Urea-induced inducible nitric oxide synthase inhibition and macrophage proliferation

Atherosclerosis is a major cause of morbidity and mortality in chronic renal failure and is associated with the proliferation of macrophages within atherosclerotic lesions.

Because the progression of atherosclerosis as a consequence of decreased nitric oxide (NO) synthesis has been described, we investigated the correlation between the inhibition of inducible NO synthase (iNOS) by urea, macrophage proliferation as assayed by cell counting, tritiated-thymidine incorporation and measurement of cell protein, and macrophage apoptosis.

Urea induces a dose-dependent inhibition of inducible NO synthesis in lipopolysaccharide-stimulated mouse macrophages (RAW 264.7) with concomitant macrophage proliferation. Macrophage proliferation as determined by cell counting became statistically significant at 60 mmol/L urea corresponding to a blood urea nitrogen level of 180 mg/100 mL, concentrations seen in uremic patients. iNOS protein expression showed a dose-dependent reduction, as revealed by immunoblotting when cells were incubated with increasing amounts of urea. The decrease of cytosolic DNA fragments in stimulated macrophages incubated with urea shows that the proliferative actions of urea are associated with a decrease of diminished NO-mediated apoptosis.

These data demonstrate that inhibition of iNOS-dependent NO production caused by urea enhances macrophage proliferation as a consequence of diminished NO-mediated apoptosis. This fact may be important for the development of atherosclerotic lesions during chronic renal failure and is in accordance with recently published studies showing that under conditions with decreased constitutive NOS activity, iNOS might substitute the synthesis of NO. iNOS expression in vascular smooth muscle cells and macrophages is supposed to prevent restenosis following angioplasty or heart transplant vasculopathy. This is supported by the fact that specific inhibition of endogenous iNOS activity with L-N6-(1-iminoethly)-lysine accelerates the progression of vasculopathy in transplantation atherosclerosis.