Regulation of nitric oxide synthesis in uraemia

Nitric oxide (NO) is a cell-to-cell mediator involved in theregulation of vascular tone and in the mechanisms of host defence.Since uraemic syndrome is characterized by abnormalities inblood pressure and flow and by impairment of white cell function,we studied the regulation of nitric oxide synthase (NOS) activityby uraemic plasma. We used three different cellular types havingdifferent levels of NOS activity: tEnd. 1 murine endothelialcell line transformed by mT oncogene of polyomavirus had a highNOS activityand expressed endothelial-NOS (eNOS) and inducible-NOS(iNOS) isoforms; human endothelial cells from cord umbilicalvein (HUVEC) had low enzymatic activity and expressed only eNOS;finally, J774 murine macrophage line was characterised by iNOSinduced after treatment with cytokines. We demonstrated thatmost (79%) of end-stage uraemic plasma studied inhibited NOSactivity in tEnd.1 and in cytokine induced -J774, whereas theywere ineffective on HUVEC. Twenty percent of plasma samples(14 of 67) activated NOS activity in tEnd.1 and in J774 cells,but not in HUVEC, suggesting the presence of molecule(s) whichinfluence iNOS. The effect of plasma was not dependent on thetype of haemodialysis treatment. A great number of plasmas frompatients with moderate renal failure also inhibited NOS activityin tEnd.1, suggesting that the accumulation of molecules affectingNOS was caused by the renal failure rather than the haemodialytictreatment. However, the haemodialysis modified the effect of plasmas onNOS activity. Plasma taken after haemodialysis session showeda reduced inhibitory activity in tEnd.1 and in some cases itenhanced NOS activity. Simultaneously, molecules reducing NOSactivity accumulated in the ultrafiltrate. The plasma concentrationof N^G-N^G dimethyl-L-arginine (asymmetrical dimethylarginine,ADMA), an inhibitor of NOS, increased in end-stage uraemic patientsand was reduced by haemodialysis. However, the concentrationsreached in uraemic plasmas were lower than the ADMA ICM₅₀ ontEnd.1 NOS, indicating that this compound contributes with othermolecules to the inhibitory effect of uraemic plasma. Haemodialysisreduced also the enhanced effect exerted by some plasmas onNOS in J774. Therefore, the effect of endstage uraemic plasmaon NOS activity derive from the balance between inhibitors andactivators.