Immune profiling and functional analysis of NK and T cells in ataxia telangiectasia.

Ataxia telangiectasia (AT) is a rare autosomal-recessive disorder characterized by profound neurodegeneration, combined immunodeficiency, and an increased risk for malignant diseases. Treatment options for AT are limited, and the long-term survival prognosis for patients remains grim, primarily due to the emergence of chronic respiratory pathologies, malignancies, and neurological complications. Understanding the dysregulation of the immune system in AT is fundamental for the development of novel treatment strategies. In this context, we performed a retrospective longitudinal immunemonitoring of lymphocyte subset distribution in a cohort of AT patients (n = 65). Furthermore, we performed FACS analyses of peripheral blood mononuclear cells from a subgroup of 12 AT patients to examine NK and T cells for the expression of activating and functional markers. We observed reduced levels of peripheral blood CD3+CD8+ cytotoxic T cells, CD3+CD4+ T helper cells, and CD19+ B cells, whereas the amount of CD3--CD56+ NK cells and CD3+CD56+ NKT-like cells was similar compared with age-matched controls. Notably, there was no association between the age-dependent kinetic of T-, B-, or NK-cell counts and the occurrence of malignancy in AT patients. Additionally, our results indicate an altered NK- and T-cell response to cytokine stimulation in AT with increased levels of TRAIL, FasL, and CD16 expression in NK cells, as well as an elevated activation level of T cells in AT with notably higher expression levels of IFN-γ, CD107a, TRAIL, and FasL. Together, these findings imply function alterations in AT lymphocytes, specifically in T and NK cells, shedding light on potential pathways for innovative therapies.

Dual Role of microRNA-146a in Experimental Inflammation in Human Pulmonary Epithelial and Immune Cells and Expression in Inflammatory Lung Diseases.

microRNA (miR)-146a emerges as a promising post-transcriptional regulator in various inflammatory diseases with different roles for the two isoforms miR-146a-5p and miR-146a-3p. The present study aimed to examine the dual role of miR-146a-5p and miR-146a 3p in the modulation of inflammation in human pulmonary epithelial and immune cells in vitro as well as their expression in patients with inflammatory lung diseases. Experimental inflammation in human A549, HL60, and THP1 via the NF-kB pathway resulted in the major upregulation of miR-146a-5p and miR-146a-3p expression, which was partly cell-specific. Modulation by transfection with miRNA mimics and inhibitors demonstrated an anti-inflammatory effect of miR-146a-5p and a pro-inflammatory effect of miR-146a-3p, respectively. A mutual interference between miR-146a-5p and miR-146a-3p was observed, with miR-146a-5p exerting a predominant influence. In vivo NGS analyses revealed an upregulation of miR-146a-3p in the blood of patients with cystic fibrosis and bronchiolitis obliterans, while miR-146a-5p levels were downregulated or unchanged compared to controls. The reverse pattern was observed in patients with SARS-CoV-2 infection. In conclusion, miR-146a-5p and miR-146a-3p are two distinct but interconnected miRNA isoforms with opposing functions in inflammation regulation. Understanding their interaction provides important insights into the progression and persistence of inflammatory lung diseases and might provide potential therapeutic options.

Impulse oscillometry bronchodilator response in preschool children.

