Homocysteine induced impairment of nitric oxide-dependent vasorelaxation is reversible by the superoxide dismutase mimetic TEMPOL.
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 microM) were mounted on a myograph, pre-constricted with phenylephrine (10 microM) and responses to acetylcholine-dependent vasorelaxation examined. The incubations were repeated in the presence of indomethacin (10 microM), omega-nitro-L-arginine methyl ester (100 microM), L-arginine (100 microM), tetrahydrobiopterin (1 microM), catalase (1200 U/ml), ebselen, a peroxynitrite chelator (20 microM) 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 EC(50) (EC(50) 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 microM 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 omega-nitro-L-arginine methyl ester, l-arginine and tetrahydrobiopterin, catalase and ebselen had no effect on the EC(50) 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.
Hucks
Davy
D
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. d.hucks@qmul.ac.uk
Thuraisingham
Raj C
RC
Raftery
Martin J
MJ
Yaqoob
Magdi M
MM
D000818:Animals; D001682:Biological Availability; D003497:Cyclic N-Oxides / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000502:physiology; D005268:Femoral Vein; D016166:Free Radical Scavengers / Q000494:pharmacology*; D006710:Homocysteine / Q000494:pharmacology*; D008297:Male; D009569:Nitric Oxide / Q000502:physiology; D009924:Organ Culture Techniques; D051381:Rats; D011921:Rats, Inbred WKY; D013113:Spin Labels; D015854:Up-Regulation / Q000502:physiology; D014664:Vasodilation / Q000187:drug effects* / Q000502:physiology
D003497:Cyclic N-Oxides; D016166:Free Radical Scavengers; D013113:Spin Labels; D006710:Homocysteine; D009569:Nitric Oxide; C001803:tempol