Comparative toxicity of the horse eosinophil peroxidase-H2O2-halide system and granule basic proteins

Abstract

Stimulated eosinophils release cytotoxic granule constituents, including eosinophil peroxidase (EPO) and a group of granule basic proteins (GBP). EPO reacts with H2O2 formed by the respiratory burst and a halide to form cytotoxic oxidants. The relative potency of the EPO-H2O2-halide system and the GBP is considered here. Horse eosinophils were induced to degranulate, the degranulation products were separated by chromatography on Sephadex G-50 and comparable volumes of the column fractions were tested for toxicity to Escherichia coli and the schistosomula of Schistosoma mansoni in the presence and absence of H2O2 and halides. Both the EPO system and GBP were toxic. However, the peak EPO fraction could be diluted 1000-fold at pH 7.0 and 5000-fold at pH 5.0, and with a 10-fold dilution at pH 7.0 incubation time could be reduced to 5 s, with retention of bactericidal activity in the presence of H2O2 and halides, whereas the peak GBP fractions diluted 10-fold had a small bactericidal effect at 1 h which increased with prolongation of incubation to 24 h. A less than 1 log fall in E. coli viable cell count was produced by the GBP fractions under all conditions as compared to total destruction (greater than 5 log fall) with the EPO system. A 1000-fold dilution of the peak EPO fraction was schistosomulocidal in the presence of H2O2 and halides, with toxicity observed at 2 h with a 10-fold dilution. In contrast, no schistosomulocidal activity was observed at 18 h with a 10-fold dilution of the GBP fractions. However, toxicity was observed with a 5- or 50-fold increase in GBP concentration with maximum toxicity observed with fractions between the two major protein peaks. Thus, under the conditions employed, the EPO-H2O2-halide system contributed to a considerably greater degree to the toxic activity of the granule components than did the GBP.