BACKGROUND: In preschoolers, performing an acceptable spirometry and measuring bronchodilator response (BDR) is challenging; in this context, impulse oscillometry (IOS) represents a valid alternative. However, more studies on the standardization of BDR for IOS in young children are required. OBJECTIVE: The objective of the study was to identify optimal thresholds to define a positive BDR test with IOS in preschoolers with suspected asthma. METHODS: Children aged 3-6 years with suspected asthma and their lung function investigated with both IOS and spirometry pre- and post-BDR were retrospectively analyzed. The spirometric BDR was defined as positive when the change of FEV1 was ≥12% or ≥200 mL. The oscillometric BDR was defined as positive in case of change of at least -40% in R5, +50% in X5, and -80% in AX. RESULTS: Among 72 patients, 36 (age 5.2 ± 1 years; 64% boys) were selected for the subsequent analysis according to ATS/ERS quality criteria of measurements; specifically, 19 patients did not meet IOS and 36 did not meet spirometry criteria. The spirometric BDR was found positive in seven subjects (19.4%); conversely, a positive oscillometric BDR was identified in four patients (11.1%). No patient presented a positive BDR response with both methods. In IOS, the mean decrease in R5 and AX was 19.9% ± 10% and 44% ± 22.1%, and the mean increase in X5 was 23.3% ± 17.8%, respectively. A decrease in R5 of 25.7% (AUC 0.77, p = .03) and an increase in X5 of 25.7% (AUC 0.75, p = .04) showed the best combination of sensitivity and specificity to detect an increase of FEV1 ≥ 12% and/or ≥200 mL. CONCLUSION: The IOS represents a valid alternative to spirometry to measure BDR in preschool children and should be the gold standard in this age group. We are considering a decrease of 26% in R5 and an increase of 26% in X5 as diagnostic threshold for BDR.

Are Questionnaires Helpful To Predict Exercise-Induced Bronchoconstriction (EIB) And Exercise-Induced Laryngeal Obstruction (EILO)?

OBJECTIVE: Exercise induced laryngeal obstruction (EILO) is an important differential diagnosis to exercise induced bronchoconstriction (EIB) and diagnosed via continuous laryngoscopy while exercising (CLE). However, availability of CLE is limited to specialized centres. And without CLE EILO is often misdiagnosed as EIB. Therefore it is essential to carefully preselect potential EILO candidates. Aim of this study was to investigate whether two short questionnaires -Asthma Control Test (ACT) and Dyspnea Index (DI) evaluating upper airway-related dyspnea- can differentiate between EIB and EILO. METHODS: Patients with dyspnea while exercising were analysed with an exercise challenge in the cold chamber (ECC) to diagnose EIB in visit 1 (V1), as appropriate a CLE in visit 2 (V2, 4-6 weeks after V1) and ACT and DI in V1 and V2. EIB patients were treated with asthma medication after V1. RESULTS: Complete dataset of 36 subjects were gathered. The ACT showed lower values in V2 in EILO compared to EIB patients. A lack of improvement in ACT in V2 after asthma medication of EIB patients is suspicious for additional EILO diagnosis. The DI showed higher values in V1 in EILO compared to EIB patients. A score≥30 can predict a positive CLE reaction. CONCLUSION: ACT and DI are valuable tools in preselecting CLE candidates to assure timely diagnostic despite limited diagnostic capabilities.

MicroRNA Profiling of the Inflammatory Response after Early and Late Asthmatic Reaction.

A high proportion of house dust mite (HDM)-allergic asthmatics suffer from both an early asthmatic reaction (EAR) and a late asthmatic reaction (LAR) which follows it. In these patients, allergic inflammation is more relevant. MiRNAs have been shown to play an important role in the regulation of asthma's pathology. The aim of this study was to analyze the miRNA profile in patients with mild asthma and an HDM allergy after bronchial allergen provocation (BAP). Seventeen patients with EAR/no LAR and 17 patients with EAR plus LAR, determined by a significant fall in FEV1 after BAP, were differentially analyzed. As expected, patients with EAR plus LAR showed a more pronounced allergic inflammation and FEV1 delta drop after 24 h. NGS-miRNA analysis identified the down-regulation of miR-15a-5p, miR-15b-5p, and miR-374a-5p after BAP with the highest significance in patients with EAR plus LAR, which were negatively correlated with eNO and the maximum decrease in FEV1. These miRNAs have shared targets like CCND1, VEGFA, and GSK3B, which are known to be involved in airway remodeling, basement membrane thickening, and Extracellular Matrix deposition. NGS-profiling identified miRNAs involved in the inflammatory response after BAP with HDM extract, which might be useful to predict a LAR.

