Kidney glomerular explants in serum-free media: demonstration of intracellular antioxidant enzymes and active oxygen metabolites.

Abstract

Guinea pig glomeruli were grown for 22 d in a serum-free medium composed of Waymouth's MB 752/l supplemented with sodium pyruvate, nonessential amino acids, and antibiotics (the basic medium). Intracellular cellular activity of the antioxidant enzymes superoxide dismutase (SOD; both copper-zinc [Cu,Zn] and manganese [Mn] forms) and catalase, and intracellular active oxygen metabolites (hydrogen peroxide [H2O2] and superoxide [O2-.]) were measured with time in culture. The results were compared to results obtained from glomeruli grown in different serum-free media, including the basic medium plus fibronectin (FN), the basic medium plus transferrin and FN, and a complex medium containing insulin, transferrin, selenium (Se), triiodothyronine, and FN (complete medium). In general, although the intracellular activity of antioxidant enzymes and active oxygen metabolites varied over time in culture in all media, there were only a few statistically significant differences among different media. Both CuZn SOD and Mn SOD activity were demonstrated in isolated glomeruli. The CuZn SOD activity per DNA ratio decreased slightly with time in culture in all media tested except the complete medium, in which CuZn SOD activity per DNA ratio remained more constant. The Mn SOD activity per DNA ratio did not vary significantly over time in culture. Catalaselike activity was very low in isolated glomeruli and declined sharply with time in culture in all media except the complete medium. Both H2O2 and O2-. were detected intracellularly in glomerular culture. Our results indicate that intracellular antioxidant enzymes and active oxygen metabolites in glomeruli vary with time in culture and, in some instances, with culture conditions.