Analysis of the effects of nitric oxide and oxygen on nitric oxide production by macrophages

The interactions between NO and O2 in activated macrophages were analysed by incorporating previous cell culture and enzyme kinetic results into a novel reaction–diffusion model for plate cultures. The kinetic factors considered were: (i) the effect of O2 on NO production by inducible NO synthase (iNOS); (ii) the effect of NO on NO synthesis by iNOS; (iii) the effect of NO on respiratory and other O2 consumption; and (iv) the effects of NO and O2 on NO consumption by a possible NO dioxygenase (NOD). Published data obtained by varying the liquid depth in macrophage cultures provided a revealing test of the model, because varying the depth should perturb both the O2 and the NO concentrations at the level of the cells. The model predicted that the rate of NO2− production should be nearly constant, and that the net rate of NO production should decline sharply with increases in liquid depth, in excellent agreement with the experimental findings. In further agreement with available results for macrophage cultures, the model predicted that net NO synthesis should be more sensitive to liquid depth than to the O2 concentration in the headspace. The main reason for the decrease in NO production with increasing liquid depth was the modulation of NO synthesis by NO, with O2 availability playing only a minor role. The model suggests that it is the ability of iNOS to consume NO, as well as to synthesize it, that creates very sensitive feedback control, setting an upper bound on the NO concentration of ∼1 μM. The effect of NO consumption by other possible pathways (e.g., NOD) would be similar to that of iNOS, in that it would help limit net NO production. The O2 utilized during enzymatic NO consumption is predicted to make the O2 demands of activated macrophages much larger than those of unactivated ones (where iNOS is absent); this remains to be tested experimentally.