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Andrographolide Sulfonate Attenuates Acute Lung Injury by Reducing Expression of Myeloperoxidase and Neutrophil-Derived Proteases in Mice.

ABSTRACT: Andrographolide sulfonate (Andro-S), a sulfonation derivative of andrographolide, is known to be effective in treating inflammation-related diseases, while the underlying mechanisms and global protein alterations in response to Andro-S remain unknown. This study aimed to investigate the pharmacological effects and potential targets of Andro-S in a murine model of acute lung injury (ALI). ALI was induced by aerosolized lipopolysaccharide (LPS) exposure before treatment with Andro-S. Inflammatory state of each treatment group was determined by histological analysis and quantification of inflammatory markers. Differentially expressed proteins in lung tissues were identified by an iTRAQ-based quantitative proteomic approach and further confirmed by immunohistochemistry analysis. Administration of Andro-S alleviated LPS-induced histological changes in the lung and reduced the expression of inflammatory markers in serum, bronchoalveolar fluid and lung tissues. Proteomic analysis identified 31 differentially expressed proteins from a total of 2,234 quantified proteins in the lung. According to bioinformatics analysis, neutrophil elastase (ELANE), cathepsin G (CTSG) and myeloperoxidase (MPO), three neutrophil-derived proteases related to immune system process and defense responses to fungi were chosen as potential targets of Andro-S. Further immunohistochemistry analysis confirmed the inhibitory effects of Andro-S on LPS-induced ELANE, CTSG and MPO up-regulation. These results indicate that Andro-S suppressed the severity of LPS-induced ALI, possibly by attenuating the expression of and neutrophil-derived proteases.

SUBMITTER: Gao F

PROVIDER: S-EPMC6107831 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Andrographolide Sulfonate Attenuates Acute Lung Injury by Reducing Expression of Myeloperoxidase and Neutrophil-Derived Proteases in Mice.

Gao Fei F Liu Xing X Shen Ziying Z Jia Xiaohui X He Han H Gao Jing J Wu Jianhong J Jiang Chunhong C Zhou Hu H Wang Yiping Y

Frontiers in physiology 20180817

Andrographolide sulfonate (Andro-S), a sulfonation derivative of andrographolide, is known to be effective in treating inflammation-related diseases, while the underlying mechanisms and global protein alterations in response to Andro-S remain unknown. This study aimed to investigate the pharmacological effects and potential targets of Andro-S in a murine model of acute lung injury (ALI). ALI was induced by aerosolized lipopolysaccharide (LPS) exposure before treatment with Andro-S. Inflammatory ...[more]

PMID: 30174607

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Project description:To test the hypothesis that restoration of antithrombin plasma concentrations attenuates vascular leakage by inhibiting neutrophil activation through syndecan-4 receptor inhibition in an established ovine model of acute lung injury.Randomized controlled laboratory experiment.University animal research facility.Eighteen chronically instrumented sheep.Following combined burn and smoke inhalation injury (40% of total body surface area, third-degree flame burn; 4 × 12 breaths of cold cotton smoke), chronically instrumented sheep were randomly assigned to receive an IV infusion of 6 IU/kg/hr recombinant human antithrombin III or normal saline (n = 6 each) during the 48-hour study period. In addition, six sham animals (not injured, continuous infusion of vehicle) were used to obtain reference values for histological and immunohistochemical analyses.Compared to control animals, recombinant human antithrombin III reduced the number of neutrophils per hour in the pulmonary lymph (p < 0.01 at 24 and 48 hr), alveolar neutrophil infiltration (p = 0.04), and pulmonary myeloperoxidase activity (p = 0.026). Flow cytometric analysis revealed a significant reduction of syndecan-4-positive neutrophils (p = 0.002 vs control at 24 hr). Treatment with recombinant human antithrombin III resulted in a reduction of pulmonary nitrosative stress (p = 0.002), airway obstruction (bronchi: p = 0.001, bronchioli: p = 0.013), parenchymal edema (p = 0.044), and lung bloodless wet-to-dry-weight ratio (p = 0.015). Clinically, recombinant human antithrombin III attenuated the increased pulmonary transvascular fluid flux (12-48 hr: p ? 0.001 vs control each) and the deteriorated pulmonary gas exchange (12-48 hr: p < 0.05 vs control each) without increasing the risk of bleeding.The present study provides evidence for the interaction between antithrombin and neutrophils in vivo, its pathophysiological role in vascular leakage, and the therapeutic potential of recombinant human antithrombin III in a large animal model of acute lung injury.

