title: Neutrophils generate microparticles during exposure to inert gases due to cytoskeletal oxidative stress.
abstract: This investigation was to elucidate the mechanism for microparticle (MP) formation triggered by exposures to high pressure inert gases. Human neutrophils generate MPs at a threshold of ∼186 kilopascals with exposures of 30 min or more. Murine cells are similar, but MP production occurs at a slower rate and continues for ∼4 h, whether or not cells remain under pressure. Neutrophils exposed to elevated gas but not hydrostatic pressure produce MPs according to the potency series: argon ≃ nitrogen > helium. Following a similar pattern, gases activate type-2 nitric-oxide synthase (NOS-2) and NADPH oxidase (NOX). MP production does not occur with neutrophils exposed to a NOX inhibitor (Nox2ds) or a NOS-2 inhibitor (1400W) or with cells from mice lacking NOS-2. Reactive species cause S-nitrosylation of cytosolic actin that enhances actin polymerization. Protein cross-linking and immunoprecipitation studies indicate that increased polymerization occurs because of associations involving vasodilator-stimulated phosphoprotein, focal adhesion kinase, the H(+)/K(+) ATPase β (flippase), the hematopoietic cell multidrug resistance protein ABC transporter (floppase), and protein-disulfide isomerase in proximity to short actin filaments. Using chemical inhibitors or reducing cell concentrations of any of these proteins with small inhibitory RNA abrogates NOS-2 activation, reactive species generation, actin polymerization, and MP production. These effects were also inhibited in cells exposed to UV light, which photoreverses S-nitrosylated cysteine residues and by co-incubations with the antioxidant ebselen or cytochalasin D. The autocatalytic cycle of protein activation is initiated by inert gas-mediated singlet O2 production.
authors.affiliation: From the Department of Emergency Medicine, University of Maryland, Baltimore, Maryland 21201 sthom@smail.umaryland.edu.
authors.forename: Stephen R
authors.initials: SR
authors.lastname: Thom
authors.affiliation: From the Department of Emergency Medicine, University of Maryland, Baltimore, Maryland 21201.
authors.forename: Veena M
authors.initials: VM
authors.lastname: Bhopale
authors.affiliation: From the Department of Emergency Medicine, University of Maryland, Baltimore, Maryland 21201.
authors.forename: Ming
authors.initials: M
authors.lastname: Yang
mesh_terms: D000199:Actins / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D055162:Biocatalysis / Q000187:drug effects; D003599:Cytoskeleton / Q000187:drug effects* / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D051379:Mice; D008863:Microspheres; D009504:Neutrophils / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism; D005741:Noble Gases / Q000494:pharmacology*; D018384:Oxidative Stress / Q000187:drug effects*; D010539:Permeability / Q000187:drug effects; D011312:Pressure; D055503:Protein Multimerization / Q000187:drug effects; D020836:Protein Structure, Quaternary; D026082:Singlet Oxygen / Q000378:metabolism
chemical_list: D000199:Actins; D005741:Noble Gases; D026082:Singlet Oxygen; D019001:Nitric Oxide Synthase
keywords: Actin; Decompression; Focal Adhesion Kinase; NADPH Oxidase; Nitrosative Stress; Reactive Nitrogen Species (RNS); Reactive Oxygen Species (ROS); S-Nitrosylation; Singlet Oxygen