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Objectives: Monocytes are plastic cells that assume different polarization states that can either promote inflammation or tissue repair and inflammation resolution. Polarized monocytes are partially defined by their transcriptional profiles that are influenced by environmental stimuli. The airway monocyte response in pediatric acute respiratory distress syndrome (PARDS) is undefined. To identify differentially expressed genes and networks using a novel transcriptomic reporter assay with donor monocytes exposed to the airway fluid of intubated children with and at-risk for PARDS. To determine differences in gene expression at two time points using the donor monocyte assay exposed to airway fluid from intubated children with PARDS obtained 48-96 hours following initial tracheal aspirate sampling., Design: In vitro pilot study carried out using airway fluid supernatant., Setting: Academic 40-bed PICU., Participants: Fifty-seven children: 44 children with PARDS and 13 children at-risk for PARDS., Interventions: None., Measurements and Main Results: We performed bulk RNA sequencing using a transcriptomic reporter assay of monocytes exposed to airway fluid from intubated children to discover gene networks differentiating PARDS from at-risk for PARDS and those differentiating mild/moderate from severe PARDS. We also report differences in gene expression in children with PARDS 48-96 hours following initial tracheal aspirate sampling. We found that interleukin (IL)-10, IL-4, and IL-13, cytokine/chemokine signaling, and the senescence-associated secretory phenotype are upregulated in monocytes exposed to airway fluid from intubated children with PARDS compared with those at-risk for PARDS. Signaling by NOTCH, histone deacetylation/acetylation, DNA methylation, chromatin modifications (B-WICH complex), and RNA polymerase I transcription and its associated regulatory apparatus were upregulated in children with PARDS 48-96 hours following initial tracheal aspirate sampling., Conclusions: We identified gene networks important to the PARDS airway immune response using bulk RNA sequencing from a monocyte reporter assay that exposed monocytes to airway fluid from intubated children with and at-risk for PARDS. Mechanistic investigations are needed to validate our findings., Competing Interests: The authors have disclosed that they do not have any potential conflicts of interest., (Copyright © 2024 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of the Society of Critical Care Medicine.)

CD4 + T cells contribute to lung inflammation in acute respiratory distress syndrome. The CD4 + T-cell response in pediatric acute respiratory distress syndrome (PARDS) is unknown., Objectives: To identify differentially expressed genes and networks using a novel transcriptomic reporter assay with donor CD4 + T cells exposed to the airway fluid of intubated children with mild versus severe PARDS., Design: In vitro pilot study., Setting: Laboratory-based study using human airway fluid samples admitted to a 36-bed university-affiliated pediatric intensive care unit., Patients/subjects: Seven children with severe PARDS, nine children with mild PARDS, and four intubated children without lung injury as controls., Interventions: None., Measurements and Main Results: We performed bulk RNA sequencing using a transcriptomic reporter assay of CD4 + T cells exposed to airway fluid from intubated children to discover gene networks differentiating severe from mild PARDS. We found that innate immunity pathways, type I (α and β), and type II (γ) interferon response and cytokine/chemokine signaling are downregulated in CD4 + T cells exposed to airway fluid from intubated children with severe PARDS compared with those with mild PARDS., Conclusions: We identified gene networks important to the PARDS airway immune response using bulk RNA sequencing from a novel CD4 + T-cell reporter assay that exposed CD4 + T cells to airway fluid from intubated children with severe and mild PARDS. These pathways will help drive mechanistic investigations into PARDS. Validation of our findings using this transcriptomic reporter assay strategy is needed., Competing Interests: The authors have disclosed that they do not have any potential conflicts of interest., (Copyright © 2023 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of the Society of Critical Care Medicine.)

Pediatric acute respiratory distress syndrome (PARDS) is a heterogeneous illness affecting 6% of mechanically ventilated children and with an overall mortality of 17%. Studies in PARDS have mainly focused on plasma biomarkers which may not reflect airway biomarkers. We lack adequate understanding of the inflammatory mediators and underlying immune responses in the airways of PARDS patients. Our objective was to compare the levels of cytokines in the airway fluid of intubated children with severe versus nonsevere acute respiratory distress syndrome., Design: Prospective observational cohort study., Setting: Single 36-bed quaternary care academic safety-net hospital PICU., Patients: Children intubated for acute respiratory failure between January 2018 and November 2021 stratified by Pediatric Acute Lung Injury Consensus Conference-1 criteria for PARDS., Interventions: None., Measurements and Main Results: We measured levels of 23 cytokines, chemokines, and protein biomarkers in the tracheal aspirate from 82 intubated children, between 14 days and 17 years old, at risk for or with PARDS. Levels of interleukin-4, -5, -7, -8, -12(p-70), -17a, -21, and fractalkine were higher in patients with severe versus nonsevere PARDS. There were no associations between airway and plasma cytokines., Conclusions: Proinflammatory cytokines are elevated in the airway fluid from intubated children with severe PARDS and reflect diverse patterns of airway inflammation., Competing Interests: The authors have disclosed that they do not have any potential conflicts of interest., (Copyright © 2022 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of the Society of Critical Care Medicine.)