REGULATION OF NITRIC-OXIDE SYNTHESIS IN UREMIA

Nitric oxide (NO) is a cell-to-cell mediator involved in the regulatio n of vascular tone and in the mechanisms of host defence. Since uraemi c syndrome is characterized by abnormalities in blood pressure and flo w and by impairment of white cell, function, we studied the regulation of nitric oxide synthase (NOS) activity by uraemic plasma. We used th ree different cellular types having different levels of NOS activity: tEnd.1 murine endothelial cell line transformed by mT oncogene of poly omavirus had a high NOS activity and expressed endothelial-NOS (eNOS) and inducible-NOS (iNOS) isoforms; human endothelial cells from cord u mbilical vein (HUVEC) had low enzymatic activity and expressed only eN OS; finally, J774 murine macrophage line was characterised by iNOS ind uced after treatment with cytokines. We demonstrated that most (79%) o f end-stage uraemic plasma studied inhibited NOS activity in tEnd.1 an d in cytokine induced -J774, whereas they were ineffective on HUVEC. T wenty percent of plasma samples (14 of 67) activated NOS activity in t End.1 and in J774 cells, but not in HUVEC, suggesting the presence of molecule(s) which influence iNOS. The effect of plasma was not depende nt on the type of haemodialysis treatment. A great number of plasmas f rom patients with moderate renal failure also inhibited NOS activity i n tEnd.1, suggesting that the accumulation of molecules affecting NOS was caused by the renal failure rather than the haemodialytic treatmen t. However, the haemodialysis modified the effect of plasmas on NOS ac tivity. Plasma taken after haemodialysis session showed a reduced inhi bitory activity in tEnd.1 and in some cases it enhanced NOS activity. Simultaneously, molecules reducing NOS activity accumulated in the ult rafiltrate. The plasma concentration of N-G-N-G dimethyl-L-arginine (a symmetrical dimethylarginine, ADMA), an inhibitor of NOS, increased in end-stage uraemic patients and was reduced by haemodialysis. However, the concentrations reached in uraemic plasmas were fewer than the ADM A IC50 on tEnd.1 NOS, indicating that this compound contributes with o ther molecules to the inhibitory effect of uraemic plasma. Haemodialys is reduced also the enhanced effect exerted by some plasmas on NOS in J774. Therefore, the effect of endstage uraemic plasma on NOS activity derive from the balance between inhibitors and activators.