Severe bronchial hyperresponsiveness along with house dust mite allergy indicates persistence of asthma in young children.

BACKGROUND: Significant risk factors for persistence of asthma later in life are family history of allergies, early allergic sensitization and bronchial hyperresponsiveness (BHR). The evolution of BHR in young children without allergic sensitization and with house dust mite allergy (HDM) was investigated. METHODS: In this retrospective analysis, electronic charts of 4850 young children with asthma and wheezy bronchitis between 2005 and 2018 were reviewed in order to study all patients ≤6 years with BHR assessed by methacholine provocation tests (MCT) at least once (n = 1175). Patients with more than two follow-up measurements were divided in group 1 (no allergic sensitization; n = 110) and group 2 (HDM allergy; n = 88). Additionally, skin prick test, exhaled nitrite oxide (eNO), and asthma treatment were analyzed. RESULTS: Forty-seven patients of group 1 aged median 4.3 years and 48 patients of group 2 aged median 4.7 years showed initially severe BHR <0.1 mg. At follow-up, patients with HDM were more likely to show persistence of severe BHR than non-sensitized patients (severe BHR group 1: n = 5 (10.6%) vs. group 2: n = 21 (43.8%), p < .001). In addition, 89.4% of group 1 had mild to moderate or no BHR, compared to only 56.2% of group 2. There was a significant difference in eN0 (median group 1: 9 ppb vs. group 2: 26 ppb, p < .001), at last follow-up. Age, sex, and asthma therapy had no effect on BHR. CONCLUSION: In young children without sensitization BHR normalizes, whereas HDM allergy indicates a persistence of asthma beyond infancy.

Methacholine and FeNO Measurement in Patients with Habit Cough.

BACKGROUND: Chronic cough is one of the most common symptoms in childhood. Making a definite diagnosis is a challenge for all pediatricians especially in patients when cough is without an organic cause like in habit cough. PATIENTS AND METHODS: In this retrospective analysis, all electronic outpatient charts of the Division of Allergology and Pneumology, between January 1, 2010 and December 31, 2019 were reviewed in order to study all children with potential habit cough. All children underwent the following diagnostic algorithms, skin prick test (SPT), measurement of fractional exhaled nitric oxide (FeNO), spirometry and methacholine challenge test (MCT). The value of a normal MCT and FeNO measurement for diagnosing habit cough was investigated. RESULTS: The chart review revealed 482 patients with chronic cough>4 weeks. Of these, 99 (20.5%) with suspected habit cough were collected. 13 patients had to be excluded for other diagnosis and a complete data set was available in 55 patients. 33 (60.0%) of 55 patients were SPT negative and 22 (40.0%) had sensitization to common allergens. Five patients had elevated FeNO≥20 ppb and three showed severe bronchial hyperresponsiveness<0.1 mg methacholine, challenging the diagnosis of habit cough. CONCLUSION: A normal FeNO and MCT can help confirm the clinical diagnosis of habit cough. However, in patients with positive MCT and/or elevated FeNO habit cough can be present. Especially in patients with elevated FeNO and severe BHR cough variant asthma and eosinophilic bronchitis have to be ruled out.

Antibiotic Use in Paediatric Patients Hospitalized with Acute Severe Asthma.

