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Platelets are essential for normal hemostasis and thrombosis but become hyperactive in hemolytic disorders. Cell-free heme is known to be toxic to platelets and endothelial cells, playing a significant role in the progression of pathological complications in various hemolytic conditions. The abnormal activation of circulatory platelets results in micro/macrovascular thrombosis and clot formation in the lungs, worsening the disease. This work aimed to establish the potent bioactive molecule that can regulate the heme-induced toxicity in platelets. We found that unconjugated bilirubin (UCB), an endogenous antioxidant and a byproduct of heme degradation, exhibited a higher protective effect against hemin-induced platelet aggregation and activation. This protective effect could mainly be due to reducing ROS and lipid peroxidation-mediated ferroptosis in hemin-treated platelets. Further experiments suggested that by blocking the interaction between hemin and the CLEC-2 receptor, UCB regulates the downstream Syk phosphorylation, a key event in hemin-induced platelet toxicity. Thus, UCB is emerging as a natural regulatory molecule that mitigates hemin-induced platelet toxicity and holds promise as an adjunctive therapy for managing platelet-associated complications, particularly in hemolytic disorders., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Ltd.)

Traffic-related air pollution (TRAP) exposure is associated with systemic health effects, which can be studied using blood-based markers. Although we have previously shown that high TRAP concentrations alter the plasma proteome, the concentration-response relationship between blood proteins and TRAP is unexplored in controlled human exposure studies. We aimed to identify concentration-dependent plasma markers of diesel exhaust (DE), a model of TRAP. Fifteen healthy non-smokers were enrolled into a double-blinded, crossover study where they were exposed to filtered air (FA) and DE at 20, 50 and 150 μg/m 3 PM 2.5 for 4h, separated by ≥ 4-week washouts. We collected blood at 24h post-exposure and used label-free mass spectrometry to quantify proteins in plasma. Proteins exhibiting a concentration-response, as determined by linear mixed effects models (LMEMs), were assessed for pathway enrichment using WebGestalt. Top candidates, identified by sparse partial least squares discriminant analysis and LMEMs, were confirmed using enzyme-linked immunoassays. Thereafter, we assessed correlations between proteins that showed a DE concentration-response and acute inflammatory endpoints, forced expiratory volume in 1 s (FEV 1 ) and methacholine provocation concentration causing a 20% drop in FEV 1 (PC 20 ). DE exposure was associated with concentration-dependent alterations in 45 proteins, which were enriched in complement pathways. Of the 9 proteins selected for confirmatory immunoassays, based on complementary bioinformatic approaches to narrow targets and availability of high-quality assays, complement factor I (CFI) exhibited a significant concentration-dependent decrease (-0.02 μg/mL per μg/m 3 of PM 2.5 , p = 0.04). Comparing to FA at discrete concentrations, CFI trended downward at 50 (-2.14 ± 1.18, p = 0.08) and significantly decreased at 150 μg/m 3 PM 2.5 (-2.93 ± 1.18, p = 0.02). CFI levels were correlated with FEV 1 , PC 20 and nasal interleukin (IL)-6 and IL-1β. This study details concentration-dependent alterations in the plasma proteome following DE exposure at concentrations relevant to occupational and community settings. CFI shows a robust concentration-response and association with established measures of airway function and inflammation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Introduction: TNFα-inducible matrix metalloproteinases play a critical role in the process of airway remodeling in respiratory inflammatory disease including asthma. The cationic host defense peptide LL-37 is elevated in the lungs during airway inflammation. However, the impact of LL-37 on TNFα-driven processes is not well understood. Here, we examined the effect of LL-37 on TNFα-mediated responses in human bronchial epithelial cells (HBECs)., Methods: We used a slow off-rate modified aptamer-based proteomics approach to define the HBEC proteome altered in response to TNFα. Abundance of selected protein candidates and signaling intermediates was examined using immunoassays, ELISA and Western blots, and mRNA abundance was examined by qRT-PCR., Results: Proteomics analysis revealed that 124 proteins were significantly altered, 12 proteins were enhanced by ≥2-fold compared to unstimulated cells, in response to TNFα. MMP9 was the topmost increased protein in response to TNFα, enhanced by ∼10-fold, and MMP13 was increased by ∼3-fold, compared to unstimulated cells. Furthermore, we demonstrated that LL-37 significantly suppressed TNFα-mediated MMP9 and MMP13 in HBEC. Mechanistic data revealed that TNFα-mediated MMP9 and MMP13 production is controlled by SRC kinase and that LL-37 enhances related upstream negative regulators, namely, phospho-AKT (T308) and TNFα-mediated TNFAIP3 or A20., Conclusions: The findings of this study suggest that LL-37 may play a role in intervening in the process of airway remodeling in chronic inflammatory respiratory disease such as asthma., (© 2024 The Author(s). Published by S. Karger AG, Basel.)