Homocysteine-induced endothelin-1 release is dependent on hyperglycaemia and reactive oxygen species production in bovine aortic endothelial cells.
BACKGROUND
Elevated plasma homocysteine (Hcy) is a risk factor for coronary disease. The objective of this study was to investigate whether Hcy either alone or in high glucose conditions induces endothelin-1 (ET-1) synthesis via the production of reactive oxygen species (ROS).


METHODS
Bovine aortic endothelial cells were grown in high (25 mmol/l) and low (5 mmol/l) glucose medium.


RESULTS
In high glucose, Hcy caused a time-dependent increase in ET-1 release, which was greatest with 50 micromol/l Hcy at 24 h (p < 0.01). This effect was not seen in low glucose conditions. In high glucose and 50 micromol/l Hcy, ET-1 mRNA levels were maximal after 1 h (p < 0.05). Tissue factor mRNA levels were raised at 4 h (p < 0.05) and functional activity was raised at 6 h (p < 0.01). Intracellular ROS production was increased by 50 micromol/l Hcy after 24 h (p < 0.05) but only in high glucose. To investigate the role of mitochondrial metabolism in ROS production, cells were incubated with thenoyltrifluoroacetone (inhibitor of complex II) or carbonyl cyanide m-chlorophenylhydrazone (uncoupler of oxidative phosphorylation). Both compounds abolished the Hcy-induced increase in ROS production and ET-1 release. There was an alteration in intracellular glutathione (GSH) levels with Hcy treatment with more oxidised GSH present.


CONCLUSIONS
The combined metabolic burden of Hcy and high glucose stimulates ET-1 synthesis in bovine aortic endothelial cells via a mechanism dependent on the production of mitochondrial ROS, but may not be generalisable to all types of endothelial cells.
Sethi
Amarjit S
AS
Department of Experimental Therapeutics, William Harvey Research Institute, London, UK. Amarjit.Sethi@eht.nhs.uk
Lees
Delphine M
DM
Douthwaite
Julie A
JA
Dawnay
Anne B
AB
Corder
Roger
R
D000818:Animals; D001011:Aorta / Q000166:cytology; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002417:Cattle; D002478:Cells, Cultured; D042963:Electron Transport Complex II / Q000037:antagonists & inhibitors; D019332:Endothelin-1 / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D005947:Glucose / Q000008:administration & dosage; D005978:Glutathione / Q000378:metabolism; D006710:Homocysteine / Q000494:pharmacology*; D006943:Hyperglycemia / Q000503:physiopathology*; D018384:Oxidative Stress / Q000502:physiology; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013804:Thenoyltrifluoroacetone / Q000494:pharmacology; D013925:Thromboplastin / Q000378:metabolism; D014475:Uncoupling Agents / Q000494:pharmacology
D019332:Endothelin-1; D017382:Reactive Oxygen Species; D014475:Uncoupling Agents; D006710:Homocysteine; D013804:Thenoyltrifluoroacetone; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D013925:Thromboplastin; D042963:Electron Transport Complex II; D005978:Glutathione; D005947:Glucose