BACKGROUND: Antibiotic use during asthma exacerbations in paediatric patients is not routinely recommended but common practise in out-patient and in-patient settings. Objective of this study was to analyse frequency of antibiotic use during acute severe asthma exacerbations, antibiotic classes utilized and clinical decision-making. METHODS: All in-patient admissions over 10 years in a single German Children's University hospital due to acute severe asthma were included in this retrospective analysis. Age, length of stay, oxygen supplementation, treatment, laboratory parameters and chest x-rays of all patients ranging from 1 to 17 years were analysed. RESULTS: 580 hospital admissions were included in this study. Overall antibiotic use was high but decreased with age (1-5 years 69,6%, 6-11 years 57,6% and 12-17 years 39,7%, p<0.001). Analysis of antibiotic treatment without clear indication showed a consistently lower treatment rate of 28.3%, with macrolides being the most common antibiotic class. Younger age significantly decreased, whereas, increase of CrP value, use of oxygen supplementation and concomitant fever all significantly increased the odds ratio (OR 0.967; 4.366, 2.472 and 2.011 respectively) of receiving antibiotic treatment without clear indication. CONCLUSION: Antibiotic treatment without clear indication during acute severe asthma is common in this German single-centre cohort. Clinical parameters of more severe disease affect clinician's decision to administer antibiotics despite evidence of bacterial infection or improved outcome.

Evaluating Children and Adolescents with Suspected Exercise Induced Asthma: Real Life Data.

OBJECTIVE: Exercise-induced bronchoconstriction (EIB) occurs frequently in children and adolescents and may be a sign of insufficient asthma control. EIB is often evaluated by respiratory symptoms, spirometry, eNO measurement and methacholine testing (MCT) instead of time consuming exercise test. Aim of this study was to analyse the amount of patients for which an exercise challenge in a cold chamber (ECC) was needed for a clear EIB diagnosis, to characterize EIB phenotypes and the incidence of exercise induced laryngeal obstruction (EILO) in a large cohort of patients with EIB. METHODS: A retrospective analysis was performed in 595 children and adolescents (mean age 12.1 years) with suspected EIB from January 2014 to December 2018. Complete data sets of skin prick test, spirometry, eNO and MCT were available from 336 patients. RESULTS: An ECC to confirm the EIB diagnosis was performed in 125 (37.2%) of patients. Three EIB phenotypes were detected: group 1: EIB without allergic sensitization (n=159); group 2: EIB with other than house dust mite (HDM) sensitization (n=87) and group 3: EIB with HDM sensitization (n=90). MCT and eNO showed significant differences between the subgroups: An eNO>46 ppb and/or a MCT<0.1 mg was found in 23.9% vs. 50.6% vs. 57.8% in group 1-3, respectively. Significantly more patients suffered from EILO in group 1 compared to group 2 and 3 (n=13 vs. n=1). CONCLUSION: EIB without sensitization is as often as EIB with sensitization. In patients without sensitization, EILO has to be considered as a possible cause of symptoms during exercise.

Ligelizumab treatment for severe asthma: learnings from the clinical development programme.

OBJECTIVE: Ligelizumab is a humanised IgG1 anti-IgE antibody that binds IgE with higher affinity than omalizumab. Ligelizumab had greater efficacy than omalizumab on inhaled and skin allergen provocation responses in mild allergic asthma. This multi-centre, randomised, double-blind study was designed to test ligelizumab in severe asthma patients not adequately controlled with high-dose inhaled corticoids plus long-acting ß2-agonist. METHODS: Patients received 16 weeks ligelizumab (240 mg q2w), omalizumab or placebo subcutaneously, and ACQ-7 was measured as primary outcome at Week 16. In addition, the study generated dose-ranging data of ligelizumab and safety data. RESULTS: A total of 471 patients, age 47.4 ± 13.36 years, were included in the study. Treatment with ligelizumab did not significantly improve asthma control (ACQ-7) and exacerbation rates compared to omalizumab and placebo. Therefore, primary and secondary objectives of the study were not met. The compound was well tolerated, and the safety profile showed no new safety findings. Pharmacokinetic data demonstrated faster clearance and lower serum concentrations of ligelizumab than historical omalizumab data, and exploratory in vitro data showed differential IgE blocking properties relative to FcεRI and FcεRII/CD23 between the two compounds. CONCLUSION: Ligelizumab failed to demonstrate superiority over placebo or omalizumab. Although ligelizumab is more potent than omalizumab at inhibiting IgE binding to the high-affinity FcεRI, there is differential IgE blocking properties relative to FcεRI and FcεRII/CD23 between the two compounds. Therefore, the data suggest that different anti-IgE antibodies might be selectively efficacious for different IgE-mediated diseases.

