Beneficial effects of red wine polyphenols on cardiovascular health are well known. The aim of our research was an interaction
study of four red wine polyphenols – resveratrol (R), quercetin (Q), kaempferol (KF) and isorhamnetin (IR) of their ability to activate
endothelial NO synthase (eNOS) promoter when used alone and in equimolar mixtures. To determine their activity, we performed
a luciferase reporter gene assay on EA.hy926 cells stably transfected with a luciferase reporter gene construct containing
eNOS promoter. The Bradford assay was also performed to account the cytotoxicity and/or the cell number differences. The
median effect equation, as an interaction analysis evaluating synergy or antagonism of the combinations was done according to
mass-action law principle. Isobolographic method was performed on selected double mixtures and dose reduction index was
calculated for all mixtures. All single polyphenols activated eNOS promoter. The EC50 values were in micromolar concentrations
ranging from 3.44 μM (R2 = 0.96) for kaempferol to 9.89 μM for isorhamnetin (R2 = 0.94). All mixtures activated eNOS promoter,
but their interactions varied from synergy (Q+R, Q+IR+KF, Q+R+KF and Q+R+IR+KF), through additive (R+IR+KF) to antagonistic
interaction (R+IR, R+KF, Q+IR, Q+KF, IR+KF and R+Q+IR). In this study, we show for the first time that red wine polyphenols activated
eNOS promoter when used alone and in mixtures with different type of interactions.
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