KF4 anti-CELA1 Antibody and Purified α1-Antitrypsin Have Similar but Not Additive Efficacy in Preventing Emphysema in Murine α1-Antitrypsin Deficiency.

Alpha-1 antitrypsin (AAT) deficiency is the most common genetic cause of emphysema. Chymotrypsin-like Elastase 1 (CELA1) is a serine protease neutralized by AAT and is important in emphysema progression. Cela1 -deficiency is protective in a murine models of AAT-deficient emphysema. KF4 anti-CELA1 antibody prevented emphysema in PPE and cigarette smoke models in wild type mice. We evaluated potential toxicities of KF4 and its ability to prevent emphysema in AAT deficiency. We found Cela1 protein expression in mouse lung, pancreas, small intestine, and spleen. In toxicity studies, mice treated with KF4 25 mg/kg weekly for four weeks showed an elevation in blood urea nitrogen and slower weight gain compared to lower doses or equivalent dose IgG. In histologic grading of tissue injury of the lung, kidney, liver, and heart, there was some evidence of liver injury with KF4 25 mg/kg, but in all tissues, injury was less than in control mice subjected to cecal ligation and puncture. In efficacy studies, KF4 doses as low as 0.5 mg/kg reduced the lung elastase activity of AAT -/- mice treated with 0.2 units of PPE. In this injury model, AAT -/- mice treated with KF4 1 mg/kg weekly, human purified AAT 60 mg/kg weekly, and combined KF4 and AAT treatment had less emphysema than mice treated with IgG 1 mg/kg weekly. However, the efficacy of KF4, AAT, or KF4 & AAT was similar. While KF4 might be an alternative to AAT replacement, combined KF4 and AAT replacement does not confer additional benefit.

Inflammatory and tissue injury marker dynamics in pediatric acute respiratory distress syndrome.

BACKGROUNDThe molecular signature of pediatric acute respiratory distress syndrome (ARDS) is poorly described, and the degree to which hyperinflammation or specific tissue injury contributes to outcomes is unknown. Therefore, we profiled inflammation and tissue injury dynamics over the first 7 days of ARDS, and associated specific biomarkers with mortality, persistent ARDS, and persistent multiple organ dysfunction syndrome (MODS).METHODSIn a single-center prospective cohort of intubated pediatric patients with ARDS, we collected plasma on days 0, 3, and 7. Nineteen biomarkers reflecting inflammation, tissue injury, and damage-associated molecular patterns (DAMPs) were measured. We assessed the relationship between biomarkers and trajectories with mortality, persistent ARDS, or persistent MODS using multivariable mixed effect models.RESULTSIn 279 patients (64 [23%] nonsurvivors), hyperinflammatory cytokines, tissue injury markers, and DAMPs were higher in nonsurvivors. Survivors and nonsurvivors showed different biomarker trajectories. IL-1α, soluble tumor necrosis factor receptor 1, angiopoietin 2 (ANG2), and surfactant protein D increased in nonsurvivors, while DAMPs remained persistently elevated. ANG2 and procollagen type III N-terminal peptide were associated with persistent ARDS, whereas multiple cytokines, tissue injury markers, and DAMPs were associated with persistent MODS. Corticosteroid use did not impact the association of biomarker levels or trajectory with mortality.CONCLUSIONSPediatric ARDS survivors and nonsurvivors had distinct biomarker trajectories, with cytokines, endothelial and alveolar epithelial injury, and DAMPs elevated in nonsurvivors. Mortality markers overlapped with markers associated with persistent MODS, rather than persistent ARDS.FUNDINGNIH (K23HL-136688, R01-HL148054).

Biomarker-Based Risk Stratification Tool in Pediatric Acute Respiratory Distress Syndrome: Single-Center, Longitudinal Validation in a 2014-2019 Cohort.

