Alveolar epithelial cells (A549) exposed at the air-liquid interface to diesel exhaust: First study in TNO's powertrain test center.

Air-liquid interface (ALI) exposures enable in vitro testing of mixtures of gases and particles such as diesel exhaust (DE). The main objective of this study was to investigate the feasibility of exposing human lung epithelial cells at the ALI to complete DE generated by a heavy-duty truck in the state-of-the-art TNO powertrain test center. A549 cells were exposed at the air-liquid interface to DE generated by a heavy-duty Euro III truck for 1.5h. The truck was tested at a speed of ∼70kmh(-1) to simulate free-flowing traffic on a motorway. Twenty-four hours after exposure, cells were analyzed for markers of oxidative stress (GSH and HO-1), cytotoxicity (LDH and Alamar Blue assay) and inflammation (IL-8). DE exposure resulted in an increased oxidative stress response (significantly increased HO-1 levels and significantly reduced GSH/GSSH ratio), and a decreased cell viability (significantly decreased Alamar Blue levels and slightly increased LDH levels). However, the pro-inflammatory response seemed to decrease (decrease in IL-8). The results presented here demonstrate that we are able to successfully expose A549 cells at ALI to complete DE generated by a heavy-duty truck in TNO's powertrain test center and show oxidative stress and cytotoxicity responses due to DE exposure.

Toxic effects following phosgene exposure of human epithelial lung cells in vitro using a CULTEX® system.

The aim of the present study was to investigate toxic effects following phosgene exposure of human epithelial lung cells (A549) in vitro using a CULTEX® system. In particular, toxic effects regarding early biomarkers emerging during the latency period following exposure might be of great value for medical treatment. Cells cultured on semi-permeable membranes were directly exposed at the liquid-air interface to different concentrations of phosgene, or dry medical air. Cell membrane integrity (leakage of LDH), metabolic activity (reduction of Alamar Blue), oxidative damage (GSH, and HO-1, in cell lysates), and release of IL-8, were studied. For most of the above-mentioned biological end-point markers, significant changes could be assessed following a 20 min exposure to 1.0 ppm and 2.0 ppm phosgene. Moreover, except for IL-8, all biological marker profiles showed to be in line with results obtained by others in animal studies. The C×t value of 40 ppm min appeared to be constant. The overall results suggest that at 4 h post-exposure a maximal level of toxicity was achieved. Our results demonstrate the suitability of a CULTEX® system to detect toxic effects induced by phosgene on human epithelial lung cells, which may contribute to the discovery of early biomarkers for new medical countermeasures.