Nrf2 positively regulates autophagy antioxidant response in human bronchial epithelial cells exposed to diesel exhaust particles

Abstract

Diesel exhaust particles (DEP) are known to generate reactive oxygen species in the respiratory system, triggering cells to activate antioxidant defence mechanisms, such as Keap1-Nrf2 signalling and autophagy. The aim of this study was to investigate the relationship between the Keap1-Nrf2 signalling and autophagy pathways after DEP exposure. BEAS-2B cells were transfected with silencing RNA (siRNA) specific to Nrf2 and exposed to DEP. The relative levels of mRNA for Nrf2, NQO1, HO-1, LC3B, p62 and Atg5 were determined using RT-PCR, while the levels of LCB3, Nrf2, and p62 protein were determined using Western blotting. The autophagy inhibitor bafilomycin caused a significant decrease in the production of Nrf2, HO-1 and NQO1 compared to DEPs treatment, whereas the Nrf2 activator sulforaphane increased the LC3B (p = 0.020) levels. BEAS-2B cells exposed to DEP at a concentration of 50 μg/mL for 2 h showed a significant increase in the expression of LC3B (p = 0.001), p62 (p = 0.008), Nrf2 (p = 0.003), HO-1 (p = 0.001) and NQO1 (p = 0.015) genes compared to control. In siRNAtransfected cells, the LC3B (p < 0.001), p62 (p = 0.001) and Atg5 (p = 0.024) mRNA levels and the p62 and LC3II protein levels were decreased, indicating that Nrf2 modulated the expression of autophagy markers (R < 1). These results imply that, in bronchial cells exposed to DEP, the Nrf2 system positively regulates autophagy to maintain cellular homeostasis.Chronic exposure to air pollution has been associated with adverse effects on the health of individuals 1 , such as inflammation 2-4 and mucociliary clearance dysfunction 5 of the respiratory system. Air pollution particulate matter (PM) induces oxidative stress in airway tissues, causing living organisms to activate their defences to prevent cell death 6-8 . One mechanism that cells use for defence against oxidative stress is autophagy, which is a homeostatic process that reduces cytoplasmic volume by degrading damaged organelles and proteins through a lysosome-dependent degradation process, and new organelles and proteins are synthesized as replacements 9-11 . Previous studies have emphasized that exposure to PM induces the generation of reactive oxygen species (ROS) and increases the levels of autophagy and cell death [12][13][14] . Some of the most harmful components of urban PM are derived from diesel exhaust particles (DEP) 15,16 . Usually, composed of carbon nuclei, DEP have large surface areas that can adsorb other chemicals in the www.nature.com/scientificreports www.nature.com/scientificreports/ environment, such as polycyclic aromatic hydrocarbons (PAHs), sulphate, nitrate, metals, carbon monoxide, aldehydes and various low molecular weight hydrocarbons 17,18 . PAHs and their derivatives are especially important because they are able to generate ROS in tissues 19 .Genes encoding key antioxidant enzymes are important for maintaining intracellular redox homeostasis through the antioxidant response element (ARE) found within the promoter regions of these genes 20 . The transcription fact...

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“…Our findings are consistent with those of several reports [33,34]. In addition, AP-SD dose dependently increased p62 expression in Nrf2 WT mice, not in Nrf2KO mice (Figure 11(b)).…”

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“…Autophagy is a cellular mechanism for the and degradation of intracellular pathogens and compromised organelles, and also clears other cellular components, such as inflammasomes and pro-inflammatory cytokines. In fact, DEP has been reported to activate autophagy in BEAS-2B cells [27]. These findings tempted us to speculate that autophagy is implicated in DEP-induced inflammation.…”

“…DEP has been widely reported to induce bronchial epithelial cell injury by regulation of autophagy, inflammation, oxidative stress, and apoptosis [27][28][29][30].…”