OBJECTIVES: The Pediatric Acute Respiratory Distress Syndrome Biomarker Risk Model (PARDSEVERE) used age and three plasma biomarkers measured within 24 hours of pediatric acute respiratory distress syndrome (ARDS) onset to predict mortality in a pilot cohort of 152 patients. However, longitudinal performance of PARDSEVERE has not been evaluated, and it is unclear whether the risk model can be used to prognosticate after day 0. We, therefore, sought to determine the test characteristics of PARDSEVERE model and population over the first 7 days after ARDS onset. DESIGN: Secondary unplanned post hoc analysis of data from a prospective observational cohort study carried out 2014-2019. SETTING: University-affiliated PICU. PATIENTS: Mechanically ventilated children with ARDS. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Between July 2014 and December 2019, 279 patients with ARDS had plasma collected at day 0, 266 at day 3 (11 nonsurvivors, two discharged between days 0 and 3), and 207 at day 7 (27 nonsurvivors, 45 discharged between days 3 and 7). The actual prevalence of mortality on days 0, 3, and 7, was 23% (64/279), 14% (38/266), and 13% (27/207), respectively. The PARDSEVERE risk model for mortality on days 0, 3, and 7 had area under the receiver operating characteristic curve (AUROC [95% CI]) of 0.76 (0.69-0.82), 0.68 (0.60-0.76), and 0.74 (0.65-0.83), respectively. The AUROC data translate into prevalence thresholds for the PARDSEVERE model for mortality (i.e., using the sensitivity and specificity values) of 37%, 27%, and 24% on days 0, 3, and 7, respectively. Negative predictive value (NPV) was high throughout (0.87-0.90 for all three-time points). CONCLUSIONS: In this exploratory analysis of the PARDSEVERE model of mortality risk prediction in a population longitudinal series of data from days 0, 3, and 7 after ARDS diagnosis, the diagnostic performance is in the "acceptable" category. NPV was good. A major limitation is that actual mortality is far below the prevalence threshold for such testing. The model may, therefore, be more useful in cohorts with higher mortality rates (e.g., immunocompromised, other countries), and future enhancements to the model should be explored.

Anti-CELA1 antibody KF4 prevents emphysema by inhibiting stretch-mediated remodeling.

There are no therapies to prevent emphysema progression. Chymotrypsin-like elastase 1 (CELA1) is a serine protease that binds and cleaves lung elastin in a stretch-dependent manner and is required for emphysema in a murine antisense oligonucleotide model of α-1 antitrypsin (AAT) deficiency. This study tested whether CELA1 is important in strain-mediated lung matrix destruction in non-AAT-deficient emphysema and the efficacy of CELA1 neutralization. Airspace simplification was quantified after administration of tracheal porcine pancreatic elastase (PPE), after 8 months of cigarette smoke (CS) exposure, and in aging. In all 3 models, Cela1-/- mice had less emphysema and preserved lung elastin despite increased lung immune cells. A CELA1-neutralizing antibody was developed (KF4), and it inhibited stretch-inducible lung elastase in ex vivo mouse and human lung and immunoprecipitated CELA1 from human lung. In mice, systemically administered KF4 penetrated lung tissue in a dose-dependent manner and 5 mg/kg weekly prevented emphysema in the PPE model with both pre- and postinjury initiation and in the CS model. KF4 did not increase lung immune cells. CELA1-mediated lung matrix remodeling in response to strain is an important contributor to postnatal airspace simplification, and we believe that KF4 could be developed as a lung matrix-stabilizing therapy in emphysema.

Comparison of 16 Pediatric Acute Respiratory Distress Syndrome-Associated Plasma Biomarkers With Changing Lung Injury Severity.

