Human iPSC-Based Model of COPD to Investigate Disease Mechanisms, Predict SARS-COV-2 Outcome, and Test Preventive Immunotherapy.

Chronic inflammation and dysregulated repair mechanisms after epithelial damage have been implicated in chronic obstructive pulmonary disease (COPD). However, the lack of ex vivo-models that accurately reflect multicellular lung tissue hinders our understanding of epithelial-mesenchymal interactions in COPD. Through a combination of transcriptomic and proteomic approaches applied to a sophisticated in vitro iPSC-alveolosphere with fibroblasts model, epithelial-mesenchymal crosstalk was explored in COPD and following SARS-CoV-2 infection. These experiments profiled dynamic changes at single-cell level of the SARS-CoV-2-infected alveolar niche that unveiled the complexity of aberrant inflammatory responses, mitochondrial dysfunction, and cell death in COPD, which provides deeper insights into the accentuated tissue damage/inflammation/remodeling observed in patients with SARS-CoV-2 infection. Importantly, this 3D system allowed for the evaluation of ACE2-neutralizing antibodies and confirmed the potency of this therapy to prevent SARS-CoV-2 infection in the alveolar niche. Thus, iPSC-alveolosphere cultured with fibroblasts provides a promising model to investigate disease-specific mechanisms and to develop novel therapeutics.

IL-13-induced STAT3-dependent signaling networks regulate esophageal epithelial proliferation in eosinophilic esophagitis.

BACKGROUND: Basal zone hyperplasia (BZH) and dilated intercellular spaces (DISs) are thought to contribute to the clinical manifestations of eosinophilic esophagitis (EoE); however, the molecular pathways that drive BZH remain largely unexplored. OBJECTIVE: We sought to define the role of IL-13-induced transcriptional programs in esophageal epithelial proliferation in EoE. METHODS: We performed RNA sequencing, bioinformatics, Western blot, reverse transcriptase quantitative PCR, and histologic analyses on esophageal biopsies from healthy control and patients with EoE, primary esophageal cells derived from patients with EoE, and IL-13-stimulated esophageal epithelial keratinocytes grown at the air-liquid interface (EPC2-ALI). Genetic (shRNA) and pharmacologic (proteolysis-targeting chimera degrader) approaches and in vivo model of IL-13-induced esophageal epithelial remodeling (Krt5-rtTA x tetO-IL-13Tg) were used to define the role of signal transducer and activator of transcription 3 (STAT3) and STAT6 and secreted frizzled-related protein 1 (SFRP1) in esophageal epithelial proliferation. RESULTS: RNA-sequencing analysis of esophageal biopsies (healthy control vs EoE) and EPC2-ALI revealed 82 common differentially expressed genes that were enriched for putative STAT3 target genes. In vitro and in vivo analyses revealed a link between IL-13-induced STAT3 and STAT6 phosphorylation, SFRP1 mRNA expression, and esophageal epithelial proliferation. In vitro studies showed that IL-13-induced esophageal epithelial proliferation was STAT3-dependent and regulated by the STAT3 target SFRP1. SFRP1 mRNA is increased in esophageal biopsies from patients with active EoE compared with healthy controls or patients in remission and identifies an esophageal suprabasal epithelial cell subpopulation that uniquely expressed the core EoE proinflammatory transcriptome genes (CCL26, ALOX15, CAPN14, ANO1, and TNFAIP6). CONCLUSIONS: These studies identify SFRP1 as a key regulator of IL-13-induced and STAT3-dependent esophageal proliferation and BZH in EoE and link SFRP1+ esophageal epithelial cells with the proinflammatory and epithelial remodeling response in EoE.

Localized strain characterization of cardiomyopathy in Duchenne muscular dystrophy using novel 4D kinematic analysis of cine cardiovascular magnetic resonance.

