Prominent role of NF-kappaB in the induction of endothelial activation by endogenous nitric oxide inhibition.
Decreased endothelial nitric oxide (NO) production and increased expression of vascular cell adhesion molecule-1 (VCAM-1) are early features of atherosclerosis. We investigated the effects of suppressing endogenous NO production by the NO synthase inhibitor l-mono-methyl-arginine (L-NMMA), given alone or in combination with interleukin(IL)-1alpha, on VCAM-1 expression by human umbilical vein endothelial cells (HUVEC). VCAM-1 expression (by enzyme immunoassay), barely detectable at baseline, was significantly increased by L-NMMA (by no more than 20% over control compared with IL-1alpha induction). This was paralleled by an increase in U937 monocytoid cell adhesion. When HUVEC incubated with L-NMMA were stimulated with low concentrations of IL-1alpha (0.05-0.5ng/mL), these determined a higher VCAM-1 expression than in the presence of L-NMMA or IL-1alpha alone. Northern analysis indicated that VCAM-1 mRNA was induced by L-NMMA alone, and that the effects of L-NMMA and IL-1alpha were, again, at least additive. Nuclear factor-kappaB (NF-kappaB), GATA, activator protein-1 (AP-1) and interferon regulatory factor-1 (IRF-1), transcription factors all involved in VCAM-1 gene expression, were all activated at electrophoretic mobility shift assay and at chromatin immunoprecipitation assay by L-NMMA, but additive effects with the combined administration of L-NMMA and IL-1alpha only occurred for NF-kappaB. These results support the view that endogenous NO mantains a normal endothelial non-reactivity towards circulating monocytes, and that suppression of this endogenous brake for endothelial activation results in the activation of multiple transcription factors even in the absence of other endothelial activators, with a prominent role of NF-kappaB in the presence or absence of other inflammatory mediators.
Lazzerini
Guido
G
C.N.R. Institute of Clinical Physiology, Pisa, Italy.
Del Turco
Serena
S
Basta
Giuseppina
G
O'Loghlen
Ana
A
Zampolli
Antonella
A
Caterina
Raffaele De
RD
D050197:Atherosclerosis / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D024202:Electrophoretic Mobility Shift Assay; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D050980:GATA Transcription Factors / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D050856:Interferon Regulatory Factor-1 / Q000187:drug effects / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism*; D009569:Nitric Oxide / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors; D011401:Promoter Regions, Genetic; D018808:Transcription Factor AP-1 / Q000187:drug effects / Q000378:metabolism; D019010:Vascular Cell Adhesion Molecule-1 / Q000096:biosynthesis* / Q000235:genetics; D019323:omega-N-Methylarginine / Q000494:pharmacology
D004791:Enzyme Inhibitors; D050980:GATA Transcription Factors; D050856:Interferon Regulatory Factor-1; D016328:NF-kappa B; D018808:Transcription Factor AP-1; D019010:Vascular Cell Adhesion Molecule-1; D019323:omega-N-Methylarginine; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase