BLOOD LEUKOCYTE LUMINESCENCE BIO ASSAY: A EUKARYOTIC APPROACH TO STUDYING THE EFFECTS OF TOXIC CHEMICALS ON METABOLISM

摘要

The potential toxic effects of industrial chemicals, pharmaceuticals, foods, and pesticides on cellular metabolism have been predicted using bacterial and tissue culture-based methods of testing. These in vitro techniques have proven to be useful in evaluating the potential mutagenic and carcinogenic effects of various chemicals, and for investigating the mechanisms of their toxic action. However, manipulation of cells in these techniques can alter their functional states and thus lead to divergent results when compared to in vivo data. The objective of the present investigation was to design a method based on ex vivo human cell functional measurement that would be relatively easy to perform, would not require long-term culture, and would produce results that more closely approximate the effects of toxic chemicals on human cellular metabolism and immune function. Ethanol, a commonly imbibed agent with well-known physiologic and toxic effects, was chosen as the test chemical. The study focused on measuring human blood phagocyte metabolic activity using a few microliters of whole blood and a commercially available luminometer. The method allows for direct functional analysis of multiple phagocyte functions, including (1) basal and chemically-stimulated phagocyte oxidase activation, which in turn is dependent on glycolytic and hexose monophosphate shunt metabolism of glucose, (2) receptor-dependent phagocytosis and degranulation, and (3) myeloperoxidase-dependent activities, using appropriate combinations of chemiluminigenic substrates (CLS) and chemical/immune stimuli.