BACKGROUND: Cardiomyopathy (CMP) is the most common cause of mortality in Duchenne muscular dystrophy (DMD), though the age of onset and clinical progression vary. We applied a novel 4D (3D + time) strain analysis method using cine cardiovascular magnetic resonance (CMR) imaging data to determine if localized strain metrics derived from 4D image analysis would be sensitive and specific for characterizing DMD CMP. METHODS: We analyzed short-axis cine CMR image stacks from 43 DMD patients (median age: 12.23 yrs [10.6-16.5]; [interquartile range]) and 25 male healthy controls (median age: 16.2 yrs [13.3-20.7]). A subset of 25 male DMD patients age-matched to the controls (median age: 15.7 yrs [14.0-17.8]) was used for comparative metrics. CMR images were compiled into 4D sequences for feature-tracking strain analysis using custom-built software. Unpaired t-test and receiver operator characteristic area under the curve (AUC) analysis were used to determine statistical significance. Spearman's rho was used to determine correlation. RESULTS: DMD patients had a range of CMP severity: 15 (35% of total) had left ventricular ejection fraction (LVEF) > 55% with no findings of myocardial late gadolinium enhancement (LGE), 15 (35%) had findings of LGE with LVEF > 55% and 13 (30%) had LGE with LVEF < 55%. The magnitude of the peak basal circumferential strain, basal radial strain, and basal surface area strain were all significantly decreased in DMD patients relative to healthy controls (p < 0.001) with AUC values of 0.80, 0.89, and 0.84 respectively for peak strain and 0.96, 0.91, and 0.98 respectively for systolic strain rate. Peak basal radial strain, basal radial systolic strain rate, and basal circumferential systolic strain rate magnitude values were also significantly decreased in mild CMP (No LGE, LVEF > 55%) compared to a healthy control group (p < 0.001 for all). Surface area strain significantly correlated with LVEF and extracellular volume (ECV) respectively in the basal (rho = - 0.45, 0.40), mid (rho = - 0.46, 0.46), and apical (rho = - 0.42, 0.47) regions. CONCLUSION: Strain analysis of 3D cine CMR images in DMD CMP patients generates localized kinematic parameters that strongly differentiate disease from control and correlate with LVEF and ECV.

Youth Anxiety: The Moderating Effects of Accommodation and Emotional Warmth.

Parental accommodation (i.e., modifying behavior to reduce child distress) is among the most empirically supported anxiety enhancing parenting practices; while emotional warmth (i.e., support and affection) has demonstrated a less clear link to anxiety. The current study aims to explore the interactive nature of emotional warmth within the context of accommodation. We hypothesized that accommodation would moderate the relationship between emotional warmth and anxiety. The sample included parents of youth (N = 526) ages 7-17. A simple moderation analysis was conducted. Accommodation significantly moderated the relationship [B = 0.03, C.I. (0.01, 0.05), p = 0.01]. Additional variance was accounted for by adding the interaction term to the model (R2 = 0.47, p < 0.001). At high levels of accommodation, emotional warmth significantly predicted child anxiety symptoms. This study affirms that emotional warmth is significantly related to anxiety in the context of high accommodation. Future work ought to build upon these findings to explore these relationships. Limitations of the study include sampling and parent-report data.

Sleep Disturbances in Pediatric Body-Focused Repetitive Behaviors: A Preliminary Investigation.

Body-focused repetitive behaviors (BFRBs) are repeated actions to one's body resulting in physical damage. Limited research has examined sleep, a known factor in psychological health, within the context of pediatric BFRBs. The current study sought to explore the connection between disordered sleep and BFRBs in a community sample. Aim 1 of the study was to determine the predictive power of group membership [control group (no BFRB symptoms reported), subthreshold BFRB group (mild BFRB symptoms reported; severity score of 2 or less out of 9), and those with symptoms characteristic of BFRBs (more than mild BFRB symptoms reported; severity score of 3 or higher out of 9)] for level of sleep disturbance. A hierarchical regression revealed that there was a significant effect of group membership after controlling for anxiety (F (3, 410) = 152.976, p < .001). Aim 2 of the study was to test whether there was a relationship between sleep disturbance and BFRB severity. The hierarchical regression revealed that at Step 1, anxiety accounted for 23.1% of the variance in BFRB severity (ß = 0.48, t = 8.87, p < 0.001). At Step 2, sleep disturbance total score accounted for an additional 7.2% of the variance, suggesting this variable makes a unique contribution to overall BFRB severity (SDSC: ß = 0.40, t = 5.18, p < 0.001). The findings of this study suggest that sleep could be a clinical factor to consider when conceptualizing a child with BFRBs.

