Toxicity of diesel engine exhausts in an in vitro model of lung slices in biphasic organotypic culture: induction of a proinflammatory and apoptotic response

Precision-cut rat lung slices in organotypic culture placed in a biphasic a ir/liquid system were used for this study. This model allowed pathological as well as cellular and molecular biology investigations to be carried out. Slices were exposed to a continuous flow of diluted diesel exhaust, with a PO2 adjusted to 20% to avoid hypoxia-induced effects. The exposure system allowed five exhaust concentrations from the same diesel engine to be studi ed concomitantly, and also allowed the impact of removing the particulate m atter using a filter cap on the exposure vials to be evaluated. Lung slices were exposed for 3 or 6 h to whole or filtered diesel exhaust. DNA integri ty was characterized by two different techniques: (1) an ELISA for the dete rmination of nucleosomes, and (2) the histochemical TUNEL method. By the TU NEL method, apoptotic cells were detected after a 6-h exposure followed by an incubation period of 18 h in a controlled atmosphere comprising 5% CO2/9 5% O-2. Under these conditions, apoptotic nuclei were more frequent in slic es exposed to diesel exhaust than in control slices. Cytokine production (t umor necrosis factor alpha, interleukin-1 beta) in the culture medium was m easured using an ELISA technique. After a 3-h exposure only TNF-alpha was d etected and increased in the culture medium of lung slices exposed to diese l exhaust. Under the same conditions, nucleosome levels in the slices incre ases in a dose-dependent way. In conclusion, whole diesel exhaust induced a n inflammatory response and DNA alterations which were reduced by filtratio n, thus indicating the important role of the particulate matter in diesel e xhaust.