Homocysteine-induced decrease in endothelin-1 production is initiated at the extracellular level and involves oxidative products

The increased cardiovascular risk associated with hyperhomocysteinemia has been partly related to homocysteine (Hcy)-induced endothelial cell dysfunct ion. However, the intra or extracellular starting point of the interaction between Hcy and endothelial cells, leading to cellular dysfunction, has not yet been identified. We investigated the effects of both intracellular and extracellular Hcy accumulation on endothelin-1 (ET-1) synthesis by culture d human endothelial cells. Incubation of cultures with methionine (1.0 mmol .L-1) for 2 It induced a slight increase in cellular Hcy content but no cha nge in ET-1 production. Incubation of cells with Hcy (0.2 mmol.L-1) led to a significant fall in ET-1 generation, accompanied by a significant increas e in cellular Hcy content. Addition of the amino-acid transport system L su bstrate 2-amino-2-norbornane carboxylic acid had no effect on the Hcy-induc ed decrease in ET-1 production but significantly inhibited the Hcy-induced increase in the cellular Hcy content. Incubation of cells with a lower Hcy concentration (0.05 mmol.L-1) also reduced ET-1 production without increasi ng the cellular Hcy content. Co-incubation with extracellular free-radical inhibitors (superoxide dismutase, catalase and mannitol) markedly reduced t he effect of Hcy on ET-1 production. Thus, it is extracellular Hcy accumula tion that triggers the decrease in ET-1 production by endothelial cells thr ough oxidative products.