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

• Lung cells were successfully exposed at the air–liquid interface to diesel exhaust.
• Cells showed a decreased cell viability and increased oxidative stress response.
• Diesel exhaust was obtained from a heavy-duty truck in a climatic–altitude chamber.
• This chamber offers unique capability to measure real world emission of a truck.

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 1.5 h. The truck was tested at a speed of ∼70 km h−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.