the respiratory mucosa (epithelial function and microbiota). Current evidence supports the notion that early infancy is characterized by heightened susceptibility to airway genetic reprogramming in response to the first exposures in life, some of which can have life-long consequences. Here, we summarize and analyze the latest insights from studies that support a novel epigenetic paradigm centered on human maturational and developmental programs including three cardinal elements: genes, environment, and developmental timing. The combination of these factors is likely responsible for the functional trajectory of the respiratory system at the molecular, functional, and clinical levels.

OBJECTIVE Initiation of continuous positive airway pressure (CPAP) in children with severe obstructive sleep apnea syndrome (OSAS) is challenging and the distinct features of the subset of children requiring CPAP are poorly defined. Asthma often coexists with OSAS in children. The goal of this study was to explore the influence of asthma in the need for CPAP therapy in children with severe OSAS. HYPOTHESIS Asthmatic children with severe OSAS have higher probability of needing CPAP than children with severe OSAS without asthma. METHODS Cross-sectional study of clinical presentation, individual risk factors, and initial overnight polysomnogram (PSG) parameters in children with severe OSAS. Severe OSAS was defined as an obstructive apnea hypopnea index ≥10/h. The association between asthma and CPAP initiation was studied individually and adjusted by pertinent covariates. RESULTS Four hundred eligible children (mean age 7 years, ±SD 5.3) with severe OSAS were enrolled and 133 individuals (33%) were identified to have asthma. The proportion of children needing CPAP was significantly higher in asthmatics with severe OSAS (29%) compared to those with OSAS alone (14%) (P

Background Human Rhinovirus (HRV) has been identified as the most common cause of acute respiratory infections and hospitalizations in premature children. It is unclear if premature children are more susceptible to HRV due to their decreased pulmonary reserve or because they have enhanced lower airway reactivity to HRV. Methods We conducted a retrospective analysis of the clinical respiratory presentation of all PCR-confirmed HRV infections in full-term and premature children aged ≤3 years in our institution. Standardized respiratory distress scores were developed to examine lower airway obstruction (i.e., wheezing, hyperinflation, and sub-costal retractions) along with markers of decreased pulmonary reserve (hypoxemia and tachypnea) in young children with HRV infections. Demographic and clinical variables were obtained from reviewing electronic medical records (EMR). Results This study included a total of 205 children; 71% of these children were born full-term (>37 weeks gestation), 10% preterm (32–37 weeks) and 19% severely premature (

Background Airway epithelial cells (AEC) are quite difficult to access in newborns and infants. It is critically important to develop robust life-extended models to conduct translational studies in this age group. We propose the use of a recently described cell-culture technology (conditionally reprogrammed cells - CRC) to generate continuous primary cell cultures from nasal and bronchial AEC of young children. Methods We collected nasal and/or bronchial AEC from a total of 23 subjects of different ages including newborns/infants/toddlers (0-2 years; N=9), school age children (4-11 years; N=6), and a group of adolescent/adult donors (N=8). For CRC generation, we used conditioned medium from mitotically inactivated 3T3 fibroblasts and Rho-associated kinase (ROCK) inhibitor (Y-27632). Antiviral immune responses were studied using 25 key antiviral genes and protein production of type III epithelial interferon (IFN λ1) after double stranded (ds) RNA exposure. Results CRC derived from primary AEC of neonates/infants and young children exhibited: 1) augmented proliferative capacity and life-extension, 2) preserved airway epithelial phenotype after multiple passages, 3) robust immune responses characterized by expression of innate antiviral genes and parallel nasal/bronchial production of IFN λ1 after exposure to dsRNA, and 4) induction of airway epithelial inflammatory and remodeling responses to dsRNA (e.g. CXCL8 and MMP9). Conclusion Conditional reprogramming of AEC from young children is a feasible and powerful translational approach to investigate early-life airway epithelial immune responses in humans. This article is protected by copyright. All rights reserved.

