Nitric oxide and superoxide radical production by human mononuclear leukocytes

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/10(6) cells and 0.25+/-0.02 nmol of superoxide anion (O2-)/min/10(6) 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/10(6) cells and 0.76+/-0.12 nmol of O2-/min/10(6) 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.