Homocysteine induces VCAM-1 gene expression through NF-kappaB and NAD(P)H oxidase activation: protective role of Mediterranean diet polyphenolic antioxidants.
Hyperhomocysteinemia is a recognized risk factor for vascular disease, but pathogenetic mechanisms involved in its vascular actions are largely unknown. Because VCAM-1 expression is crucial in monocyte adhesion and early atherogenesis, we evaluated the NF-kappaB-related induction of VCAM-1 by homocysteine (Hcy) and the possible inhibitory effect of dietary polyphenolic antioxidants, such as trans-resveratrol (RSV) and hydroxytyrosol (HT), which are known inhibitors of NF-kappaB-mediated VCAM-1 induction. In human umbilical vein endothelial cells (HUVEC), Hcy, at 100 micromol/l, but not cysteine, induced VCAM-1 expression at the protein and mRNA levels, as shown by enzyme immunoassay and Northern analysis, respectively. Transfection studies with deletional VCAM-1 promoter constructs demonstrated that the two tandem NF-kappaB motifs in the VCAM-1 promoter are necessary for Hcy-induced VCAM-1 gene expression. Hcy-induced NF-kappaB activation was confirmed by EMSA, as shown by the nuclear translocation of its p65 (RelA) subunit and the degradation of the inhibitors IkappaB-alpha and IkappaB-beta by Western analysis. Hcy also increased intracellular reactive oxygen species by NAD(P)H oxidase activation, as shown by the membrane translocation of its p47(phox) subunit. NF-kappaB inhibitors decreased Hcy-induced intracellular reactive oxygen species and VCAM-1 expression. Finally, we found that nutritionally relevant concentrations of RSV and HT, but not folate and vitamin B6, reduce (by >60% at 10(-6) mol/l) Hcy-induced VCAM-1 expression and monocytoid cell adhesion to the endothelium. These data indicate that pathophysiologically relevant Hcy concentrations induce VCAM-1 expression through a prooxidant mechanism involving NF-kappaB. Natural Mediterranean diet antioxidants can inhibit such activation, suggesting their possible therapeutic role in Hcy-induced vascular damage.
Carluccio
Maria Annunziata
MA
Consiglio Nazionale delle Ricerca Institute of Clinical Physiology, University of Lecce, Lecce, Italy.
Ancora
Maria Assunta
MA
Massaro
Marika
M
Carluccio
Marisa
M
Scoditti
Egeria
E
Distante
Alessandro
A
Storelli
Carlo
C
De Caterina
Raffaele
R
D000975:Antioxidants / Q000494:pharmacology* / Q000627:therapeutic use; D002448:Cell Adhesion / Q000187:drug effects; D002478:Cells, Cultured; D038441:Diet, Mediterranean; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D005419:Flavonoids / Q000494:pharmacology* / Q000627:therapeutic use; D006710:Homocysteine / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D020138:Hyperhomocysteinemia / Q000201:enzymology / Q000378:metabolism / Q000517:prevention & control; D020672:I-kappa B Proteins / Q000378:metabolism; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism; D009154:Mutation; D019255:NADPH Oxidases / Q000378:metabolism*; D010636:Phenols / Q000494:pharmacology* / Q000627:therapeutic use; D010626:Phenylethyl Alcohol / Q000031:analogs & derivatives / Q000494:pharmacology; D010766:Phosphorylation; D059808:Polyphenols; D011401:Promoter Regions, Genetic / Q000187:drug effects; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D021381:Protein Transport; D012333:RNA, Messenger / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D000077185:Resveratrol; D013267:Stilbenes / Q000494:pharmacology; D051996:Transcription Factor RelA / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D015854:Up-Regulation; D019010:Vascular Cell Adhesion Molecule-1 / Q000235:genetics / Q000378:metabolism*
D000975:Antioxidants; D005419:Flavonoids; D020672:I-kappa B Proteins; D010636:Phenols; D059808:Polyphenols; C498143:RELA protein, human; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D013267:Stilbenes; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha; D019010:Vascular Cell Adhesion Molecule-1; D006710:Homocysteine; C005975:3,4-dihydroxyphenylethanol; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1; D046988:Proteasome Endopeptidase Complex; D010626:Phenylethyl Alcohol; D000077185:Resveratrol