Tiotropium as an Add-on Treatment Option for Severe Uncontrolled Asthma in Preschool Patients.

BACKGROUND: Toddlers with asthma suffer disproportionally more than school-aged children from exacerbations with emergency visits and hospital admissions despite inhaled corticosteroid (ICS) treatment. A recent trial for children ≤5 years showed tolerability of tiotropium and potential to reduce asthma-related events. METHODS: We conducted a retrospective analysis of electronic outpatient records (2017‒2019) of children <6 years treated with ICS plus long-acting ß2-agonists (LABAs) plus tiotropium as an add-on for uncontrolled severe asthma. The primary endpoint was a comparison of systemic corticosteroid (SCS) prescriptions 6 months before and after ICS/LABA/tiotropium start. Secondary endpoints included physician visits, hospitalisations and antibiotic prescriptions. We compared outcomes with children without asthma matched for age, sex, season and screening date. RESULTS: Compared with a mean 2.42 (95% CI: 1.75, 3.36) SCS courses per patient within 6 months prior to ICS/LABA/tiotropium, 0.74 (95% CI: 0.25, 1.08) SCS courses per patient were prescribed within 6 months after starting ICS/LABA/tiotropium (P<0.001). Physician visits dropped from 9.23 (95% CI: 7.15, 12.72) to 5.76 (95% CI: 3.10, 7.70) per patient (P<0.01). Nineteen hospitalisations were recorded 6 months before ICS/LABA/tiotropium compared with one hospitalisation after (P<0.01). A mean 1.79 antibiotic courses (95% CI: 1.22, 2.23) per patient were prescribed before ICS/LABA/tiotropium compared with 0.74 (95% CI: 0.22, 1.00) after ICS/LABA/tiotropium (P<0.001). Hospitalisation rates for patients at observation end were not statistically different from healthy controls before/after matching. INTERPRETATION: Our retrospective study showed that adding tiotropium to ICS/LABA is a new treatment option for patients with severe preschool asthma; however, larger confirmatory studies are needed.

Hospitalization, Asthma Phenotypes, and Readmission Rates in Pre-school Asthma.

Objective: Children with pre-school asthma suffer disproportionally more often from severe asthma exacerbations with emergency visits and hospital admissions compared to school children. Despite this high disease burden, there are only a few reports looking at this particular severe asthma cohort. Similarly, there is little real-life research on the distribution of asthma phenotypes and personalized treatment at discharge in this age group. Patients and Methods: Retrospective analysis of the electronic charts of all children aged 1-5 years with asthma hospitalizations (ICD J45) at the Frankfurt University between 2008 and 2017. An acute severe asthma exacerbation was defined as dyspnea, oxygen demand, and/or systemic steroid therapy. Age, gender, duration of hospitalization, asthma phenotype, treatment, and readmission rate were analyzed. Results: Of 572 patients, 205 met the definition of acute severe asthma. The phenotypic characterization showed 56.1% had allergic asthma, 15.2% eosinophilic asthma and 28.7% non-allergic asthma. Of these patients, 71.7% were discharged with inhaled corticosteroids (ICS) or ICS + long-acting-beta-agonists (LABA), 15.1% with leukotriene antagonists (LTRA) and 7.3% salbutamol on demand. The rate of emergency presentations (emergency department and readmission) within 12 months after discharge was high (n = 42; 20.5%). No phenotype tailored treatment was detectable. Neither the number of eosinophils (>300/µl) nor the treatment at discharge had an effect on emergency visits and readmission rate. Conclusion: Despite protective therapy with ICS, ICS + LABA, or LTRA, the readmission rate was high. Thus, current care and treatment strategies should be reevaluated continuously, in order to better control asthma in pre-school children and prevent hospitalization.

