The Gillings Sampler – An electrostatic air sampler as an alternative method for aerosol in vitro exposure studies

Abstract

There is growing interest in studying the toxicity and health risk of exposure to multi-pollutant mixtures found in ambient air, and the U.S. Environmental Protection Agency (EPA) is moving towards setting standards for these types of mixtures. Additionally, the Health Effects Institute's strategic plan aims to develop and apply next-generation multi-pollutant approaches to understanding the health effects of air pollutants. There's increasing concern that conventional in vitro exposure methods are not adequate to meet EPA's strategic plan to demonstrate a direct link between air pollution and health effects. To meet the demand for new in vitro technology that better represents direct air-to-cell inhalation exposures, a new system that exposes cells at the air-liquid interface was developed. This new system, named the Gillings Sampler, is a modified two-stage electrostatic precipitator that provides a viable environment for cultured cells. Polystyrene latex spheres were used to determine deposition efficiencies (38-45%), while microscopy and imaging techniques were used to confirm uniform particle deposition. Negative control A549 cell exposures indicated the sampler can be operated for up to 4 hours without inducing any significant toxic effects on cells, as measured by lactate dehydrogenase (LDH) and interleukin-8 (IL-8). A novel positive aerosol control exposure method, consisting of a p-tolualdehyde (TOLALD) impregnated mineral oil aerosol (MOA), was developed to test this system. Exposures to the toxic MOA at a 1 ng/cm2 dose of TOLALD yielded a reproducible 1.4 and 2 fold increase in LDH and IL-8 mRNA levels over controls. This new system is intended to be used as an alternative research tool for aerosol in vitro exposure studies. While further testing and optimization is still required to produce a “commercially ready” system, it serves as a stepping-stone in the development of cost-effective in vitro technology that can be made accessible to researchers in the near future.

Assessment of biological responses of EpiAirway 3-D cell constructs versus A549 cells for determining toxicity of ambient air pollution

“…In ALI conditions, the apical surface of the cells is exposed to the air while the basolateral surface of the cells is fed with culture medium through a porous membrane (Akhtar et al, 2011, Maier et al, 2008), similar to what occurs in vivo . To conduct cell exposures at ALI conditions, new in vitro exposure technologies were developed both at research universities and commercially (Blank et al, 2006, de Bruijne et al, 2009, Aufderheide and Mohr, 1999, Aufderheide and Mohr, 2004, Lenz et al, 2009, Tippe et al, 2002, Cooney and Hickey, 2011, Aufderheide et al, 2013, Savi et al, 2008, Volckens et al, 2009, Ning et al, 2008, Zavala et al, 2014). …”

In vitro exposure to isoprene-derived secondary organic aerosol by direct deposition and its effects on <i>COX-2</i> and <i>IL-8</i> gene expression

“…Compared to exposure of cells in culture media to resuspended particles, direct particle deposition likely provides a more biologically relevant exposure model and enhances sensitivity of cells to air pollution particle exposures (Volckens et al, 2009;Lichtveld et al, 2012;Hawley et al, 2014a, b;Zavala et al, 2014;Hawley and Volckens, 2013). The Electrostatic Aerosol in Vitro Exposure System (EAVES) used in this study deposits particles, generated in our outdoor photochemical chamber, directly onto lung cells by electrostatic precipitation (de Bruijne et al, 2009).…”

“…In this study we chose to examine the gene expression levels of interleukin-8 (IL-8) and cyclooxygenase-2 (COX-2), not only for their links to inflammation and oxidative stress (Kunkel et al, 1991;Uchida, 2008) but also because both have been examined in previous studies using the EAVES for fresh and aged diesel exhaust (Lichtveld et al, 2012). Other studies on air pollution mixtures have also examined IL-8 as a biological endpoint due to its involvement with inflammation (Zavala et al, 2014;Ebersviller et al, 2012a, b;Doyle et al, 2004Doyle et al, , 2007. We compared the gene expression levels in cells exposed to SOA generated in an outdoor chamber from photochemical oxidation of isoprene in the presence of NO and acidified sulfate seed aerosol to cells exposed to a dark control mixture of isoprene, NO, and acidified sulfate seed aerosol to isolate the effects of the isoprene-derived SOA on the cells using the EAVES.…”

Cellular RNA is chemically modified by exposure to air pollution mixtures

“…significant cell death occurred), IL-8 and miR-10b measurements either did not change significantly or decreased. This is not surprising, since significant cell death can reduce the measurable levels of transcriptional and translational signals, and since downstream signals like IL-8 and miR-10b can take several hours after exposure to change significantly (Doyle et al, 2007;Rager et al, 2011a;Zavala et al, 2014). In stark contrast, trends in measured levels of 8OG were highly consistent, even as reflected by the linear additive effect in 8OG accumulation resulting from combining O 3 and unsaturated carbonyls.…”

“…Briefly, cells were grown on 30 mm Millicell membranes (EMD Millipore Corp., Billerica, MA) in F12-K media with 10% fetal bovine serum (FBS) plus 0.01% penicillin/streptomycin at a density of 850 000 cells/ membrane, 28 h prior to exposure as described (Jaspers et al, 1997;Zavala et al, 2014). Four hours before exposure, when cells reached $80% confluency, FBS-containing media was replaced with serum-free media (F12-K media + 1.5 mg/ml bovine serum albumin + 0.01% penicillin/streptomycin).…”