OBJECTIVES: Pediatric acute respiratory distress syndrome (PARDS) is a source of substantial morbidity and mortality in the PICU, and different plasma biomarkers have identified different PARDS and ARDS subgroups. We have a poor understanding of how these biomarkers change over time and with changing lung injuries. We sought to determine how biomarker levels change over PARDS course, whether they are correlated, and whether they are different in critically ill non-PARDS patients. DESIGN: Two-center prospective observational study. SETTING: Two quaternary care academic children's hospitals. PATIENTS: Subjects under 18 years of age admitted to the PICU who were intubated and met the Second Pediatric Acute Lung Injury Consensus Conference-2 PARDS diagnostic criteria and nonintubated critically ill subjects without apparent lung disease. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Plasma samples were obtained on study days 1, 3, 7, and 14. The levels of 16 biomarkers were measured using a fluorometric bead-based assay. Compared with non-PARDS subjects, on day 1 PARDS subjects had increased concentrations of tumor necrosis factor-alpha, interleukin (IL)-8, interferon-γ, IL17, granzyme B, soluble intercellular adhesion molecule-1 (sICAM1), surfactant protein D, and IL18 but reduced matrix metalloproteinase 9 (MMP-9) concentrations (all p < 0.05). Day 1 biomarker concentrations and PARDS severity were not correlated. Over PARDS course, changes in 11 of the 16 biomarkers positively correlated with changing lung injury with sICAM1 ( R = 0.69, p = 2.2 × 10 -16 ) having the strongest correlation. By Spearman rank correlation of biomarker concentrations in PARDS subjects, we identified two patterns. One had elevations of plasminogen activator inhibitor-1, MMP-9, and myeloperoxidase, and the other had higher inflammatory cytokines. CONCLUSIONS: sICAM1 had the strongest positive correlation with worsening lung injury across all study time points suggesting that it is perhaps the most biologically relevant of the 16 analytes. There was no correlation between biomarker concentration on day 1 and day 1 PARDS severity; however, changes in most biomarkers over time positively correlated with changing lung injury. Finally, in day 1 samples, 7 of the 16 biomarkers were not significantly different between PARDS and critically ill non-PARDS subjects. These data highlight the difficulty of using plasma biomarkers to identify organ-specific pathology in critically ill patients.

Machine learning-driven identification of the gene-expression signature associated with a persistent multiple organ dysfunction trajectory in critical illness.

BACKGROUND: Multiple organ dysfunction syndrome (MODS) disproportionately drives morbidity and mortality among critically ill patients. However, we lack a comprehensive understanding of its pathobiology. Identification of genes associated with a persistent MODS trajectory may shed light on underlying biology and allow for accurate prediction of those at-risk. METHODS: Secondary analyses of publicly available gene-expression datasets. Supervised machine learning (ML) was used to identify a parsimonious set of genes associated with a persistent MODS trajectory in a training set of pediatric septic shock. We optimized model parameters and tested risk-prediction capabilities in independent validation and test datasets, respectively. We compared model performance relative to an established gene-set predictive of sepsis mortality. FINDINGS: Patients with a persistent MODS trajectory had 568 differentially expressed genes and characterized by a dysregulated innate immune response. Supervised ML identified 111 genes associated with the outcome of interest on repeated cross-validation, with an AUROC of 0.87 (95% CI: 0.85-0.88) in the training set. The optimized model, limited to 20 genes, achieved AUROCs ranging from 0.74 to 0.79 in the validation and test sets to predict those with persistent MODS, regardless of host age and cause of organ dysfunction. Our classifier demonstrated reproducibility in identifying those with persistent MODS in comparison with a published gene-set predictive of sepsis mortality. INTERPRETATION: We demonstrate the utility of supervised ML driven identification of the genes associated with persistent MODS. Pending validation in enriched cohorts with a high burden of organ dysfunction, such an approach may inform targeted delivery of interventions among at-risk patients. FUNDING: H.R.W.'s NIHR35GM126943 award supported the work detailed in this manuscript. Upon his death, the award was transferred to M.N.A. M.R.A., N.S.P, and R.K were supported by NIHR21GM151703. R.K. was supported by R01GM139967.

