Homocysteine stimulates inducible nitric oxide synthase expression in macrophages: antagonizing effect of ginkgolides and bilobalide.

Hyperhomocysteinemia is an independent risk factor for atherosclerotic diseases. Inducible nitric oxide synthase (iNOS) is mainly expressed in macrophages upon stimulation. Overproduction of nitric oxide (NO) by iNOS can exacerbate the development of atherosclerosis. Our previous studies demonstrated that the extract of ginkgo biloba leaves (EGb) inhibited the iNOS-mediated NO production in monocyte-derived macrophage. We also reported that homocysteine could stimulate monocyte chemoattractant protein-1 (MCP-1) expression in vascular cells causing enhanced monocyte chemotaxis. The objective of the present study was to investigate the effect of homocysteine on iNOS-mediated NO production in macrophages and the antagonizing effect of EGb. Human monocytic cell (THP-1)-derived macrophages were incubated with homocysteine for various time periods. Homocysteine at concentrations of 0.05-0.1 mM significantly stimulated NO production and iNOS activity in macrophages via increased expression of iNOS mRNA and protein. The increased iNOS expression was associated with activation of nuclear factor-kappa B (NF-kappaB) arising from reduced expression of inhibitor protein (IkappaB alpha) mRNA as well as increased phosphorylation of IkappaB alpha protein in homocysteine-treated cells. EGb and its terpenoids (ginkgolide A, ginkgolide B and bilobalide) could antagonize the homocysteine effect on iNOS expression in macrophages via their antioxidant effect resulting in attenuation of NF-kappaB activation. Taken together, our results have demonstrated that homocysteine, at pathophysiological concentrations, stimulates iNOS-mediated NO production in macrophages. EGb and its terpenoids can antagonize such stimulatory effect via antioxidation and attenuation of NF-kappaB activation.