IRON, CALCIUM AND OXIDATIVE STRESS IN LUNG INJURY

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Striking differences in the prevalence of coal workers' pneumoconiosis and chronic obstructive bronchopneumopathy exist between different coal mine regions despite comparable levels of dust exposure. The goal of this proposal is to test the hypothesis that bio-available Fe2+ is the active component in coal dust-induced cell injury through oxidative stress pathway, and that calcite (CaCO) may play a protective role in such injury by inhibiting solubilization of Fe2+, making Fe2+ less bio- available. Primary cultures of guinea pig alveolar macrophages (AMs) and human lung type II cells (A549) will be used for coal treatments. Total of thirty coal samples from three coal mine regions with a CWP prevalence of 4 percent (Utah), 10 percent (West Virginia), and 26 percent (Pennsylvania), respectively, will be tested. The bio-availability of Fe2+ and Ca2+, as well as other trace metals (i.e. As, Cd, Ni, Zn, Co, Cu) and quartz released from coal dusts in these cells will be determined. Ferritin and metallothionein will be measured as indices of the bio-availability of Fe2+ and trace metals in these cells. Cellular Fe2+ and Ca2+ levels will also be determined using colorimetric methods. Quartz content in the cells will be quantitated using X-ray diffraction after low temperature ashing of cell matrix. Lipid peroxidation of cell membranes, H2O2 formation, and AP-1 expression will be measured before and after treatment with coals. The levels of tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), and fibronectin will also be determined as a part of this study. Using TNF-alpha, IL-1, and other biochemical parameters as independent variables, the independent effect of bio-available Fe2+, trace metals, and quartz will be identified by multiple regression analyses. A pre-determined percentage of calcite will be added to the Pennsylvania and West Virginia coals before cell treatment. AMs and A549 cells will also be pretreated with Ca2+ before exposure to the untreated coals. The above mentioned end-points will be measured and effects of treatments assessed. We expect that, by confirming the role of Fe2+ in coal-induced injury, we will be able to predict which coal is likely to be more toxic, even before mining. By determining the role of calcite, it is possible to propose alternative methods for lung disease prevention.