Infants requiring hospitalization due to a viral lower respiratory tract infection (LRTI) have a high risk of developing recurrent respiratory illnesses in early life and asthma beyond childhood. Notably, all validated clinical scales for viral LRTI have focused on predicting acute severity instead of recurrence. We present a novel clinical approach combining individual risk factors with bedside clinical parameters to predict recurrence after viral LRTI hospitalization in young children. A retrospective longitudinal cohort of young children (≤3 years) designed to define clinical predictive factors of recurrent respiratory illnesses within 12 months after hospitalization due to PCR-confirmed viral LRTI. Data collection was through electronic medical record. We included 138 children hospitalized with viral LRTI. Using automatic stepwise logistic model selection, we found that the strongest predictors of recurrence in infants hospitalized for the first time were severe prematurity (≤32 weeks’ gestational age, OR=5.19; 95% CI 1.76 to 15.32; p=0.002) and a clinical score that weighted hypoxemia, subcostal retractions and wheezing (OR=3.33; 95% CI 1.59 to 6.98; p

Background: Human metapneumovirus (HMPV) is a recently discovered respiratory pathogen of the family Paramyxoviridae, the same family as that of respiratory syncytial virus (RSV). Premature children are at high risk of severe RSV infections, however, it is unclear whether HMPV infection is more severe in hospitalized children with a history of severe prematurity. Methods: We conducted a retrospective analysis of the clinical respiratory presentation of all polymerase chain reaction-confirmed HMPV infections in preschool-age children ( 5 years) with and without history of severe prematurity (

This paper presents a quantitative imaging method and software technology to predict the risk and assess the severity of respiratory diseases in premature babies by fusing information from multiple sources: non-invasive low-radiation chest X-ray (CXR) imaging and clinical parameters. Prematurity is the largest single cause of death in children under five in the world. Lower respiratory tract infections (LRTI) are the top cause of hospitalization and mortality in prematurity. However, there is no objective clinical marker to predict and prevent severe LRTI in the 15 million babies born prematurely every year worldwide. Traditionally, imaging biomarkers of lung disease from computed tomography have been successfully used in adults, but they entail heightened risks for children due to cumulative radiation and the need for sedation. The proposed technology is the first approach that uses low-radiation CXR imaging to predict hospitalization due to LRTI in prematurity. The method uses deep learning to quantify heterogeneous patterns (air trapping and irregular opacities) in the chest, which are combined with clinical parameters to predict the risk of LRTI. Our preliminary results obtained using a data obtained from ten premature subjects with LRTI showed high correlation between our imaging biomarkers and the rehospitalization of these subjects R2=0.98).

Background Rhinitis and obstructive sleep apnea (OSA) often coexist during childhood. To delineate this clinical association, we examined OSA severity and polysomnogram (PSG) features in children with rhinitis and OSA. Given that rapid-eye-movement (REM) sleep is characterized by nasal congestion, we hypothesized that children with rhinitis have more REM-related breathing abnormalities. Methods We conducted a retrospective cross-sectional analysis of 145 children with PSG-diagnosed A. Outcomes included PSG parameters and obstructive apnea–hypopnea index (OAHI) during REM and non-REM. Linear multivariable models examined the joint effect of rhinitis and OSA parameters with control for potential confounders. Results Rhinitis was present in 43% of children with OSA (n = 63) but overall OAHI severity was unaffected by the presence of rhinitis. In contrast, OAHI during REM sleep in children with moderate-severe OSA was significantly increased subjects with rhinitis and OSA (44.1/hr; SE = 6.4) compared with those with OSA alone (28.2/hr; SE = 3.8). Conclusion Rhinitis is highly prevalent in children with OSA. Although OSA is not more severe in children with rhinitis, they do have a distinct OSA phenotype characterized by more REM-related OSA. Further research is needed to delineate the link between REM-sleep and the physiology of the nose during health and disease.

Background Innate immune responses are fine-tuned by small noncoding RNA molecules termed microRNAs (miRs) that modify gene expression in response to the environment. During acute infections, miRs can be secreted in extracellular vesicles (EV) to facilitate cell-to-cell genetic communication. The purpose of this study was to characterize the baseline population of miRs secreted in EVs in the airways of young children (airway secretory microRNAome) and examine the changes during rhinovirus (RV) infection, the most common cause of asthma exacerbations and the most important early risk factor for the development of asthma beyond childhood. Methods Nasal airway secretions were obtained from children (≤3 yrs. old) during PCR-confirmed RV infections (n = 10) and age-matched controls (n = 10). Nasal EVs were isolated with polymer-based precipitation and global miR profiles generated using NanoString microarrays. We validated our in vivo airway secretory miR data in an in vitro airway epithelium model using apical secretions from primary human bronchial epithelial cells (HBEC) differentiated at air-liquid interface (ALI). Bioinformatics tools were used to determine the unified (nasal and bronchial) signature airway secretory miRNAome and changes during RV infection in children. Results Multiscale analysis identified four signature miRs comprising the baseline airway secretory miRNAome: hsa-miR-630, hsa-miR-302d-3p, hsa- miR-320e, hsa-miR-612. We identified hsa-miR-155 as the main change in the baseline miRNAome during RV infection in young children. We investigated the potential biological relevance of the airway secretion of hsa-mir-155 using in silico models derived from gene datasets of experimental in vivo human RV infection. These analyses confirmed that hsa-miR-155 targetome is an overrepresented pathway in the upper airways of individuals infected with RV. Conclusions Comparative analysis of the airway secretory microRNAome in children indicates that RV infection is associated with airway secretion of EVs containing miR-155, which is predicted in silico to regulate antiviral immunity. Further characterization of the airway secretory microRNAome during health and disease may lead to completely new strategies to treat and monitor respiratory conditions in all ages.

