Nitric Oxide and Superoxide Radical Production by Human Mononuclear Leukocytes

Abstract

Human mononuclear cells (90% lymphocytes, 9% monocytes, and 1% polymorphonuclear leukocytes) produced spontaneously in resting state 0.11 ± 0.01 nmol of nitric oxide (NO)/min/106 cells and 0.25 ± 0.02 nmol of superoxide anion (O₂ ^-)/min/106 cells, as primary products. When these cells were stimulated with phorbol 12-myristate 13-acetate (PMA), the NO and O₂ ^- production increased by 82% and 204% to 0.25 ± 0.02 nmol of NO/min/106 cells and 0.76 ± 0.12 nmol of O₂ ^-/min/106 cells, respectively. Oxygen uptake reasonably accounted for the sum of the rates of NO and hydrogen peroxide (H₂O₂), the latter calculated as 0.5 O₂ ^- production, in nonstimulated and in PMA-stimulated cells. H₂O₂ and peroxynitrite formation were detected and measured as secondary products of the primary products O₂ ^- and NO. An original assay to determine H₂O₂ steady-state concentration and production rates is described. The determined production rates of the involved reactive species are in good agreement with known chemical equations. It is apparent that NO and O₂ ^- production by human mononuclear cells may constitute the basis of intercellular signaling and cell toxicity.