PLUNC Is a Novel Airway Surfactant Protein with Anti-Biofilm Activity

The PLUNC ("Palate, lung, nasal epithelium clone") protein is an abundant secretory product of epithelia present throughout the conducting airways of humans and other mammals, which is evolutionarily related to the lipid transfer/lipopolysaccharide binding protein (LT/LBP) family. Two members of this family--the bactericidal/permeability increasing protein (BPI) and the lipopolysaccharide binding protein (LBP)--are innate immune molecules with recognized roles in sensing and responding to Gram negative bacteria, leading many to propose that PLUNC may play a host defense role in the human airways.Based on its marked hydrophobicity, we hypothesized that PLUNC may be an airway surfactant. We found that purified recombinant human PLUNC greatly enhanced the ability of aqueous solutions to spread on a hydrophobic surface. Furthermore, we discovered that PLUNC significantly reduced surface tension at the air-liquid interface in aqueous solutions, indicating novel and biologically relevant surfactant properties. Of note, surface tensions achieved by adding PLUNC to solutions are very similar to measurements of the surface tension in tracheobronchial secretions from humans and animal models. Because surfactants of microbial origin can disperse matrix-encased bacterial clusters known as biofilms [1], we hypothesized that PLUNC may also have anti-biofilm activity. We found that, at a physiologically relevant concentration, PLUNC inhibited biofilm formation by the airway pathogen Pseudomonas aeruginosa in an in vitro model.Our data suggest that the PLUNC protein contributes to the surfactant properties of airway secretions, and that this activity may interfere with biofilm formation by an airway pathogen

A P53-TLR3 Axis Ameliorates Pulmonary Hypertension by Inducing BMPR2 Via IRF3

Pulmonary arterial hypertension (PAH) features pathogenic and abnormal endothelial cells (ECs), and one potential origin is clonal selection. We studied the role of p53 and toll-like receptor 3 (TLR3) in clonal expansion and pulmonary hypertension (PH) via regulation of bone morphogenetic protein (BMPR2) signaling. ECs of PAH patients had reduced p53 expression. EC-specific p53 knockout exaggerated PH, and clonal expansion reduced p53 and TLR3 expression in rat lung CD117+ ECs. Reduced p53 degradation (Nutlin 3a) abolished clonal EC expansion, induced TLR3 and BMPR2, and ameliorated PH. Polyinosinic/polycytidylic acid [Poly(I:C)] increased BMPR2 signaling in ECs via enhanced binding of interferon regulatory factor-3 (IRF3) to the BMPR2 promoter and reduced PH in p53−/− mice but not in mice with impaired TLR3 downstream signaling. Our data show that a p53/TLR3/IRF3 axis regulates BMPR2 expression and signaling in ECs. This link can be exploited for therapy of PH

Serum metabolomic signatures of fatty acid oxidation defects differentiate host-response subphenotypes of acute respiratory distress syndrome

BACKGROUND: Fatty acid oxidation (FAO) defects have been implicated in experimental models of acute lung injury and associated with poor outcomes in critical illness. In this study, we examined acylcarnitine profiles and 3-methylhistidine as markers of FAO defects and skeletal muscle catabolism, respectively, in patients with acute respiratory failure. We determined whether these metabolites were associated with host-response ARDS subphenotypes, inflammatory biomarkers, and clinical outcomes in acute respiratory failure. METHODS: In a nested case-control cohort study, we performed targeted analysis of serum metabolites of patients intubated for airway protection (airway controls), Class 1 (hypoinflammatory), and Class 2 (hyperinflammatory) ARDS patients (N = 50 per group) during early initiation of mechanical ventilation. Relative amounts were quantified by liquid chromatography high resolution mass spectrometry using isotope-labeled standards and analyzed with plasma biomarkers and clinical data. RESULTS: Of the acylcarnitines analyzed, octanoylcarnitine levels were twofold increased in Class 2 ARDS relative to Class 1 ARDS or airway controls (P = 0.0004 and \u3c 0.0001, respectively) and was positively associated with Class 2 by quantile g-computation analysis (P = 0.004). In addition, acetylcarnitine and 3-methylhistidine were increased in Class 2 relative to Class 1 and positively correlated with inflammatory biomarkers. In all patients within the study with acute respiratory failure, increased 3-methylhistidine was observed in non-survivors at 30 days (P = 0.0018), while octanoylcarnitine was increased in patients requiring vasopressor support but not in non-survivors (P = 0.0001 and P = 0.28, respectively). CONCLUSIONS: This study demonstrates that increased levels of acetylcarnitine, octanoylcarnitine, and 3-methylhistidine distinguish Class 2 from Class 1 ARDS patients and airway controls. Octanoylcarnitine and 3-methylhistidine were associated with poor outcomes in patients with acute respiratory failure across the cohort independent of etiology or host-response subphenotype. These findings suggest a role for serum metabolites as biomarkers in ARDS and poor outcomes in critically ill patients early in the clinical course

RNA sequencing identifies common pathways between cigarette smoke exposure and replicative senescence in human airway epithelia

Abstract Background Aging is affected by genetic and environmental factors, and cigarette smoking is strongly associated with accumulation of senescent cells. In this study, we wanted to identify genes that may potentially be beneficial for cell survival in response to cigarette smoke and thereby may contribute to development of cellular senescence. Results Primary human bronchial epithelial cells from five healthy donors were cultured, treated with or without 1.5% cigarette smoke extract (CSE) for 24 h or were passaged into replicative senescence. Transcriptome changes were monitored using RNA-seq in CSE and non-CSE exposed cells and those passaged into replicative senescence. We found that, among 1534 genes differentially regulated during senescence and 599 after CSE exposure, 243 were altered in both conditions, representing strong enrichment. Pathways and gene sets overrepresented in both conditions belonged to cellular processes that regulate reactive oxygen species, proteasome degradation, and NF-κB signaling. Conclusions Our results offer insights into gene expression responses during cellular aging and cigarette smoke exposure, and identify potential molecular pathways that are altered by cigarette smoke and may also promote airway epithelial cell senescence

Masking of a Nuclear Signal Motif by Monoubiquitination Leads to Mislocalization and Degradation of the Regulatory Enzyme Cytidylyltransferase▿ †

Monoubiquitination aids in the nuclear export and entrance of proteins into the lysosomal degradative pathway, although the mechanisms are unknown. Cytidylyltransferase (CCTα) is a proteolytically sensitive lipogenic enzyme containing an NH2-terminal nuclear localization signal (NLS). We show here that CCTα is monoubiquitinated at a molecular site (K57) juxtaposed near its NLS, resulting in disruption of its interaction with importin-α, nuclear exclusion, and subsequent degradation within the lysosome. Cellular expression of a CCTα-ubiquitin fusion protein that mimics the monoubiquitinated enzyme resulted in cytoplasmic retention. A CCTα K57R mutant exhibited an extended half-life, was retained in the nucleus, and displayed proteolytic resistance. Importantly, by using CCTα-ubiquitin hybrid constructs that vary in the intermolecular distance between ubiquitin and the NLS, we show that CCTα monoubiquitination masks its NLS, resulting in cytoplasmic retention. These results unravel a unique molecular mechanism whereby monoubiquitination governs the trafficking and life span of a critical regulatory enzyme in vivo