Accurate assessment of severity of viral respiratory illnesses (VRIs) allows early interventions to prevent morbidity and mortality in young children. This paper proposes a novel imaging biomarker framework with chest X-ray image for assessing VRI's severity in infants, developed specifically to meet the distinct challenges for pediatric population. The proposed framework integrates three novel technical contributions: a) lung segmentation using weighted partitioned active shape model, b) obtrusive object removal using graph cut segmentation with asymmetry constraint, and c) severity quantification using information-theoretic heterogeneity measures. This paper presents our pilot experimental results with a dataset of 148 images and the ground-truth severity scores given by a board-certified pediatric pulmonologist, demonstrating the effectiveness and clinical relevance of the presented framework.

Background: It is unknown why human metapneumovirus (HMPV) and respiratory syncytial virus (RSV) cause severe respiratory infection in children, particularly in premature infants. Our aim was to investigate if there are defective airway antiviral responses to these viruses in young children with history of prematurity. Methods: Nasal airway secretions were collected from 140 children ≤3 y old without detectable virus (n = 80) or with PCR-confirmed HMPV or RSV infection (n = 60). Nasal protein levels of IFNγ, CCL5/RANTES, IL-10, IL-4, and IL-17 were determined using a multiplex magnetic bead immunoassay. Results: Full-term children with HMPV and RSV infection had increased levels of nasal airway IFNγ, CCL5, and IL-10 along with an elevation in Th1 (IFNγ)/Th2 (IL-4) ratios, which is expected during antiviral responses. In contrast, HMPV-infected premature children (< 32 wk gestation) did not exhibit increased Th1/Th2 ratios or elevated nasal airway secretion of IFNγ, CCL5, and IL-10 relative to uninfected controls. Conclusion: Our study is the first to demonstrate that premature infants have defective IFNγ, CCL5/RANTES, and IL-10 airway responses during HMPV infection and provides novel insights about the potential reason why HMPV causes severe respiratory disease in children with history of prematurity.

To the Editor: Rhinovirus wheezing illnesses during early childhood are strongly linked with development of asthma later in life [1]. Indeed, rhinovirus infection in the first 3 years of life is associated with an almost 10-fold increase in risk for asthma at age 6 years [1]. The exact mechanism by which rhinovirus elicits a pro-asthmatic propensity in young children is largely unknown, but is purportedly related to a viral-induced T-helper cell (Th)2 airway inflammatory response [2]. Interestingly, the discovery of interactions between epithelial innate immunity and adaptive allergic responses has unveiled new potential links between rhinovirus and asthma. Of particular interest is the epithelial-derived cytokine thymic stromal lymphopoietin (TSLP), which is considered a “master Th2 cytokine” because it primes the differentiation of naive T0 cells into Th2 lymphocytes via activation of antigen presenting cells [3]. TSLP is induced by rhinovirus infection or by exposure to double stranded (ds)RNA (viral surrogate) in the lungs of allergic mice [4], and in human bronchial epithelial cells (HBEC) [5]. Together, these data suggest that TSLP may be the missing link between innate antiviral epithelial immunity and the Th2 immune response characteristic of asthma. This cross-sectional preliminary study aimed to investigate whether rhinovirus infections that occur naturally during the first 3 years of life are associated with elevated airway TSLP levels and enhanced Th2 responses, which may potentially facilitate the establishment of rhinovirus-induced pro-asthmatic changes during early childhood. We measured nasal airway TSLP, Th2 cytokines and antiviral responses in nasal washes obtained from newborns, infants and toddlers (≤3 years) with PCR-confirmed acute …

