Homocysteine-induced inhibition of endothelium-dependent relaxation in rabbit aorta - Role for superoxide anions

Hyperhomocysteinemia is associated with endothelial dysfunction, although i ts mechanism is unknown. Isometric tension recordings and lucigenin chemilu minescence were used to assess the effects of homocysteine exposure on endo thelium-dependent and -independent relaxation in isolated rabbit aortic rin gs and superoxide anion (O-2(-)) production by cultured porcine aortic endo thelial cells, respectively. Homocysteine (0.1 to 10 mmol/L) produced a sig nificant (P<0.001) concentration- and time-dependent inhibition of endothel ium-dependent relaxation in response to both acetylcholine and the calcium ionophore A23187, Only the: intracellular O-2(-) scavenger 4,5-dihydroxy-1, 3-benzene disulfonic acid (Tiron, 10 mmol/L) significantly (P<0.001) inhibi ted the effect of homocysteine on acetylcholine- and A23187-induced relaxat ion, Incubation of porcine aortic endothelial cells with homocysteine (0.03 to 1 mmol/L for up to 72 hours) caused a significant (P<0.001) time-depend ent increase in the O-2(-) released by these cells on the addition of Trito n X-100 (1% [vol/vol]), with levels returning to values comparable to those of control cells at the 72-hour time point. These changes in O-2(-) levels were associated with a time-dependent increase in endothelial cell superox ide dismutase activity, becoming significant (P<0.001) after 72 hours. Furt hermore, the homocysteine-induced increase in endothelial cell O-2(-) level s was completely inhibited (P<0.001) by the concomitant incubation with eit her Tiron (10 mmol/L), vitamin C (10 mu mol/L), or vitamin E (10 mu mol/L). These data suggest that the inhibitory effect of homocysteine on endotheli um-dependent relaxation is due to an increase in the endothelial cell intra cellular levels of O-2(-) and provide a possible mechanism for the endothel ial dysfunction associated with hyperhomocysteinemia.