Quantification of murine myocardial infarct size using 2-D and 4-D high-frequency ultrasound.

Ischemic heart disease is the leading cause of death in the United States, Canada, and worldwide. Severe disease is characterized by coronary artery occlusion, loss of blood flow to the myocardium, and necrosis of tissue, with subsequent remodeling of the heart wall, including fibrotic scarring. The current study aims to demonstrate the efficacy of quantitating infarct size via two-dimensional (2-D) echocardiographic akinetic length and four-dimensional (4-D) echocardiographic infarct volume and surface area as in vivo analysis techniques. We further describe and evaluate a new surface area strain analysis technique for estimating myocardial infarction (MI) size after ischemic injury. Experimental MI was induced in mice via left coronary artery ligation. Ejection fraction and infarct size were measured through 2-D and 4-D echocardiography. Infarct size established via histology was compared with ultrasound-based metrics via linear regression analysis. Two-dimensional echocardiographic akinetic length (r = 0.76, P = 0.03), 4-D echocardiographic infarct volume (r = 0.85, P = 0.008), and surface area (r = 0.90, P = 0.002) correlate well with histology. Although both 2-D and 4-D echocardiography were reliable measurement techniques to assess infarct, 4-D analysis is superior in assessing asymmetry of the left ventricle and the infarct. Strain analysis performed on 4-D data also provides additional infarct sizing techniques, which correlate with histology (surface strain: r = 0.94, P < 0.001, transmural thickness: r = 0.76, P = 0.001). Two-dimensional echocardiographic akinetic length, 4-D echocardiography ultrasound, and strain provide effective in vivo methods for measuring fibrotic scarring after MI.NEW & NOTEWORTHY Our study supports that both 2-D and 4-D echocardiographic analysis techniques are reliable in quantifying infarct size though 4-D ultrasound provides a more holistic image of LV function and structure, especially after myocardial infarction. Furthermore, 4-D strain analysis correctly identifies infarct size and regional LV dysfunction after MI. Therefore, these techniques can improve functional insight into the impact of pharmacological interventions on the pathophysiology of cardiac disease.

Psychometric Properties of a Measure Assessing Anxiogenic Parenting Practices in Food Allergy.

AIM: Pediatric food allergy represents a significant public health burden. In order to avoid allergen consumption, adequate management requires daily vigilance and involvement from parents, frequently leading to increased parental anxiety. While specific anxiogenic parenting practices (i.e., parenting behaviors which may aid in the development and/or exacerbation of childhood anxiety) have been documented within this population, to this point, these behaviors have not been systematically measured. OBJECTIVES: The current study aimed to develop and examine a parent-report scale designed to measure anxiogenic parenting behaviors related to food allergy. METHODS: Participants included 177 parents of children with food allergy recruited online using Amazon Mechanical Turk (mTurk). An exploratory factor analysis was conducted to determine the factor structure of the newly developed scale. Subsequently, psychometric properties (e.g., construct validity) were examined via correlational analyses. RESULTS: Results indicated a 24-item, 3 factor (Factor 1: Involvement in Food Allergy; Factor 2: Worry about Food Allergy; Factor 3: Autonomy Promotion) scale, which accounted for 53.11% of the total variance. The Kaiser-Meyer-Olkin measure was acceptable, KMO = 0.872 and Bartlett's test of sphericity indicated sufficient correlations between items (χ2(378)=2568.95). All subscales demonstrated strong internal consistency (Involvement in Food Allergy: α = .880; Worry about Food Allergy: α = .892; Autonomy Promotion α = .796) as well as convergent and discriminant validity. CONCLUSIONS: Results support the overall psychometric properties of the scale. Interpretations, limitations, and future directions are discussed.