Cytomegalovirus (CMV) is a prevalent pathogen in the immunocompromised host and invasive pneumonia is a feared complication of the virus in this population. In this pediatric case series we characterized CMV lung infection in 15 non-HIV infected children (median age 3 years; IQR 0.2–4.9 years), using current molecular and imaging diagnostic modalities, in combination with respiratory signs and symptoms. The most prominent clinical and laboratory findings included cough (100%), hypoxemia (100%), diffuse adventitious breath sounds (100%) and increased respiratory effort (93%). All patients had abnormal lung images characterized by ground glass opacity/consolidation in 80% of cases. CMV was detected in the lung either by CMV PCR in bronchoalveolar lavage (82% detection rate) or histology/immunohistochemistry in lung biopsy (100% detection rate). CMV caused respiratory failure in 47% of children infected and the overall mortality rate was 13.3%. Conclusion: CMV pneumonia is a potential lethal disease in non-HIV infected children that requires a high-index of suspicion. Common clinical and radiological patterns such as hypoxemia, diffuse adventitious lung sounds and ground-glass pulmonary opacities may allow early identification of CMV lung infection in the pediatric population, which may lead to prompt initiation of antiviral therapy and better clinical outcomes.

Background Thymic stromal lymphoproetin (TSLP) is a cytokine secreted by the airway epithelium in response to respiratory viruses and it is known to promote allergic Th2 responses in asthma. This study investigated whether virally-induced secretion of TSLP is directional in nature (apical vs. basolateral) and/or if there are TSLP-mediated effects occurring at both sides of the bronchial epithelial barrier in the asthmatic state. Methods Primary human bronchial epithelial cells (HBEC) from control (n = 3) and asthmatic (n = 3) donors were differentiated into polarized respiratory tract epithelium under air-liquid interface (ALI) conditions and treated apically with dsRNA (viral surrogate) or TSLP. Sub-epithelial effects of TSLP were examined in human airway smooth muscle cells (HASMC) from normal (n = 3) and asthmatic (n = 3) donors. Clinical experiments examined nasal airway secretions obtained from asthmatic children during naturally occurring rhinovirus-induced exacerbations (n = 20) vs. non-asthmatic uninfected controls (n = 20). Protein levels of TSLP, CCL11/eotaxin-1, CCL17/TARC, CCL22/MDC, TNF-α and CXCL8 were determined with a multiplex magnetic bead assay. Results Our data demonstrate that: 1) Asthmatic HBEC exhibit an exaggerated apical, but not basal, secretion of TSLP after dsRNA exposure; 2) TSLP exposure induces unidirectional (apical) secretion of CCL11/eotaxin-1 in asthmatic HBEC and enhanced CCL11/eotaxin-1 secretion in asthmatic HASMC; 3) Rhinovirus-induced asthma exacerbations in children are associated with in vivo airway secretion of TSLP and CCL11/eotaxin-1. Conclusions There are virally-induced TSLP-driven secretory immune responses at both sides of the bronchial epithelial barrier characterized by enhanced CCL11/eotaxin-1 secretion in asthmatic airways. These results suggest a new model of TSLP-mediated eosinophilic responses in the asthmatic airway during viral-induced exacerbations.

Rationale. The sleep-related factors that modulate the nocturnal worsening of asthma in children are poorly understood. This study addressed the hypothesis that asthmatic children have a REM sleep-related vulnerability trait that is independent of OSA.Methods. We conducted a retrospective cross-sectional analysis of pulse-oximetry signals obtained during REM and NREM sleep in control and asthmatic children (n=134). Asthma classification was based on preestablished clinical criteria. Multivariate linear regression model was built to control for potential confounders (significance levelP≤0.05).Results. Our data demonstrated that (1) baseline nocturnal respiratory parameters were not significantly different in asthmatic versus control children, (2) the maximal % of SaO2desaturation during REM, but not during NREM, was significantly higher in asthmatic children, and (3) multivariate analysis revealed that the association between asthma and REM-related maximal % SaO2desaturation was independent of demographic variables.Conclusion. These results demonstrate that children with asthma have a REM-related vulnerability trait that impacts oxygenation independently of OSA. Further research is needed to delineate the REM sleep neurobiological mechanisms that modulate the phenotypical expression of nocturnal asthma in children.