Oxidative stress-mediated epidermal growth factor receptor activation regulates PM_2.5-induced over-secretion of pro-inflammatory mediators from human bronchial epithelial cells

Highlights

• PM_2.5 exposure increased pro-inflammatory mediator expression in BEAS-2B cells.

• Exposure to PM_2.5 elevates intracellular levels of reactive oxygen species.

• Exposure to PM_2.5 induces phosphorylation of the EGFR.

• Oxidative stress is required for PM_2.5-induced EGFR activation.

Abstract

Background

Exposure to PM_2.5 has been associated with increased morbidity and mortality of lung diseases although the underlying mechanisms have not been fully uncovered. Airway inflammation is a critical event in the pathogenesis of lung diseases. This study aimed to examine the role of oxidative stress and epidermal growth factor receptor (EGFR) in PM_2.5-induced pro-inflammatory response in a human bronchial epithelial cell line, BEAS-2B.

Methods

BEAS-2B cells were exposed to 0, 20, 50, 100 and 150 μg/ml of PM_2.5. Secretion of pro-inflammatory mediators including interleukin-6 (IL-6), IL-8 and IL-1β was determined using enzyme linked immunosorbent assay. Levels of intracellular reactive oxygen species (ROS) were determined using flow cytometry. Phosphorylation of the EGFR was examined with immunoblotting.

Results

PM_2.5 exposure increased the secretion of IL-6, IL-8, and IL-1β in a concentration-dependent fashion. Moreover, exposure to PM_2.5 elevated intracellular levels of ROS, and phosphorylation of the EGFR (Y1068). Pretreatment of BEAS-2B cells with either an antioxidant or a specific EGFR inhibitor significantly reduced PM_2.5-induced IL-6, IL-8 and IL-1β secretion, implying that both oxidative stress and EGFR activation were involved in PM_2.5-induced pro-inflammatory response. Furthermore, pre-treatment of BEAS-2B cells with an antioxidant significantly blunted PM_2.5-induced EGFR activation, suggesting that oxidative stress was required for PM_2.5-induced EGFR activation.

Conclusion

PM_2.5 exposure induces pro-inflammatory response in human bronchial epithelial cells through oxidative stress-mediated EGFR activation.

Introduction

Air pollution is a complex mixture of heterogeneous components from a wide variety of sources, mainly consisting of particulate matter (PM) and gaseous pollutants. All components of air pollution are harmful to human health, but the most severe effects have been attributed to PM [1]. Though the air quality has been improving in recent years, China is still experiencing severe and persistent air pollution because of rapid urbanization and climate change. Fine PM pollution in winter time is still severe, especially in Northern and Central China [2]. Inhaled PM can induce inflammatory alterations and severe health consequences including pulmonary and cardiovascular effects [1,[3], [4], [5], [6]]. Recent assessment of health outcomes of fine PM has suggested that the fine particles may be more closely associated with adverse respiratory health effects than larger particles [[7], [8], [9]]. Acute and long-term exposure to higher concentrations of ambient airborne fine particles increased morbidity and mortality of pulmonary and cardiovascular diseases [10].

PM can be classified into the following categories based on aerodynamic diameter: coarse PM (2.5–10 μm, PM_2.5–10), fine PM (< 2.5 μm, PM_2.5), and ultrafine PM (< 0.1 μm, PM_0.1) [11]. The size of PM is directly linked to their potential for causing health problems. Small particles less than 10 μm in diameter pose the greatest problems, because they can get deep into lungs, and some may even get into bloodstream. PM_2.5 is capable of reaching alveoli and deposits deeper in the lung while PM₁₀ mostly stops at the upper bronchi [12]. Inhaled PM_2.5 can penetrate pulmonary surfactant and attack the underling pulmonary epithelial cells [13]. Accumulated evidence indicates airway epithelium lining the respiratory airways not only acts as a frontline barrier against PM but also is actively involved in innate and acquired immune responses as well as airway inflammation, the latter has been proposed to play a critical role in the pathogenesis of diverse lung diseases [14]. In a recent review [6], we have presented the evidence showing that short- or long-term exposure to PM_2.5 is associated with significant increases in hospitalizations, emergency department visits, or the incidence of inflammatory lung diseases including chronic obstructive pulmonary disease (COPD), asthma, and pneumonia, which is supported by the experimental results from toxicological studies with animal and cellular models [6].

Thus far, the exact mechanisms for PM_2.5-induced airway inflammation have not been fully elaborated. Previous studies have demonstrated that oxidative stress and epidermal growth factor receptor (EGFR) are involved in airway inflammation induced by environmental insults [[15], [16], [17], [18], [19]]. Oxidative stress refers to the excessive production of ROS (reactive oxygen species) in the cells and tissues and antioxidant system cannot be able to neutralize them. The role of oxidative stress in air pollution-induced adverse health effects including lung inflammation has been acknowledged [20,21]. Evidence shows that inhaled ambient PM adversely affects the bronchial epithelium through various mechanisms including the imposition of oxidative stress, which drives the transcription of pro-inflammatory mediators relevant to asthma and other inflammatory lung diseases [22]. EGFR is a single transmembrane protein that possesses intrinsic tyrosine kinase activity. It can be directly activated or trans-activated in response to a variety of stimuli [23,24]. EGFR signaling plays an important role in the pathogenesis of inflammation through regulation of pro-inflammatory genes [25,26]. The activation of the EGFR signaling in PM- and PM component-treated human bronchial epithelial cells has been detected [27,28]. In this study, we used pro-inflammatory mediators as the biomarker of airway inflammation and determined the role of oxidative stress and EGFR in ambient PM_2.5-induced pro-inflammatory response in human bronchial epithelial cells.