Immune competence and respiratory symptoms in patients with ataxia telangiectasia: A prospective follow-up study.

Ataxia telangiectasia is a multi-system disorder characterized by progressive cerebellar ataxia, malignancies, chronic pulmonary disease and immunodeficiency. The aim of our study was to determine the immune competence and prevalence of respiratory infections and/or chronic cough in classical A-T patients compared to age-matched healthy controls. STUDY DESIGN: We recruited 20 classical A-T not treated by immunoglobulins and 21 healthy age-matched control patients. The caregivers were advised to keep a daily diary with the following items (daytime and nighttime cough, runny nose, fever), number of cold episodes, number of antibiotic treatments. RESULTS: Patients with A-T showed significant differences compared to healthy controls in symptom score, daytime and nighttime cough, days with symptoms and missed days in kindergarten/school. Severe infections with hospitalization occurred rarely. Respiratory symptoms did not correlate with immunoglobulin levels in A-T patients. CONCLUSIONS: Mild symptoms like chronic cough were present in A-T patients, possibly indicating ongoing silent crippling disease.

Single-photon emission computed tomography/computed tomography imaging of RAGE in smoking-induced lung injury.

BACKGROUND: Expression of the Receptor for Advanced Glycation Endproducts (RAGE) initiates pro-inflammatory pathways resulting in lung destruction. We hypothesized that RAGE directed imaging demonstrates increased lung uptake in smoke-exposure. METHODS: After exposure to room air or to cigarette smoke for 4-weeks or 16-weeks, rabbits were injected with 99mTc-anti-RAGE F(ab')2 and underwent Single-Photon Emission Computed Tomography/Computed Tomography (SPECT/CT) imaging. Lung radiotracer uptake was calculated as percent injected dose (%ID). Lungs were dissected for gamma well counting and histological analysis. RESULTS: 99mTc-anti-RAGE F(ab')2 SPECT/CT imaging demonstrated increased lung expression of RAGE with smoke exposure compared to room air control at 4-weeks: Room air right (R) 0.75 ± 0.38%ID, left (L) 0.62 ± 0.32%ID vs. Smoke exposed R 0.17 ± 0.03, L 0.17 ± 0.02%ID (p = 0.02 and 0.028, respectively). By 16-weeks of smoke exposure, the uptake decreased to 0.19 ± 0.05%ID R and 0.17 ± 0.05%ID L, significantly lower than 4-week imaging (p = 0.0076 and 0.0129 respectively). Staining for RAGE confirmed SPECT results, with the RAGE ligand HMGB1 upregulated in the macrophages of 4-week smoke-exposed rabbits. CONCLUSIONS: RAGE-directed imaging identified pulmonary RAGE expression acutely in vivo in an animal model of emphysema early after smoke exposure, with diminution over time. These studies document the extent and time course of RAGE expression under smoke exposure conditions and could be utilized for disease monitoring and examining response to future RAGE-targeted therapies.

A critical role for ABC transporters in persistent lung inflammation in the development of emphysema after smoke exposure.

