Resveratrol, a natural phenolic compound, inhibits cell proliferation and prevents oxidative DNA damage

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Abstract

Resveratrol (3,4′,5-trihydroxystilbene) is a naturally occurring phenolic compound which is present at high levels in wine and has been recently proposed as a potential cancer chemopreventive and chemoterapeutic agent. In this study, we evaluated the antiproliferative activity of resveratrol on a panel of cell lines of various histogenetic origin, including normal rat fibroblasts and mouse mammary epithelial cells compared to human breast, colon and prostate cancer cells. The concentration of resveratrol inhibiting cell growth by 50% (IC50) ranged from about 20 to 100 μM. At such concentration, we were unable to detect a significant increase in the apoptotic index in most of the cell lines analyzed.

We also studied the effects of resveratrol on cell cycle distribution. The most striking effect was a reduction in the percentage of cells in the G2/M phase which was most frequently associated with an increase of cells in the S phase of the cell cycle. We also found that resveratrol is able to prevent the increase in reactive oxygen species (ROS) following exposure to oxidative agents (i.e. tobacco-smoke condensate (TAR) and H2O2). Resveratrol also reduced nuclear DNA fragmentation, as assessed by single cell gel electrophoresis (comet test). Taken together our results suggest that resveratrol can act as an antimutagenic/anticarcinogenic agent by preventing oxidative DNA damage which plays a pivotal role in the carcinogenic activity of many genotoxic agents.

Introduction

Resveratrol (3,4′,5-trihydroxystilbene) is a naturally occurring phenolic compound found in the berry skins of most grape cultivars and in a variety of medicinal plants where it functions as a phytoalexin protecting against fungal infections [1]. Recent evidence suggests that this molecule can affect many biological activities. Indeed, it has been proposed that resveratrol, which is present at high levels in wine, may explain for the reduced risk of coronary heart disease associated with moderate wine consumption [2]. This effect has been attributed to the inhibition of oxidation of low-density lipoprotein cholesterol [3], platelet aggregation and coagulation [4], [5]. Resveratrol has also been reported to compete with 17β-estradiol for estrogen receptor and activate estrogen-responsive reporter genes in vitro [6] but these results have not been confirmed by in vivo studies [7].

It has been reported that diet supplementation with dealcoholized red wine solids can inhibit tumor formation in mice [8]. Several reports suggest that resveratrol might mediate this protective effect of moderate wine consumption. Indeed, it has been shown to inhibit cellular events associated with tumor initiation, promotion and progression [9]. In fact, resveratrol inhibits free-radical formation and has an antimutagenic activity [9], [10]. It also has an anti-inflammatory effect mainly due to its ability to inhibit cyclooxigenase and hydroperoxidase functions [9], [11] and it has been shown to induce differentiation of human promyelocitic leukemia [9] and osteoblastic cells [12]. Resveratrol also inhibits, in a dose-dependent manner, the development of preneoplastic lesions in a mouse mammary gland culture model of chemical carcinogenesis [9] and reduces tumor formation in a two-stage mouse skin cancer model [9]. Moreover, it inhibits tumor cell growth in a rat model of ascites hepatoma [13] and has been shown to induce cell cycle arrest [14] and apoptosis in human promyelocitic leukemia cells [15]. These data have suggested that resveratrol might be effective as both cancer chemopreventive and chemoterapeutic agent and support the need for further studies on the effects of this molecule in both normal and tumor cells.

In the present study, we evaluated the effects of resveratrol on the growth and cell cycle parameters of a series of human cancer cell lines. We found that resveratrol inhibits the in vitro growth of different types of human cancer cells and that it has specific effects on cell cycle parameters. We also found that resveratrol prevents oxidative DNA damage induced by different stimuli. The implications of these findings are discussed.

Section snippets

Cell culture

The HC11 mouse mammary epithelial cells were clonally derived from a spontaneously immortalized mammary epithelial cell culture originally established from a mid-term pregnant BALB/c mouse [16] and were grown and maintained in RPMI-1640 medium (RPMI) (Gibco, Merelbeke, Belgium) supplemented with 10% heat inactivated fetal bovine serum (FBS). The Rat-1 normal rat fibroblasts were grown and maintained in Eagle’s minimum essential medium (EMEM) (Gibco) supplemented with 10% heat inactivated FBS.

Effects of resveratrol on cell proliferation and cell cycle parameters on cell lines of various histogenetic origin

In a first feasibility study, we aimed to determine the concentration of resveratrol inhibiting cell growth by 50% (IC50) in a series of cell lines including diploid rodent fibroblasts (Rat-1), normal mouse mammary epithelial cells (HC11), three human breast (MCF-7, T47-D and BT-549), three colon (HCT116, HT-29 and SW-620), two prostate (PC3 and DU145) and one cervix (HeLa) cancer cell lines. Cytotoxicity assays were carried out by use of the MTT test. Exponentially growing cultures of each

Discussion

Epidemiological studies have suggested that dietary factors play an important role in cancer development in humans and the preventive effects of plant-based diets is well documented [21], [22]. Resveratrol is a stilbene (3,4′,5-trihydroxystilbene) with a relatively broad distribution in plants and is present in various human foods, including red wines, peanuts and mulberries [23]. It is an antioxidant and has been shown to inhibit various stages of tumor development [9].

In this study, we

Acknowledgements

This work was supported in part by a grant from “Co-finanziamento MURST” 1998. We thank O. Cantoni and P. Sestili (Università di Urbino, Italy) for initial assistance with the comet test.

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