Regulation of epithelial transitional states in murine and human pulmonary fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a progressive scarring disease arising from impaired regeneration of the alveolar epithelium after injury. During regeneration, type 2 alveolar epithelial cells (AEC2s) assume a transitional state that upregulates multiple keratins and ultimately differentiate into AEC1s. In IPF, transitional AECs accumulate with ineffectual AEC1 differentiation. However, whether and how transitional cells cause fibrosis, whether keratins regulate transitional cell accumulation and fibrosis, and why transitional AECs and fibrosis resolve in mouse models but accumulate in IPF are unclear. Here, we show that human keratin 8 (KRT8) genetic variants were associated with IPF. Krt8-/- mice were protected from fibrosis and accumulation of the transitional state. Keratin 8 (K8) regulated the expression of macrophage chemokines and macrophage recruitment. Profibrotic macrophages and myofibroblasts promoted the accumulation of transitional AECs, establishing a K8-dependent positive feedback loop driving fibrogenesis. Finally, rare murine transitional AECs were highly senescent and basaloid and may not differentiate into AEC1s, recapitulating the aberrant basaloid state in human IPF. We conclude that transitional AECs induced and were maintained by fibrosis in a K8-dependent manner; in mice, most transitional cells and fibrosis resolved, whereas in human IPF, transitional AECs evolved into an aberrant basaloid state that persisted with progressive fibrosis.

A titratable murine model of progressive emphysema using tracheal porcine pancreatic elastase.

Progressive emphysema often leads to end-stage lung disease. Most mouse models of emphysema are typically modest (i.e. cigarette smoke exposure), and changes over time are difficult to quantify. The tracheal porcine pancreatic elastase model (PPE) produces severe injury, but the literature is conflicted as to whether emphysema improves, is stable, or progresses over time. We hypothesized a threshold of injury below which repair would occur and above which emphysema would be stable or progress. We treated 8-week-old C57BL6 mixed sex mice with 0, 0.5, 2, or 4 activity units of PPE in 100 µL PBS and performed lung stereology at 21 and 84 days. There were no significant differences in weight gain or mouse health. Despite minimal emphysema at 21-days in the 0.5 units group (2.8 µm increased mean linear intercept, MLI), MLI increased by 4.6 µm between days 21 and 84 (p = 0.0007). In addition to larger MLI at 21 days in 2- and 4-unit groups, MLI increases from day 21 to 84 were 17.2 and 34 µm respectively (p = 0.002 and p = 0.0001). Total lung volume increased, and alveolar surface area decreased with time and injury severity. Contrary to our hypothesis, we found no evidence of alveolar repair over time. Airspace destruction was both progressive and accelerative. Future mechanistic studies in lung immunity, mechano-biology, senescence, and cell-specific changes may lead to novel therapies to slow or halt progressive emphysema in humans.

Chymotrypsin-like Elastase-1 Mediates Progressive Emphysema in Alpha-1 Antitrypsin Deficiency.

Chymotrypsin-like elastase 1 (CELA1) is a serine protease that is neutralized by alpha-1antitrypsin (AAT) and prevents emphysema in a murine antisense oligonucleotide model of AAT-deficient emphysema. Mice with genetic ablation of AAT do not have emphysema at baseline but develop emphysema with injury and aging. We tested the role of the CELA1 gene in emphysema development in this genetic model of AAT-deficiency following tracheal lipopolysaccharide (LPS), 10 months of cigarette smoke exposure, aging, and a low-dose tracheal porcine pancreatic elastase (LD-PPE) model we developed. In this last model, we performed proteomic analysis to understand differences in lung protein composition. We were unable to show that AAT-deficient mice developed more emphysema than wild type with escalating doses of LPS. In the LD-PPE model, AAT-deficient mice developed significant and progressive emphysema from which Cela1-/- & AAT-deficient mice were protected. Cela1-/-& AAT-deficient lungs had more matrix-associated proteins than AAT-deficientlungs but also had more leukocyte-associated proteases. With cigarette smoke exposure, Cela1-/- &AAT-deficient mice had more emphysema than AAT-deficient mice but had less myeloperoxidase activity. Cela1-/-&AAT-deficient mice had less age-related airspace simplification than AAT-deficient and were comparable to wild type. While CELA1 promotes inflammation-independent emphysema progression and its absence preserves the lung matrix in multiple models of AAT-deficient emphysema, for unclear reasons Cela1 deficiency is associated with increased emphysema with cigarette smoke. While anti-CELA1 therapies could potentially be used to prevent emphysema progression in AAT deficiency after smoking cessation, an understanding of why and how cigarette smoke exacerbates emphysema in Cela1 deficiency and whether AAT replacement therapy mitigates this effect is needed first.

