Our preliminary research objectives is to elucidate how metals associated with residual oil fly ash (ROFA) initiates and exacerbates the inflammatory response in the lung. Our overall hypothesis is: Lung macrophages and epithelial cells in susceptible individuals respond disproportionately at a genetic level to metals in ROFA. "Priming" of lung cells by a pre-existing inflammatory condition results in synergistic amplification within the signal transduction pathways regulating gene expression. Specific aims are: 1) To determine the metal constituents of ROFA that mediate toxicity and its inflammatory potential; 2) To characterize the toxicity of ROFA-associated metals and their ability to induce pro-inflammatory cytokines, individually and in combination; 3) To establish molecular mechanisms responsible for increased cytokine expressions; 4) To establish a role for oxidative stress in metal-induced cytokine expression; and 5) To characterize the overall genetic response of macrophages and epithelial cells to metals. To determine the inflammatory potential of ROFA-associated metals, we will expose rats to soluble metals by intratracheal instillation and assess pulmonary inflammation and pro-inflammatory mediators by macrophages and epithelial cell lines and primary cell cultures in response to an in vitro challenge. Cells will be either macrophages and epithelial cell lines and primary cell cultures in response to an vitro challenge. Cells will be either normal or pre-treated ("primed") with bacterial lipopolysaccharide or tumor necrosis factor (TNF)-alpha, respectively. Signal transduction pathways regulating cytokine gene expression will be elucidated at a molecular level. Differential gene expression in primed and unprimed- macrophages and epithelial cells in response to metals will be compared and differentially-expressed genes identified. A role for an oxidant stress in the signal transduction pathway controlling regulation of these genes will be examined. We predict that metals capable of inducing a cellular respiratory burst will elicit enhanced production. of pro-inflammatory cytokines in primed lung epithelium and macrophages. This inappropriate response to ambient or low levels of metals would be an important first step in understanding the observed increase in morbidity/mortality associated with elevated airborne particle concentrations.