METAL SPECIATION--EFFECT ON TRANSPORT AND INTERACTIVE TOXICITY

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It is generally recognized that metal ions do not circulate freely in biological fluids, but are bound to endogenous ligands (e.g., proteins, amino acids, etc.) in blood and cells. The metal ion may be the ultimate toxin but the dose-response relationship and/or the access into the cell is undoubtedly controlled by the form of the metal in the blood or bathing medium. The goal of this project is to establish a relationship between metal transport and metal toxicity using the chemical species that occur in blood. The hypotheses of this project are as follows: I. In the systemic circulation, metals will exist primarily as metal complexes with endogenous ligands and not as the free ions. II. The rate of metal uptake and the extent of toxicity in in vitro systems is dependent on the actual metal species which are found in blood. This project will focus on thiol binding ligands (cysteine, glutathione, albumin, metallothionein) and will examine the effect of metal complexes on cellular transport and toxicity. Metal ions which will be examined belong to Group IIB of the Periodic Table: Cd(II), Hg(II), and Zn(II). The specific aims include the following: 1). Determine an internally consistent set of values for the metal-ligand formation constants, ligand concentrations in various body compartments (including blood, extra- and intracellular fluids), and total metal concentrations found in various body compartments. Use solution equilibria calculations found on standard computer programs to predict the most prevalent metal species that should be found in the various body compartments. 2). Metal-Protein exchange reactions will be studied using recently developed chromatographic techniques. The thermodynamics, and possibly the kinetics, of the exchange reaction can be determined by the elution and/or location of the metal ion as a function of time. 3). Examine the uptake and toxicity of the metal ion complexes by using in vitro preparations of isolated cell populations in culture. The cell populations will include erythrocytes and erythrocyte ghosts; LLC-PK1 kidney cells and primary proximal kidney tubule preparations; and ARL-18 (liver) epithelial cells and primary liver hepatocyte preparations.