Hydrogen peroxide release by rat peritoneal macrophages in the presence and absence of tumor cells

Abstract

The ability of mineral oil-elicited rat peritoneal macrophages to release hydrogen peroxide (H 2O 2) to the extracellular medium was measured in the presence and absence of rat lymphoma cells grown in tissue culture, and in the presence of phorbol myristate acetate (PMA). Horseradish peroxidase (HRP)-catalyzed oxidation of scopoletin or phenol red was used to measure H 2O 2 release during incubation of cells in monolayer culture for periods up to 24 h. Macrophages appeared to release H 2O 2 with or without PMA, although PMA greatly increased the amount of H 2O 2 released in short (1 to 4 h) incubations. Tumor cells did not replace PMA as a triggering agent for H 2O 2 release. Instead, tumor cells inhibited H 2O 2 release. The probable basis for inhibition was competition between macrophages and tumor cells for the supply of oxygen (O 2). Tumor cells did not inhibit H 2O 2 release when the O 2 concentration was held constant. The rates at which macrophages took up O 2 and released H 2O 2 were proportional to the O 2 concentration, as measured with the O 2 electrode. Rates of H 2O 2 release could be calculated from the difference in the rate constants for O 2 uptake measured in the presence of two different extracellular H 2O 2-consuming systems (HRP-scopoletin vs catalase). PMA-stimulated uptake of O 2 and release of H 2O 2 were highest in a small subpopulation of macrophages, obtained at the lowest-density position on gradients of bovine serum albumin. These cells also released H 2O 2 in the absence of PMA. Tumor cells had no effect on the rate constants for O 2 uptake and H 2O 2 release by the unfractionated macrophages or the macrophage subpopulations.