Nephrol Dial Transplant (2004) 19: 1999-2005
Nephrol Dial Transplant Vol. 19 No. 8 © ERA-EDTA 2004; all rights reserved
Original Article
Homocysteine induced impairment of nitric oxide-dependent vasorelaxation is reversible by the superoxide dismutase mimetic TEMPOL
Experimental Medicine, Nephrology and Critical Care, William Harvey Research Institute, Charterhouse Square, St Bartholomew's and the Royal London Hospitals, Queen Mary, University of London, London EC1M 6BQ, UK
Correspondence and offprint requests to: Mr David Hucks, BSc, Experimental Medicine, Nephrology and Critical Care, William Harvey Research Institute, Charterhouse Square, St Bartholomew's and the Royal London Hospitals, Queen Mary, University of London, London EC1M 6BQ, UK. Email: d.hucks{at}qmul.ac.uk
Background. Elevated plasma homocysteine concentrations in renal patients are associated with accelerated cardiovascular disease. The mechanism(s) by which homocysteine acts remains unclear however, evidence implicates a role involving endothelial dysfunction.
Methods. Rat femoral arteries after acute or 4-h pre-incubation with racemic D,L-homocysteine (100 µM) were mounted on a myograph, pre-constricted with phenylephrine (10 µM) and responses to acetylcholine-dependent vasorelaxation examined. The incubations were repeated in the presence of indomethacin (10 µM), 
-nitro-L-arginine methyl ester (100 µM), L-arginine (100 µM), tetrahydrobiopterin (1 µM), catalase (1200 U/ml), ebselen, a peroxynitrite chelator (20 µM) and TEMPOL, a superoxide dismutase mimetic (1 mM). Results are shown as means±standard error, expressed as per cent relaxation to acetylcholine added (nmol/l).
Results. Increasing concentrations of homocysteine had no affect when added directly to basally relaxed or pre-constricted freshly isolated vessels. However, 4-h pre-incubation with or without homocysteine significantly shifted the acetylcholine EC50 (EC50 was defined as the concentration of acetylcholine that caused relaxation of the phenylephrine contracted tissue by 50%), control(4 h) = 74.7 nmol/l±10.5 vs 100 µM D,L-homocysteine(4 h) = 159.9 nmol/l±20.6; P<0.05) without affecting maximal relaxation. Response to endothelial independent relaxation was unaffected. Indomethacin, indomethacin and -nitro-L-arginine methyl ester, L-arginine and tetrahydrobiopterin, catalase and ebselen had no effect on the EC50 in homocysteine-exposed arteries. However, TEMPOL normalized vasorelaxation in homocysteine-treated arteries (75.2 nmol/l±14.6) but had no effect on the 4-h control group. Moreover, washing TEMPOL from the treated vessels restored endothelial dysfunction in D,L-homocysteine-treated vessels (163.9 nmol/l±34.1).
Conclusions. We conclude that homocysteine causes endothelial dysfunction by up-regulating a potential superoxide generating system resulting in reduced nitric oxide bio-availability.
Keywords: endothelial dysfunction; D,L-homocysteine; oxidation; TEMPOL
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