Macrophage infiltration is common to both emphysema and atherosclerosis, and cigarette smoke down-regulates the macrophage cholesterol efflux transporter ATP binding cassette (ABC)A1. This decreased cholesterol efflux results in lipid-laden macrophages. We hypothesize that cigarette smoke adversely affects cholesterol transport via an ABCA1-dependent mechanism in macrophages, enhancing TLR4/myeloid differentiation primary response gene 88 (Myd88) signaling and resulting in matrix metalloproteinase (MMP) up-regulation and exacerbation of pulmonary inflammation. ABCA1 is significantly down-regulated in the lung upon smoke exposure conditions. Macrophages exposed to cigarette smoke in vivo and in vitro exhibit impaired cholesterol efflux correlating with significantly decreased ABCA1 expression, up-regulation of the TLR4/Myd88 pathway, and downstream MMP-9 and MMP-13 expression. Treatment with liver X receptor (LXR) agonist restores ABCA1 expression after short-term smoke exposure and attenuates the inflammatory response; after long-term smoke exposure, there is also attenuated physiologic and morphologic changes of emphysema. In vitro, treatment with LXR agonist decreases macrophage inflammatory activation in wild-type but not ABCA1 knockout mice, suggesting an ABCA1-dependent mechanism of action. These studies demonstrate an important association between cigarette smoke exposure and cholesterol-mediated pathways in the macrophage inflammatory response. Modulation of these pathways through manipulation of ABCA1 activity effectively blocks cigarette smoke-induced inflammation and provides a potential novel therapeutic approach for the treatment of chronic obstructive pulmonary disease.-Sonett, J., Goldklang, M., Sklepkiewicz, P., Gerber, A., Trischler, J., Zelonina, T., Westerterp, M., Lemaître, V., Okada, V., D'Armiento, J. A critical role for ABC transporters in persistent lung inflammation in the development of emphysema after smoke exposure.

Biased Generation and In Situ Activation of Lung Tissue-Resident Memory CD4 T Cells in the Pathogenesis of Allergic Asthma.

Asthma is a chronic inflammatory disease mediated by allergen-specific CD4 T cells that promote lung inflammation through recruitment of cellular effectors into the lung. A subset of lung T cells can persist as tissue-resident memory T cells (TRMs) following infection and allergen induction, although the generation and role of TRM in asthma persistence and pathogenesis remain unclear. In this study, we used a mouse model of chronic exposure to intranasal house dust mite (HDM) extract to dissect how lung TRMs are generated and function in the persistence and pathogenesis of allergic airway disease. We demonstrate that both CD4+ and CD8+ T cells infiltrate into the lung tissue during acute HDM exposure; however, only CD4+ TRMs, and not CD8+ TRMs, persist long term following cessation of HDM administration. Lung CD4+ TRMs are localized around airways and are rapidly reactivated upon allergen re-exposure accompanied by the rapid induction of airway hyperresponsiveness independent of circulating T cells. Lung CD4+ TRM activation to HDM challenge is also accompanied by increased recruitment and activation of dendritic cells in the lungs. Our results indicate that lung CD4+ TRMs can perpetuate allergen-specific sensitization and direct early inflammatory signals that promote rapid lung pathology, suggesting that targeting lung CD4+ TRMs could have therapeutic benefit in alleviating recurrent asthma episodes.

Maternal smoke exposure decreases mesenchymal proliferation and modulates Rho-GTPase-dependent actin cytoskeletal signaling in fetal lungs.

The present study tested the hypothesis that maternal smoke exposure results in fetal lung growth retardation due to dysregulation in various signaling pathways, including the Wnt (wingless-related integration site)/ß-catenin pathway. Pregnant female C57BL/6J mice were exposed to cigarette smoke (100-150 mg/m3) or room air, and offspring were humanely killed on 12.5, 14.5, 16.5, and 18.5 d post coitum (dpc). We assessed lung stereology with Cavalieri estimation; apoptosis with proliferating cell nuclear antigen, TUNEL, and caspase assays; and gene expression with quantitative PCR (qPCR) and RNA sequencing on lung epithelium and mesenchyme retrieved by laser capture microdissection. Results demonstrated a significant decrease in body weight and lung volume of smoke-exposed embryos. At 16.5 dpc, the reduction in lung volume was due to loss of lung mesenchymal tissue correlating with a decrease in cell proliferation (n = 10; air: 61.65% vs. smoke: 44.21%, P < 0.05). RNA sequence analysis demonstrated an alteration in the Wnt pathway, and qPCR confirmed an increased expression of secreted frizzled-related protein 1 (sFRP-1) [n = 12; relative quantification (RQ) 1 vs. 2.33, P < 0.05] and down-regulation of Cyclin D1 (n = 7; RQ 1 vs. 0.61, P < 0.05) in mesenchymal tissue. Furthermore, genome expression studies revealed a smoke-induced up-regulation of Rho-GTPase-dependent actin cytoskeletal signaling that can lead to loss of tissue integrity.-Unachukwu, U., Trischler, J., Goldklang, M., Xiao, R., D'Armiento, J. Maternal smoke exposure decreases mesenchymal proliferation and modulates Rho-GTPase-dependent actin cytoskeletal signaling in fetal lungs.

