In vivo and in vitro correlation of pulmonary map kinase activation following metallic exposure

Residual oil fly ash (ROFA) is a particulate pollutant produced in the comb ustion of fuel oil. Exposure to ROFA is associated with adverse respiratory effects in humans, induces lung inflammation in animals, and induces infla mmatory mediator expression in cultured human airway epithelial cells (HAEC ). ROFA has a high content of transition metals, including vanadium, a pote nt tyrosine phosphatase inhibitor that we have previously shown to disregul ate phosphotyrosine metabolism and activate mitogen-activated protein kinas e (MAPK) signaling cascades in HAEC. In order to study MAPK activation in r esponse to in vivo metal exposure, we used immunohistochemical methods to d etect levels of phosphorylated protein tyrosines (P-Tyr) and the MAPKs ERK1 /2, JNK, and P38 in lung sections from rats intratracheally exposed to ROFA . After a 1-h exposure to 500 mu g ROFA, rat lungs showed no histological c hanges and no significant increases in immunostaining for either P-Tyr or p hospho-(P-) MAPKs compared to saline-instilled controls. At 4 h of exposure , there was mild and variable inflammation in the lung, which was accompani ed by an increase in specific immunostaining for P-Tyr and P-MAPKs in airwa y and alveolar epithelial cells and resident macrophages. By 24 h of exposu re, there was a pronounced inflammatory response to ROFA instillation and a marked increase in levels of P-Tyr and P-MAPKs present within the alveolar epithelium and in the inflammatory cells, while the airway epithelium show ed a continued increase in the expression of P-ERK1/2. By comparison, HAEC cultures exposed to 100 mu g/ml ROFA for 20 min resulted in marked increase s in P-Tyr and P-MAPKs, which persisted after 24 h of exposure. P-Tyr level s continued to accumulate for up to 24 h in HAEC exposed to ROFA. These res ults demonstrate in vivo activation in cell signaling pathways in response to pulmonary exposure to particulate matter, and support the relevance of i n vitro studies in the identification of mechanisms of lung injury induced by pollutant inhalation.