Persistent Nitrate in Alpine Waters with Changing Atmospheric Deposition and Warming Trends.

Nitrate concentrations in high-elevation lakes of the Colorado Front Range remain elevated despite declining trends in atmospherically deposited nitrate since 2000. The current source of this elevated nitrate in surface waters remains elusive, given shifts in additional nitrogen sources via glacial inputs and atmospheric ammonium deposition. We present the complete isotopic composition of nitrate (δ15N, δ18O, and Δ17O) from a suite of nitrate-bearing source waters collected during the summers of 2017-2018 from two alpine ecosystems to constrain the provenance of elevated nitrate in surface waters during the summer open-water season. The results indicate a consistent contribution of uncycled atmospheric nitrate throughout the summer (13-23%) to alpine lakes, despite seasonal changes in source water inputs. The balance of nitrate (as high as 87% in late summer) is likely from nitrate production within the catchment via nitrification of reduced nitrogen sources (e.g., thawed soil organic matter and ammonium deposition) and released with rock glacier meltwater. The role of microbially produced nitrate has become increasingly important over time based on historical surface water samples from the mid-90s to present, a trend coincident with increasing ammonium deposition to alpine systems.

Opposing roles for TGFß- and BMP-signaling during nascent alveolar differentiation in the developing human lung.

Alveolar type 2 (AT2) cells function as stem cells in the adult lung and aid in repair after injury. The current study aimed to understand the signaling events that control differentiation of this therapeutically relevant cell type during human development. Using lung explant and organoid models, we identified opposing effects of TGFß- and BMP-signaling, where inhibition of TGFß- and activation of BMP-signaling in the context of high WNT- and FGF-signaling efficiently differentiated early lung progenitors into AT2-like cells in vitro. AT2-like cells differentiated in this manner exhibit surfactant processing and secretion capabilities, and long-term commitment to a mature AT2 phenotype when expanded in media optimized for primary AT2 culture. Comparing AT2-like cells differentiated with TGFß-inhibition and BMP-activation to alternative differentiation approaches revealed improved specificity to the AT2 lineage and reduced off-target cell types. These findings reveal opposing roles for TGFß- and BMP-signaling in AT2 differentiation and provide a new strategy to generate a therapeutically relevant cell type in vitro.

Opposing roles for TGFß- and BMP-signaling during nascent alveolar differentiation in the developing human lung.

Alveolar type 2 (AT2) cells function as stem cells in the adult lung and aid in repair after injury. The current study aimed to understand the signaling events that control differentiation of this therapeutically relevant cell type during human development. Using lung explant and organoid models, we identified opposing effects of TGFß- and BMP-signaling, where inhibition of TGFß- and activation of BMP-signaling in the context of high WNT- and FGF-signaling efficiently differentiated early lung progenitors into AT2-like cells in vitro . AT2-like cells differentiated in this manner exhibit surfactant processing and secretion capabilities, and long-term commitment to a mature AT2 phenotype when expanded in media optimized for primary AT2 culture. Comparing AT2-like cells differentiated with TGFß-inhibition and BMP-activation to alternative differentiation approaches revealed improved specificity to the AT2 lineage and reduced off-target cell types. These findings reveal opposing roles for TGFß- and BMP-signaling in AT2 differentiation and provide a new strategy to generate a therapeutically relevant cell type in vitro .

Extra-abdominal umbilical vein varix in a newborn.