CELA1 Mediates Progressive Emphysema in Alpha-1 Antitrypsin Deficiency.

Chymotrypsin-like elastase 1 ( CELA1 ) is a serine protease that is neutralized by α1-antitrypsin (AAT) and prevents emphysema in a murine antisense oligonucleotide model of AAT-deficient emphysema. Mice with genetic ablation of AAT do not have emphysema at baseline but develop emphysema with injury and aging. We tested the role of CELA1 in emphysema development in this genetic model of AAT -deficiency following tracheal lipopolysacharide (LPS), 8 months of cigarette smoke (CS) exposure, aging, and a low-dose tracheal porcine pancreatic elastase (LD-PPE) model. In this last model, we performed proteomic analysis to understand differences in lung protein composition. We were unable to show that AAT -/ - mice developed more emphysema than wild type with LPS. In the LD-PPE model, AAT -/- mice developed progressive emphysema from which Cela1 -/- &AAT -/- mice were protected. In the CS model, Cela1 -/- &AAT -/- mice had worse emphysema than AAT -/- , and in the aging model, 72-75 week-old Cela1 -/- &AAT -/- mice had less emphysema than AAT -/- mice. Proteomic analysis of AAT -/- vs. wildtype lungs in the LD-PPE model showed reduced amounts of AAT proteins and increased amounts of proteins related to Rho and Rac1 GTPases and protein oxidation. Similar analysis of Cela1 -/- &AAT -/- vs. AAT -/- lungs showed differences in neutrophil degranulation, elastin fiber synthesis, and glutathione metabolism. Thus, Cela1 prevents post-injury emphysema progression in AAT -deficiency, but it has no effect and potentially worsens emphysema in response to chronic inflammation and injury. Prior to developing anti-CELA1 therapies for AAT-deficient emphysema, an understanding of why and how CS exacerbates emphysema in Cela1 deficiency is needed.

Multi-omic characterization of pediatric ARDS via nasal brushings.

RATIONALE: While nasal brushing transcriptomics can identify disease subtypes in chronic pulmonary diseases, it is unknown whether this is true in pediatric acute respiratory distress syndrome (PARDS). OBJECTIVES: Determine whether nasal transcriptomics and methylomics can identify clinically meaningful PARDS subgroups that reflect important pathobiological processes. METHODS: Nasal brushings and serum were collected on days 1, 3, 7, and 14 from control and PARDS subjects from two centers. PARDS duration was the primary endpoint. MEASUREMENTS AND MAIN RESULTS: Twenty-four control and 39 PARDS subjects were enrolled. Two nasal methylation patterns were identified. Compared to Methyl Subgroup 1, Subgroup 2 had hypomethylation of inflammatory genes and was enriched for immunocompromised subjects. Four transcriptomic patterns were identified with temporal patterns indicating injury, repair, and regeneration. Over time, both inflammatory (Subgroup B) and cell injury (Subgroup D) patterns transitioned to repair (Subgroup A) and eventually homeostasis (Subgroup C). When control specimens were included, they were largely Subgroup C. In comparison with 17 serum biomarkers, the nasal transcriptome was more predictive of prolonged PARDS. Subjects with initial Transcriptomic Subgroup B or D assignment had median PARDS duration of 8 days compared to 2 in A or C (p = 0.02). For predicting PARDS duration ≥ 3 days, nasal transcriptomics was more sensitive and serum biomarkers more specific. CONCLUSIONS: PARDS nasal transcriptome may reflect distal lung injury, repair, and regeneration. A combined nasal PCR and serum biomarker assay could be useful for predictive and diagnostic enrichment. Trial registration Clinicaltrials.gov NCT03539783 May 29, 2018.

