Abstract 1397: Homocysteine Induces Endothelial Activation Through Proteasome Dependent Pathway: Protective Role Of Natural Polyphenolic antioxidants
Hyperhomocysteinemia is a recognized risk factor for cardiovascular disease, although its causal role is under discussion. Since endothelial-leukocyte adhesion molecules expression is a crucial event in atherogenesis, we evaluated the effects of homocysteine on endothelial activation and investigated mechanisms involved.
Methods and Results - In HUVEC cells homocysteine (Hcy), at concentrations ≥50 μmol/L, but not cysteine, induced VCAM-1 expression at the protein and mRNA level, (2.4 ± 0.3 folds vs control, p<0.05; and 1.9 ± 0.2 folds vs control, p<0.05 at enzyme immunoassay and Northern analysis, respectively). Homocysteine-induced VCAM-1 expression was not mediated by the endothelial autocrine production of TNFα; however, hcy potentiated TNFα in increasing VCAM-1, as well as ICAM-1 and E-selectin expression. Transfection studies, with deletional VCAM-1 promoter constructs, demonstrated that homocysteine induced VCAM-1 gene expression mostly by NF-κB motifs in the VCAM-1 promoter region. Lactacystin, a specific proteasome inhibitor, abolished homocysteine-stimulated VCAM-1 expression, suggesting the involvement of the nuclear factor (NF)-kB proteasome-dependent pathway. Hcy indeed induced NF-kB activation, the nuclear translocation of its p65 subunit and the phosphorylation and subsequent degradation of inhibitor(I)kB. Hcy, in part by activating NAD(P)H oxidase, increased intracellular generation of reactive oxygen species (ROS), as assessed by dichlorofluoresceine fluorescence assays. In assessing the effects of various dietetic phenolic antioxidants, we found that nutritionally relevant concentrations of trans-resveratrol (RSV) and hydroxytyrosol (HT), but not folate and B vitamins, reduced both the production of ROS and homocysteine-induced VCAM-1 expression as well as monocyte adhesion.
Conclusion. Pathophysiologically relevant Hcyconcentrations induce endothelial activation. This can be balanced by natural Mediterranean diet antioxidants, thus suggesting a possible therapeutic role of these natural antioxidants in hyperhomocysteine-induced vascular damage.