Single-Photon Emission Computed Tomography/Computed Tomography Imaging in a Rabbit Model of Emphysema Reveals Ongoing Apoptosis In Vivo.

Evaluation of lung disease is limited by the inability to visualize ongoing pathological processes. Molecular imaging that targets cellular processes related to disease pathogenesis has the potential to assess disease activity over time to allow intervention before lung destruction. Because apoptosis is a critical component of lung damage in emphysema, a functional imaging approach was taken to determine if targeting apoptosis in a smoke exposure model would allow the quantification of early lung damage in vivo. Rabbits were exposed to cigarette smoke for 4 or 16 weeks and underwent single-photon emission computed tomography/computed tomography scanning using technetium-99m-rhAnnexin V-128. Imaging results were correlated with ex vivo tissue analysis to validate the presence of lung destruction and apoptosis. Lung computed tomography scans of long-term smoke-exposed rabbits exhibit anatomical similarities to human emphysema, with increased lung volumes compared with controls. Morphometry on lung tissue confirmed increased mean linear intercept and destructive index at 16 weeks of smoke exposure and compliance measurements documented physiological changes of emphysema. Tissue and lavage analysis displayed the hallmarks of smoke exposure, including increased tissue cellularity and protease activity. Technetium-99m-rhAnnexin V-128 single-photon emission computed tomography signal was increased after smoke exposure at 4 and 16 weeks, with confirmation of increased apoptosis through terminal deoxynucleotidyl transferase dUTP nick end labeling staining and increased tissue neutral sphingomyelinase activity in the tissue. These studies not only describe a novel emphysema model for use with future therapeutic applications, but, most importantly, also characterize a promising imaging modality that identifies ongoing destructive cellular processes within the lung.

Immune Modulation of the T Cell Response in Asthma through Wnt10b.

Asthma is a chronic inflammatory disease, which is characterized by activation of CD4(+) T helper 2 cells orchestrating an allergic airway response. Whereas the role of Wnt family members in regulating T cell maintenance and maturation is established, their contribution to T cell activation in allergic asthma is not known. We hypothesized that Wnt10b plays a role in the modulation of the allergic airway response and affects T cell activation and polarization. Using an in vivo house dust mite asthma model, Wnt10b-deficient (Wnt10b(-/-)) mice were allergen-sensitized and inflammation, as well as T cell activation, was studied in vivo and in vitro. Wnt10b(-/-) mice exhibited an augmented inflammatory phenotype with an increase in eosinophils in the bronchoalveolar lavage and IL-4 and IL-13 in the lungs when compared with wild-type mice. In vitro studies confirmed an increased T helper type 2 polarization and increased T cell activation of Wnt10b(-/-) cells. Accordingly, the percentage of naive T cells was elevated by the addition of recombinant Wnt10b protein. Finally, Wnt10b(-/-) mice exhibited an increase in the percentage of effector T cells in the lungs after house dust mite sensitization, which indicated a heightened activation state, measured by an increased percentage of CD69(hi)CD11a(hi) cells. These findings suggest that Wnt10b plays an important role in regulating asthmatic airway inflammation through modification of the T cell response and is a prospective target in the disease process.