A Kinetic Model of Superoxide Production from Single Pulmonary Alveolar Macrophages

A kinetic model was developed to describe the production of superoxide (02) by single pulmonary alveolar macrophages (PAM). Model predictions were compared to experimental results obtained from single rat PAM. The O2 - was quantified by measuring the reduction of nitroblue tetrazolium (NBT) to a diformazan precipitate (NBTH2) from video-recorded images of individual cells. The kinetic model considered three reactions: (1) the production of extracellular 0 - from the reduction of oxygen by NADPH oxidase using intracellular NADPH as the substrate, (2) the subsequent dismutation of o2- to from O2 , and (3) the reaction of o2- and NBT. NBT specificity for o2- was analyzed by comparing results in the presence and absence of superoxide dismutase (SOD) which catalyzes the dismutation of o2- to H2o2. Measured PAM heterogeneity was accounted for by varying the concentration of intracellular NADPH, its rate of depletion, and the concentration of intracellular NADPH oxidase in the kinetic model. Model predictions compared favorably with experimental results except when SOD was present. This discrepancy may be due to diffusional limitations since NBT is a relatively small molecule (818 MW) compared to SOD (34000 MW). In addition, the cell surface is both ruffled and negatively charged, which may introduce steric hindrances and/or electrostatic effects since SOD is also negatively charged.