Oxidative stress and cytokine expression in respiratory epithelial cells exposed to well-characterized aerosols from Kabul, Afghanistan

Abstract

In this study aerosol samples collected in an Asian mega-city (Kabul, Afghanistan) were compared to PM samples collected in a European location with traffic (Umeå, Sweden) and a reference urban dust material (SRM 1649b). The toxicity of each sample towards normal human bronchial epithelial (NHBE) cells and a human bronchial epithelial cell line (BEAS-2B) was tested along with their ability to induce reactive oxygen species (ROS) formation and inflammatory responses. The extracts’ morphology and elemental composition was studied by SEM-EDXRF, and filter samples were analyzed for metals and organic compounds. The PM from Kabul contained a larger fraction of fine particles, 19 times more polyaromatic hydrocarbons (PAH) and 37 times more oxygenated PAH (oxy-PAH) compared to samples from Umeå. The PM-samples from Kabul and the reference material (SRM 1649b) induced significantly stronger oxidative stress responses than the samples from Umeå. Furthermore, samples collected in Kabul induced significantly higher secretion of the cytokines IL-6, IL-8 and GM-CSF while SRM1649b induced a cytokine pattern more similar to samples collected in Umeå. Several properties of the particles could potentially explain these differences, including differences in their size distribution and contents of PAH and oxy-PAH, possibly in combination with their relative transition metal contents.

Introduction

Abatement programs have greatly reduced levels of traditional air pollutants in most European countries. However, many developing countries still have problematically high levels of species such as particulate matter (PM), ozone, NO₂, SO₂ and volatile organic compounds (Fenger, 1999, Hopke et al., 2008). Numerous studies have shown that air pollution is a very significant problem in Asian megacities such as Beijing, Shanghai, Karachi and New Delhi. We have recently demonstrated that Kabul in Afghanistan belongs to this group of cities since its air has high levels of PM and particle-borne polyaromatic hydrocarbons (PAHs) as well as oxygenated PAHs (oxy-PAH) and n-alkanes (Wingfors et al., 2011, Magnusson et al., 2012). PM from Kabul and Asian mega-cities differs from that found in modern western cities since a large proportion of the former consists of species produced by the incomplete combustion of coal and biomass used for domestic heating and food cooking. Additional factors that contribute to the abundance of compounds originating from partial combustion in Kabul include the use of low quality diesel and gas fuels, the relatively high proportion of vehicles using two-stroke engines with poor pollution characteristics, and diffuse burning. In contrast, ultrafine particles in modern western cities derive almost exclusively from traffic via vehicular exhausts and related processes such as wear on roads and brakes.

Ambient aerosols and PM are considered hazardous to humans. Epidemiological studies have shown that elevated levels of PM are associated with increased morbidity and mortality and that this association is especially pronounced for small-diameter PM (Valivanidis et al., 2008, Valivanidis et al., 2006, Pope et al., 2002). Over the last few decades, significant effort has been invested into finding toxicological evidence for a causal relationship between PM levels and health effects. In addition to the traditionally measured mass concentrations, the chemical composition, shape, size, surface area, number concentration and content of biological materials such as endotoxins in PM have also been considered in efforts to explain their toxicological and biological effects (Pope and Dockery, 2006). It has recently been reported that synergistic interactions between transition metals and oxy-PAHs such as quinones contribute to the adverse effects of PM. It is believed that inside cells, these species undergo redox cycling processes such as Fenton reactions that generate harmful reactive oxygen species (ROS) (Zhou et al., 2003, Xia et al., 2007, Guo et al., 2009). Because of their varied effects and the range of factors that influence them, it is important to characterize ambient aerosols comprehensively. However, it is not straightforward to assess the relative influence of distinct factors on the effects of ambient aerosols, and their relative contributions remain essentially unresolved.

The pulmonary epithelium is the first physical barrier against inhaled compounds and is also considered to be an integral component of the inflammatory defense response due to its production of numerous pro-inflammatory mediators following exposure to noxious compounds (Cunningham and Mahone, 2002, Takizawa et al., 1999). It has been suggested that the pro-inflammatory responses are, at least in part, driven by reactive oxygen species (ROS) and redox-sensitive transcription factors such as NF-κB and activator protein-1 (AP-1), which induce the activation of genes encoding pro-inflammatory cytokines such as neutrophil attractor, Il-8 and Il-6 following exposure to PM (Donaldson and Stone, 2003). Lung inflammation is believed to play a key role in a variety of health effects associated with PM exposure, including both pulmonary and cardiovascular pathologies (Salvi and Hogate, 1999, Frampton, 2006, HEI, 2002).

Cultured lung epithelial cells have been widely used to study the potential health effects of inhaling environmentally relevant PM, including ambient particulate matter (Ahktar et al., 2010, Becker et al., 2005, Salonen et al., 2004, Quay et al., 1998, Ortgiesen et al., 2000, Veronesi et al., 2002). Most of these studies have been performed using immortalized respiratory cell lines. However, it is also important to study primary cell cultures as these model systems more closely reproduce the conditions found in vivo.

PM from Asian mega-cities exhibit many of the unique properties that are believed to be responsible for some of the adverse health effects caused by exposure to ambient PM. For example, it contains a large proportion of ultrafine particles, has a large surface area, and is rich in associated organic compounds (including oxy-PAHs such as quinones) that originate from partial combustion in vehicle engines and incomplete combustion of coal and biomass. As such, it is a useful material for testing putative mechanisms of action in response to PM exposure.

The aim of the present study was to take advantage of previously published data on particle characteristics (Wingfors et al., 2011, Magnusson et al., 2012) to identify relationships between the physicochemical properties of the PM and the biological responses observed in respiratory epithelial cells. We compared the toxicity of and cellular responses to aerosol samples collected in Kabul, Afghanistan to those for samples from a small European location with traffic (Umeå, Sweden) and a standard urban dust reference material (SRM 1649b). These PM types were applied to normal human bronchial epithelial (NHBE) cells and cells from a human bronchial epithelial cell line (BEAS-2B). The cellular responses in each case were related to the chemical and physical properties of the aerosol samples as determined by GC–MS, ICP-MS and SEM–EDS analyses as well as optical methods.

The study involved collecting ambient PM into a liquid medium using a multistage impinger sampler. The collection efficiency of this apparatus was validated by performing control experiments in a test atmosphere.