A role for sulfhydryl groups in granulocyte aggregation

Polymorphonuclear leukocytes (PMN) stimulated by high concentrations of the complement component C5A form cellular aggregates, and both the rate and degree of aggregation are influenced by changes in the PMN plasma membrane and cytoskeleton. Since sulfhydryls are important constituents of the plasma membrane and cytoskeleton, we investigated the effect of agents that oxidize and bind sulfhydryls on C5A-induced aggregation. PMN incubated with diamide, a nonspecific sulfhydryl- oxidizing agent, had a marked increase in their aggregation response to C5A. Tertiary butyl hydroperoxide (BHP), which reacts specifically with the soluble sulfhydryl glutathione (GSH), had no effect on aggregation. The enhancement of PMN aggregation by diamide, but not BHP, suggested that oxidation of non-GSH sulfhydryls contributes to the aggregation response. To test the requirement for sulfhydryls in PMN aggregation, PMN were treated with the sulfhydryl-binding agent N-ethylmaleimide (NEM). NEM markedly impaired aggregation without affecting resting or methylene blue-stimulated [14C]-L-glucose oxidation of the granulocytes. P-chloromercuriphenyl sulfonic acid (PCMPSA), an external sulfhydryl-binding agent, had no effect on aggregation. These studies suggest that cellular sulfhydryls are required for optimal PMN aggregation and that oxidation of these sulfhydryls may be one of the biochemical changes that contributes to aggregation.