Use of a Sensitive Electro-Optical Method to Quantify Superoxide Release from Single PAM Exposed to Dusts in Vitro or In Vivo: Some Current Experimental and Model Results

This laboratory has developed a sensitive electro-optical method to quantify the initial rate (R) and total amount.(MAX) of superoxide (SO) produced by single pulmonary alveolar macrophages (PAM). The method uses a microscope to visualize PAM in culture, and to video record the images during the time the cells produce SO. MAX and R are calculated from measurement of temporal changes in optical density in the images due to precipitated diformazan formed by the reaction of SO, produced by each PAM, with nitroblue tetrazolium present in the culture medium. To date, values of Rand MAX, measured due to adherence of PAM to the dish, have been compared to values obtained when quartz, coal mine dust (CMD), and kaolin were added to the medium (in vitro). R and MAX have also been calculated for PAM lavaged from animals exposed to quartz and CMD in the WVU Inhalation Facility. Presently, experiments are being done using serum which permits PAM to be restimulated by different dusts to help establish a dose response effect, and a means to study the effects of lung surfactant on modifying the toxicity of inhaled dusts. Long term, this methodology should provide useful insight· into establishing the role SO plays in the phagocytosis of inhaled dusts and bacteria, in PAM dysfunction, and in lung diseases such as pneumoconioses. Supported by the Department of the Interior's Mineral Institute Program administered by the Bureau of Mines through the Generic Mineral Technology Center for Respirable Dust (G1135142).