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Free Radic Biol Med. 2017 Jul;108:334-344. doi: 10.1016/j.freeradbiomed.2017.03.026. Epub 2017 Mar 28.

Cigarette smoke extract induced exosome release is mediated by depletion of exofacial thiols and can be inhibited by thiol-antioxidants.

Author information

1
Department of Medical Microbiology, Maastricht University Medical Center, PO Box 5800, 6202 AZ Maastricht, The Netherlands; Department of Respiratory Medicine, Maastricht University Medical Center, PO Box 5800, 6202 AZ Maastricht, The Netherlands. Electronic address: b.benedikter@maastrichtuniversity.nl.
2
Department of Medical Microbiology, Maastricht University Medical Center, PO Box 5800, 6202 AZ Maastricht, The Netherlands. Electronic address: c.volgers@maastrichtuniversity.nl.
3
Department of Medical Microbiology, Maastricht University Medical Center, PO Box 5800, 6202 AZ Maastricht, The Netherlands. Electronic address: pascallevaneijck@gmail.com.
4
Department of Respiratory Medicine, Maastricht University Medical Center, PO Box 5800, 6202 AZ Maastricht, The Netherlands. Electronic address: e.wouters@mumc.nl.
5
Department of Medical Microbiology, Maastricht University Medical Center, PO Box 5800, 6202 AZ Maastricht, The Netherlands; Department of Medical Microbiology & Infection Control, VU University Medical Center, Van der Boechorststraat 7, 1081BT Amsterdam, The Netherlands. Electronic address: paul.savelkoul@mumc.nl.
6
Department of Respiratory Medicine, Maastricht University Medical Center, PO Box 5800, 6202 AZ Maastricht, The Netherlands. Electronic address: n.reynaert@maastrichtuniversity.nl.
7
Department of Pharmacology and Toxicology, Maastricht University, PO Box 616, 6200 MD Maastricht, The Netherlands. Electronic address: g.haenen@maastrichtuniversity.nl.
8
Department of Respiratory Medicine, Maastricht University Medical Center, PO Box 5800, 6202 AZ Maastricht, The Netherlands. Electronic address: g.rohde@mumc.nl.
9
Department of Pharmacology and Toxicology, Maastricht University, PO Box 616, 6200 MD Maastricht, The Netherlands. Electronic address: a.weseler@maastrichtuniversity.nl.
10
Department of Medical Microbiology, Maastricht University Medical Center, PO Box 5800, 6202 AZ Maastricht, The Netherlands. Electronic address: f.stassen@maastrichtuniversity.nl.

Abstract

INTRODUCTION:

Airway epithelial cells have been described to release extracellular vesicles (EVs) with pathological properties when exposed to cigarette smoke extract (CSE). As CSE causes oxidative stress, we investigated whether its oxidative components are responsible for inducing EV release and whether this could be prevented using the thiol antioxidants N-acetyl-l-cysteine (NAC) or glutathione (GSH).

METHODS:

BEAS-2B cells were exposed for 24h to CSE, H2O2, acrolein, 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB), bacitracin, rutin or the anti-protein disulfide isomerase (PDI) antibody clone RL90; with or without NAC or GSH. EVs in media were measured using CD63+CD81+ bead-coupled flow cytometry or tunable resistive pulse sensing (TRPS). For characterization by Western Blotting, cryo-transmission electron microscopy and TRPS, EVs were isolated using ultracentrifugation. Glutathione disulfide and GSH in cells were assessed by a GSH reductase cycling assay, and exofacial thiols using Flow cytometry.

RESULTS:

CSE augmented the release of the EV subtype exosomes, which could be prevented by scavenging thiol-reactive components using NAC or GSH. Among thiol-reactive CSE components, H2O2 had no effect on exosome release, whereas acrolein imitated the NAC-reversible exosome induction. The exosome induction by CSE and acrolein was paralleled by depletion of cell surface thiols. Membrane impermeable thiol blocking agents, but not specific inhibitors of the exofacially located thiol-dependent enzyme PDI, stimulated exosome release.

SUMMARY/CONCLUSION:

Thiol-reactive compounds like acrolein account for CSE-induced exosome release by reacting with cell surface thiols. As acrolein is produced endogenously during inflammation, it may influence exosome release not only in smokers, but also in ex-smokers with chronic obstructive pulmonary disease. NAC and GSH prevent acrolein- and CSE-induced exosome release, which may contribute to the clinical benefits of NAC treatment.

KEYWORDS:

Acrolein; Airway epithelial cells; COPD; Chronic inflammation; DTNB; EV; Extracellular vesicles; Lung; Oxidative stress

[Indexed for MEDLINE]

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