Mouse lung organoid responses to reduced, increased, and cyclic stretch.

Most lung development occurs in the context of cyclic stretch. Alteration of the mechanical microenvironment is a common feature of many pulmonary diseases, with congenital diaphragmatic hernia (CDH) and fetal tracheal occlusion (FETO, a therapy for CDH) being extreme examples with changes in lung structure, cell differentiation, and function. To address limitations in cell culture and in vivo mechanotransductive models, we developed two mouse lung organoid (mLO) mechanotransductive models using postnatal day 5 (PND5) mouse lung CD326-positive cells and fibroblasts subjected to increased, decreased, and cyclic strain. In the first model, mLOs were exposed to forskolin (FSK) and/or disrupted (DIS) and evaluated at 20 h. mLO cross-sectional area changed by +59%, +24%, and -68% in FSK, control, and DIS mLOs, respectively. FSK-treated organoids had twice as many proliferating cells as other organoids. In the second model, 20 h of 10.25% biaxial cyclic strain increased the mRNAs of lung mesenchymal cell lineages compared with static stretch and no stretch. Cyclic stretch increased TGF-ß and integrin-mediated signaling, with upstream analysis indicating roles for histone deacetylases, microRNAs, and long noncoding RNAs. Cyclic stretch mLOs increased αSMA-positive and αSMA-PDGFRα-double-positive cells compared with no stretch and static stretch mLOs. In this PND5 mLO mechanotransductive model, cell proliferation is increased by static stretch, and cyclic stretch induces mesenchymal gene expression changes important in postnatal lung development.

Fetal Tracheal Occlusion Increases Lung Basal Cells via Increased Yap Signaling.

Fetal endoscopic tracheal occlusion (FETO) is an emerging surgical therapy for congenital diaphragmatic hernia (CDH). Ovine and rabbit data suggested altered lung epithelial cell populations after tracheal occlusion (TO) with transcriptomic signatures implicating basal cells. To test this hypothesis, we deconvolved mRNA sequencing (mRNA-seq) data and used quantitative image analysis in fetal rabbit lung TO, which had increased basal cells and reduced ciliated cells after TO. In a fetal mouse TO model, flow cytometry showed increased basal cells, and immunohistochemistry demonstrated basal cell extension to subpleural airways. Nuclear Yap, a known regulator of basal cell fate, was increased in TO lung, and Yap ablation on the lung epithelium abrogated TO-mediated basal cell expansion. mRNA-seq of TO lung showed increased activity of downstream Yap genes. Human lung specimens with congenital and fetal tracheal occlusion had clusters of subpleural basal cells that were not present in the control. TO increases lung epithelial cell nuclear Yap, leading to basal cell expansion.

Validating in vivo hyperpolarized 129 Xe diffusion MRI and diffusion morphometry in the mouse lung.

PURPOSE: Diffusion and lung morphometry imaging using hyperpolarized gases are promising tools to quantify pulmonary microstructure noninvasively in humans and in animal models. These techniques assume the motion encoded is exclusively diffusive gas displacement, but the impact of cardiac motion on measurements has never been explored. Furthermore, although diffusion morphometry has been validated against histology in humans and mice using 3 He, it has never been validated in mice for 129 Xe. Here, we examine the effect of cardiac motion on diffusion imaging and validate 129 Xe diffusion morphometry in mice. THEORY AND METHODS: Mice were imaged using gradient-echo-based diffusion imaging, and apparent diffusion-coefficient (ADC) maps were generated with and without cardiac gating. Diffusion-weighted images were fit to a previously developed theoretical model using Bayesian probability theory, producing morphometric parameters that were compared with conventional histology. RESULTS: Cardiac gating had no significant impact on ADC measurements (dual-gating: ADC = 0.020 cm2 /s, single-gating: ADC = 0.020 cm2 /s; P = .38). Diffusion-morphometry-generated maps of ADC (mean, 0.0165 ± 0.0001 cm2 /s) and acinar dimensions (alveolar sleeve depth [h] = 44 µm, acinar duct radii [R] = 99 µm, mean linear intercept [Lm ] = 74 µm) that agreed well with conventional histology (h = 45 µm, R = 108 µm, Lm = 63 µm). CONCLUSION: Cardiac motion has negligible impact on 129 Xe ADC measurements in mice, arguing its impact will be similarly minimal in humans, where relative cardiac motion is reduced. Hyperpolarized 129 Xe diffusion morphometry accurately and noninvasively maps the dimensions of lung microstructure, suggesting it can quantify the pulmonary microstructure in mouse models of lung disease.