Umbilical vein varices are rare fetal anomalies typically found intra-abdominally and identified on ultrasound prior to birth. Intra-abdominal umbilical vein varices account for approximately 4% of umbilical cord abnormalities and are thought to be a developmental abnormality rather than a congenital malformation. The umbilical vein varix anomaly been shown to be associated with a higher incidence of adverse perinatal outcomes and there is evidence of a relationship between this and chromosomal abnormalities. There have been few case reports of extra-abdominal varices. This case reviews a multiparous Hispanic female who delivered a baby with an extra-abdominal umbilical vein varix who was admitted to the neonatal intensive care unit but had an uncomplicated hospital course. The report reviews strategies for antenatal testing and surveillance of identified varices.

Automated, High-resolution Mobile Collection System for the Nitrogen Isotopic Analysis of NOx.

Nitrogen oxides (NOx = NO + NO2) are a family of atmospheric trace gases that have great impact on the environment. NOx concentrations directly influence the oxidizing capacity of the atmosphere through interactions with ozone and hydroxyl radicals. The main sink of NOx is the formation and deposition of nitric acid, a component of acid rain and a bioavailable nutrient. NOx is emitted from a mixture of natural and anthropogenic sources, which vary in space and time. The collocation of multiple sources and the short lifetime of NOx make it challenging to quantitatively constrain the influence of different emission sources and their impacts on the environment. Nitrogen isotopes of NOx have been suggested to vary amongst different sources, representing a potentially powerful tool to understand the sources and transport of NOx. However, previous methods of collecting atmospheric NOx integrate over long (week to month) time spans and are not validated for the efficient collection of NOx in relevant, diverse field conditions. We report on a new, highly efficient field-based system that collects atmospheric NOx for isotope analysis at a time resolution between 30 min and 2 hr. This method collects gaseous NOx in solution as nitrate with 100% efficiency under a variety of conditions. Protocols are presented for collecting air in urban settings under both stationary and mobile conditions. We detail the advantages and limitations of the method and demonstrate its application in the field. Data from several deployments are shown to 1) evaluate field-based collection efficiency by comparisons with in situ NOx concentration measurements, 2) test the stability of stored solutions before processing, 3) quantify in situ reproducibility in a variety of urban settings, and 4) demonstrate the range of N isotopes of NOx detected in ambient urban air and on heavily traveled roadways.

Presence and incidence of izumo antibodies in sera of immunoinfertile women and men.

PROBLEM: Sperm antigens that are relevant to fertilization and fertility, and are involved in human immunoinfertility, are being investigated in several laboratories. Izumo is a sperm-specific protein that has shown to be essential for sperm-egg fusion. The present study was conducted to examine the expression of Izumo protein in human sperm. Also, the presence and incidence of Izumo antibodies in sera of immunoinfertile women and men were investigated. METHOD OF STUDY: The expression of Izumo protein in human sperm was examined by the Western blot procedure. Sera from immunoinfertile women (n = 25) and fertile women (n = 23), as well as sera from immunoinfertile men (n = 20) and fertile men (n = 15), were collected and analyzed for immunoreactivity with Izumo peptides in the enzyme-linked immunosorbent assay (ELISA). Three epitopes of human Izumo, namely one epitope of Izumo1, and two epitopes of Izumo2 [Izumo2 (K-15), and Izumo2 (T-17)], respectively, were selected for the present study. RESULTS: In the Western blot procedure, the Izumo1 antibody recognized a specific protein band of ~39 kDa in human sperm extract. In ELISA, 56% of the immunoinfertile female sera reacted positively (≧3 S.D. units) with Izumo1, 40% with Izumo2 (K-15), and 20% with Izumo2 (T-17) peptide. None of the sera (0%) from fertile women reacted positively (≦3 S.D. units) with any of the three Izumo peptides. None of the sera from immunoinfertile (0%) or fertile men (0%) reacted positively with any of the three Izumo peptides. CONCLUSION: Our findings indicate that human sperm expresses Izumo protein, and the immunoinfertile female sera have circulating isoantibodies against this protein. This is the first study to our knowledge, which examined the presence and incidence of Izumo antibodies in female and male immunoinfertility. These findings may find clinical applications in specific diagnosis and treatment of infertility, and contraceptive vaccine development.