Neutrophils Generate Microparticles During Exposure to Inert Gases Due to Cytoskeletal Oxidative Stress [Molecular Bases of Disease]

May 27th, 2014 by Thom, S. R., Bhopale, V. M., Yang, M.

This investigation was to elucidate the mechanism for microparticle (MPs) formation triggered by exposures to high pressure inert gases. Human neutrophils generate MPs at a threshold of ~ 186 kPa with exposures of 30 minutes or more. Murine cells are similar but MPs production occurs at a slower rate and continues for ~ 4 hours whether cells remain under pressure, or not. 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). MPs production does not occur with neutrophils exposed to a NOX inhibitor (Nox2ds), 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 crosslinking 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 MPs production. These effects were also inhibited in cells exposed to ultraviolet light which photo-reverses S-nitrosylated cysteine residues and by co-incubations with the antioxidant ebselen or cytochalasin D. The auto-catalytic cycle of protein activation is initiated by inert gas-mediated singlet O2 production.