Peripheral blood transcriptomic sub-phenotypes of pediatric acute respiratory distress syndrome.

BACKGROUND: Acute respiratory distress syndrome (ARDS) is heterogeneous and may be amenable to sub-phenotyping to improve enrichment for trials. We aimed to identify subtypes of pediatric ARDS based on whole blood transcriptomics. METHODS: This was a prospective observational study of children with ARDS at the Children's Hospital of Philadelphia (CHOP) between January 2018 and June 2019. We collected blood within 24 h of ARDS onset, generated expression profiles, and performed k-means clustering to identify sub-phenotypes. We tested the association between sub-phenotypes and PICU mortality and ventilator-free days at 28 days using multivariable logistic and competing risk regression, respectively. RESULTS: We enrolled 106 subjects, of whom 96 had usable samples. We identified three sub-phenotypes, dubbed CHOP ARDS Transcriptomic Subtypes (CATS) 1, 2, and 3. CATS-1 subjects (n = 31) demonstrated persistent hypoxemia, had ten subjects (32%) with immunocompromising conditions, and 32% mortality. CATS-2 subjects (n = 29) had more immunocompromising diagnoses (48%), rapidly resolving hypoxemia, and 24% mortality. CATS-3 subjects (n = 36) had the fewest comorbidities and also had rapidly resolving hypoxemia and 8% mortality. The CATS-3 subtype was associated with lower mortality (OR 0.18, 95% CI 0.04-0.86) and higher probability of extubation (subdistribution HR 2.39, 95% CI 1.32-4.32), relative to CATS-1 after adjustment for confounders. CONCLUSIONS: We identified three sub-phenotypes of pediatric ARDS using whole blood transcriptomics. The sub-phenotypes had divergent clinical characteristics and prognoses. Further studies should validate these findings and investigate mechanisms underlying differences between sub-phenotypes.

Lung microstructure in adolescent idiopathic scoliosis before and after posterior spinal fusion.

Adolescent idiopathic scoliosis (AIS) is associated with decreased respiratory quality of life and impaired diaphragm function. Recent hyperpolarized helium (HHe) MRI studies show alveolarization continues throughout adolescence, and mechanical forces are known to impact alveolarization. We therefore hypothesized that patients with AIS would have alterations in alveolar size, alveolar number, or alveolar septal dimensions compared to adolescents without AIS, and that posterior spinal fusion (PSF) might reverse these differences. We conducted a prospective observational trial using HHe MRI to test for changes in alveolar microstructure in control and AIS subjects at baseline and one year. After obtaining written informed consent from subjects' legal guardians and assent from the subjects, we performed HHe and proton MRI in 14 AIS and 16 control subjects aged 8-21 years. The mean age of control subjects (12.9 years) was significantly less than AIS (14.9 years, p = 0.003). At baseline, there were no significant differences in alveolar size, number, or alveolar duct morphometry between AIS and control subjects or between the concave (compressed) and convex (expanded) lungs of AIS subjects. At one year after PSF AIS subjects had an increase in alveolar density in the formerly convex lung (p = 0.05), likely reflecting a change in thoracic anatomy, but there were no other significant changes in lung microstructure. Modeling of alveolar size over time demonstrated similar rates of alveolar growth in control and AIS subjects in both right and left lungs pre- and post-PSF. Although this study suffered from poor age-matching, we found no evidence that AIS or PSF impacts lung microstructure. Trial registration: Clinical trial registration number NCT03539770.