Project description:IntroductionSepsis refers to severe systemic inflammation leading to acute lung injury (ALI) and death. Introducing novel therapies can reduce the mortality in ALI. Osteopontin (OPN), a secretory glycoprotein produced by immune reactive cells, plays a deleterious role in various inflammatory diseases. However, its role in ALI caused by sepsis remains unexplored. We hypothesize that treatment with an OPN-neutralizing antibody (anti-OPN Ab) protects mice against ALI during sepsis.MethodsSepsis was induced in 8-week-old male C57BL/6 mice by cecal ligation and puncture (CLP). Anti-OPN Ab or non-immunized IgG as control, at a dose of 50 μg/mouse, was intravenously injected at the time of CLP. After 20 hours, the expression of OPN and proinflammatory cytokines in tissues and plasma was examined by real-time PCR, Western blot, and ELISA. Plasma levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and lactate dehydrogenase (LDH) and the lung myeloperoxidase (MPO) levels were determined by colorimetric assays. Lung damage and neutrophil infiltrations were determined by histological H&E and Gr-1 staining, respectively. The effect of recombinant mouse OPN (rmOPN) on human neutrophil-like cell (HL-60) migration was performed by Boyden chamber assays and the involvement of intracellular signaling molecules in HL-60 cells was revealed by Western blot.ResultsAfter 20 hours of sepsis, mRNA and protein levels of OPN were significantly induced in lungs, spleen, and plasma. Treatment with an anti-OPN Ab in septic mice significantly reduced the plasma levels of ALT, AST, and LDH, and the proinflammatory cytokines IL-6, IL-1β and the chemokine MIP-2, compared with the vehicle group. Similarly, the lung mRNA and protein expressions of proinflammatory cytokines and chemokine were greatly reduced in anti-OPN Ab-treated animals. The lung histological architecture, MPO and neutrophil infiltration were significantly improved in anti-OPN Ab-treated mice compared with the vehicle animals. Treatment of rmOPN in HL-60 cells significantly increased their migration, in vitro. The neutrophils treated with rmOPN remarkably increased the levels of phospho focal adhesion kinase (pFAK), phospho extracellular signal-regulated kinase (pERK) and phospho p38.ConclusionsOur findings clearly demonstrate the beneficial outcomes of anti-OPN Ab treatment in protecting against ALI, implicating a novel therapeutic strategy in sepsis.

Project description:BackgroundMyeloperoxidase released after neutrophil and monocyte activation can generate reactive oxygen species, leading to host tissue damage. Extracellular glomerular myeloperoxidase deposition, seen in ANCA-associated vasculitis, may enhance crescentic GN through antigen-specific T and B cell activation. Myeloperoxidase-deficient animals have attenuated GN early on, but augmented T cell responses. We investigated the effect of myeloperoxidase inhibition, using the myeloperoxidase inhibitor AZM198, to understand its potential role in treating crescentic GN.MethodsWe evaluated renal biopsy samples from patients with various forms of crescentic GN for myeloperoxidase and neutrophils, measured serum myeloperoxidase concentration in patients with ANCA-associated vasculitis and controls, and assessed neutrophil extracellular trap formation, reactive oxygen species production, and neutrophil degranulation in ANCA-stimulated neutrophils in the absence and presence of AZM198. We also tested the effect of AZM198 on ANCA-stimulated neutrophil-mediated endothelial cell damage in vitro, as well as on crescentic GN severity and antigen-specific T cell reactivity in the murine model of nephrotoxic nephritis.ResultsAll biopsy specimens with crescentic GN had extracellular glomerular myeloperoxidase deposition that correlated significantly with eGFR and crescent formation. In vitro, AZM198 led to a significant reduction in neutrophil extracellular trap formation, reactive oxygen species production, and released human neutrophil peptide levels, and attenuated neutrophil-mediated endothelial cell damage. In vivo, delayed AZM198 treatment significantly reduced proteinuria, glomerular thrombosis, serum creatinine, and glomerular macrophage infiltration, without increasing adaptive T cell responses.ConclusionsMyeloperoxidase inhibition reduced neutrophil degranulation and neutrophil-mediated endothelial cell damage in patients with ANCA-associated vasculitis. In preclinical crescentic GN, delayed myeloperoxidase inhibition suppressed kidney damage without augmenting adaptive immune responses, suggesting it might offer a novel adjunctive therapeutic approach in crescentic GN.

Dataset Information

Andrographolide Sulfonate Attenuates Acute Lung Injury by Reducing Expression of Myeloperoxidase and Neutrophil-Derived Proteases in Mice.

Publications

Andrographolide Sulfonate Attenuates Acute Lung Injury by Reducing Expression of Myeloperoxidase and Neutrophil-Derived Proteases in Mice.

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