Recombinant arginine deiminase as a differential modulator of inducible (iNOS) and endothelial (eNOS) nitric oxide synthetase activity in cultured endothelial cells

Modulation of the extracellular level of arginine, substrate for nitric oxide synthetases, is a promising modality to alleviate certain pathological conditions where excess nitric oxide (NO) is produced. However, complications arise, as only preferential inhibition of the inducible nitric oxide synthetase (iNOS), but not endothelial nitric oxide synthetase (eNOS), is desired for the treatment of NO over-production. We investigated the effect of arginine deprivation mediated by a recombinant arginine deiminase (rADI) on the activity of iNOS and eNOS in an endothelial cell line, TR-BBB. Our results demonstrated that cytokine-induced NO production depends on the extracellular arginine as substrate. However, if sufficient citrulline is present in the medium, A23187-activated NO production by eNOS does not rely on extracellular arginine. Treatment with rADI can markedly inhibit cytokine-induced NO production via iNOS, but not A23187-activated NO production via eNOS. Our results also showed that the decrease of NO production by iNOS could be achieved by depleting arginine from the medium even under the conditions that would up-regulate iNOS expression. Thus, rADI appears to be a novel selective modulator of iNOS activity that may be a used as a tool in the study of pathological disorders where NO over-production plays a key role.