Resveratrol, a polyphenolic compound found in grapes and wine, is an agonist for the estrogen receptor

Abstract

The phytochemical resveratrol, which is found in grapes and wine, has been reported to have a variety of anti-inflammatory, anti-platelet, and anti-carcinogenic effects. Based on its structural similarity to diethylstilbestrol, a synthetic estrogen, we examined whether resveratrol might be a phytoestrogen. At concentrations (approximately 3-10 microM) comparable to those required for its other biological effects, resveratrol inhibited the binding of labeled estradiol to the estrogen receptor and it activated transcription of estrogen-responsive reporter genes transfected into human breast cancer cells. This transcriptional activation was estrogen receptor-dependent, required an estrogen response element in the reporter gene, and was inhibited by specific estrogen antagonists. In some cell types (e.g., MCF-7 cells), resveratrol functioned as a superagonist (i.e., produced a greater maximal transcriptional response than estradiol) whereas in others it produced activation equal to or less than that of estradiol. Resveratrol also increased the expression of native estrogen-regulated genes, and it stimulated the proliferation of estrogen-dependent T47D breast cancer cells. We conclude that resveratrol is a phytoestrogen and that it exhibits variable degrees of estrogen receptor agonism in different test systems. The estrogenic actions of resveratrol broaden the spectrum of its biological actions and may be relevant to the reported cardiovascular benefits of drinking wine.

Figure 1

Resveratrol competes for estrogen receptor binding. (A) Structures of resveratrol, DES, and estradiol. (B) Estrogen receptor binding assays were performed as described in Experimental Procedures by using 0.1 nM (circles), 0.3 nM (triangles), or 1.0 nM (squares). ¹²⁵I-estradiol competed with the indicated concentrations of resveratrol (open symbols) or unlabeled estradiol (filled circles). Each point represents the mean and range of duplicate assays after subtraction of nonspecific binding. All results are shown as percentage of binding in the absence of competitor.

Figure 2

Activation of estrogen-responsive reporter plasmids by resveratrol in MCF-7 cells. (A) MCF-7 cells were transfected with ERE-tk109-luc, incubated with the indicated amounts of estradiol or resveratrol, and assayed for luciferase activity as described above. (B) Comparisons of estradiol, resveratrol, and DES using different reporter genes. (C) Effect of combined estradiol and resveratrol in the activation of ERE-tk109-luc. Results are shown as mean ± SEM for triplicate transfections.

Figure 3

Transcriptional activation by resveratrol is estrogen receptor-mediated. (A) Dependence on an ERE. MCF-7 cells were transfected with tk109-luc or ERE-tk109-luc, incubated with the indicated concentrations of resveratrol or estradiol, and assayed for luciferase activity. Results are shown as fold induction compared with control-treated cells. (B) Inhibition by estrogen antagonists. MCF-7 cells were transfected with ERE-tk109-luc, incubated with no agonist, estradiol (0.1 nM), or resveratrol (10 μM) or with the agonists plus the antagonists tamoxifen (1 μM) or ICI 182780 (100 nM). (C) Requirement for the estrogen receptor. MDA-MB-231 cells were transfected with 1 μg/well ERE2-tk109-luc and 10 ng/well of pSG5 or pSG5-HEGO, incubated with the indicated concentrations of resveratrol or estradiol, and assayed for luciferase activity. All values are means ± SEM of triplicate transfections.

Figure 4

Agonistic activity of resveratrol is cell type-dependent. MCF-7 and BG-1 cells were transfected with ERE2-tk109-luc, incubated with maximally activating concentrations of estradiol and resveratrol (determined separately for each cell type), and assayed for luciferase activity. Estradiol and resveratrol concentrations were 10 pM and 10 μM, respectively, for MCF-7 cells and 1 nM and 33 μM for BG-1 cells. Results are mean ± SEM for three (MCF-7) or four (BG-1) replicate transfections.

Figure 5

Effect of resveratrol on estrogen-regulated transcription and growth. (A) MCF-7 cells were treated with resveratrol or estradiol, and progesterone receptor (PR) mRNA was measured by RT-PCR. Results are normalized to glyceraldehyde-3-phosphate dehydrogenase mRNA and shown as fold induction relative to control (0.1% ethanol). Similar results were obtained in an independent experiment. (B) T47D cells were cultured in the presence of 0.1 nM estradiol or 10 μM resveratrol with or without 100 nM ICI 182780 or in control medium. Cell density was determined at the indicated intervals. Each point represents the mean ± SEM of eight replicate wells (n = 16 for day 0).