METAL ENRICHED QUASI-ULTRAFINE PARTICLES FROM GAS METAL ARC WELDING- STAINLESS STEEL INDUCED GENETIC AND EPIGENETIC ALTERATIONS IN BEAS-2B CELLS

International audience

English. Purpose: Worldwide, an estimated 11 million workers have a job title of welder and around 110 million additional workers probably incur welding-related exposures. Recent evidence has identified welding fumes (WF)-derived ultrafine particles (UFP) as the driving force of their adverse health effects. However, UFP have not yet been extensively studied and are currently not included in present air quality standards/guidelines. Here, attention was focused on the underlying genetic and epigenetic mechanisms by which the quasi-UFP (Q-UFP, i.e., ≤ 0.25 µm) of the WF emitted by gas metal arc welding-stainless steel (GMAW-SS) exert their toxicity in human bronchial epithelial cells (BEAS-2B).Methods: Q-UFP were produced by GMAW-SS by an automatic welding bench and collected by a cascade impactor. Their size distribution and zeta potential were studied by Zetasizer Nano ZSP™, and their metal composition by inductively coupled plasma-mass spectrometry. BEAS-2B cells (n =5) were exposed acutely (24 h) or repeatedly (3 x 24 h) to Q-UFP at concentrations ranging from 0.1 to 50 µg/cm² to study their cytotoxicity (ATP, luminescence). Thereafter, BEAS-2B cells were exposed to Q-UFP at 1.5 and 9 µg/cm² for acute exposure, and 0.25 and 1.5 µg/cm² for repeated exposure to study the other toxicological endpoints. Oxidative stress and inflammation were studied by nuclear factor erythroid 2-related factor 2 (NRF2) and nuclear factor-kappa B (NFB) signaling pathway activation, glutathione status, oxidative damage (i.e., 8-hydroxy-2’-deoxyguanosine, carbonylated protein, and 4-hydroxynonenal), and cytokine secretion (i.e., tumor necrosis factor-alpha, interleukin-1 beta, and interleukin-6) by TransAM, RT-qPCR, and ELISA/MSD/Luminex. Primary DNA damages were studied by in vitro comet assay. Histone H3 acetylation (i.e., H3K9ac and H3K27ac), histone acetyltransferase (HAT) and deacetylase (HDAC) activities, and miRNA profiles were studied by ELISA and TaqMan® OpenArray® Human MicroRNA Panel. Other cell signaling pathways were studied by TaqMan® OpenArray® Human Signal Transduction Panel. Student t-test was used to look at statistical differences versus controls (p < 0.05).Results: Q-UFP showed a monomodal size distribution in number centered on 104.4 ± 52.3 nm and a zeta potential of 13.8 ± 0.3 mV. They were enriched in Fe > Cr, and also CrVI > Mn > Si. Dose-dependent activation of NRF2 and NFB signaling pathways, glutathione alteration, and DNA, protein and lipid oxidative damages were reported in BEAS-2B cells acutely, and to a lesser extent, repeatedly exposed (p < 0.05). However, no cytokine secretion nor primary DNA damage were seen. Histone H3 acetylation and HDAC/HAT activity decreases were reported for any exposure (p < 0.05). Differentially regulated miRNA and mRNA (FC < 0.66 or > 1.5, p < 0.05) closely indicated the activation of critical cell signaling pathways related to oxidative stress, inflammation, and cell cycle deregulation towards apoptosis.