Resveratrol Inhibits the Growth of Gastric Cancer by Inducing G1 Phase Arrest and Senescence in a Sirt1-Dependent Manner

Resveratrol, a naturally occurring polyphenolic compound, has been reported to exert anticancer activity by affecting diverse molecular targets. In this study, we examined the effects and the underlying mechanisms of resveratrol on gastric cancer. We found that resveratrol inhibited the proliferation of gastric cancer cells in a dose-dependent manner. At the concentration of 25 and 50 µM, resveratrol inhibited the cell viability and diminished the clonogenic potential of gastric cancer cells. Resveratrol treatment arrested gastric cancer cells in the G1 phase and led to senescence instead of apoptosis. Regulators of the cell cycle and senescence pathways, including cyclin D1, cyclin-dependent kinase (CDK4 and 6), p21 and p16, were dysregulated by resveratrol treatment. The inhibitory effects of resveratrol on gastric cancer were also verified in vivo using a nude mice xenograft model. Resveratrol (40 mg/kg/d) exerted inhibitory activities on gastric cancer development and significantly decreased the fractions of Ki67-positive cells in the tumor specimens from the nude mice. After resveratrol treatment, the induction of senescence and the changes in the expression of the regulators involved in the cell cycle and senescence pathways were similar to what we observed in vitro. However, the depletion of Sirtuin (Sirt)1 reversed the above-described effects of resveratrol both in vitro and in vivo. Our data suggest that resveratrol inhibits gastric cancer in a Sirt1-dependent manner and provide detailed evidence for the possibility of applying resveratrol in gastric cancer prevention and therapy.     

Effect of resveratrol on intimal hyperplasia after endothelial denudation in an experimental rabbit model

The ability of resveratrol to inhibit vascular intimal thickening was tested in an experimental model in which endothelial denudation was performed in the normal rabbit iliac artery. Resveratrol (2 approximately 4mg/ kg/d) was administered intragastrically for 5 weeks beginning 1 week before denudation. At the higher concentration of resveratrol, the intimal hyperplasia of injured vascular wall was effectively inhibited; the intimal proliferation index also was significantly less than that in the untreated control group (0.28 +/- 0.07 vs 0.41 +/- 0.13, respectively, p<0.01); the relative luminal area increased from 0.38 +/- 0.06 in the untreated control group to 0.53 +/- 0.10 in the resveratrol treatment group (p < 0.001); and the count of smooth muscle cells in the thickened intima was statistically significantly reduced in the high dose resveratrol treatment group than that in the untreated group (1,115 +/- 510 vs 1,796 +/- 963, respectively, p < 0.05). Resveratrol added to the culture media of cultured rabbit vascular smooth muscle cells inhibited DNA synthesis in a dose-dependent manner. These results showing that resveratrol is capable of inhibiting intimal hyperplasia of injured artery raise the possibility that this polyphenol might have clinical potential in prevention and treatment of restenosis after angioplasty.     

Treatment of mammalian cells with mimosine generates DNA breaks

Exponentially growing mouse erythroleukemia (MEL) cells and quiescent human peripheral blood lymphocytes (PBL) were treated with different concentrations of the nonprotein amino acid mimosine for 16 h. The treatment of the cycling cell population with 400 microM mimosine caused inhibition of DNA replication, changes in the progression of the cells in the cell cycle, and apoptosis. Nucleoid sedimentation analysis and comet assay were used to monitor the appearance and accumulation of DNA breaks. The rate of break accumulation was dose-dependent, did not depend on the stage of the cell cycle and was not connected with the mechanism of DNA replication. The data indicate that the effects of mimosine on DNA synthesis and the cell cycle may be a result of introduction of breaks into DNA.     

Resveratrol attenuates angiotensin II-induced senescence of vascular smooth muscle cells

Resveratrol (3,5,4'-trihydroxystilbene), a polyphenol abundant in red wine, is known to extend the life span of diverse species. On the contrary, it was reported that angiotensin (Ang) II enhances senescence of vascular smooth muscle cells (VSMCs). We, therefore, examined whether resveratrol attenuates Ang II-induced senescence of VSMC. Senescence-associated β-galactosidase (SA β-gal) assay showed that Ang II induced senescence of VSMC. The Ang II-induced senescence was inhibited by losartan, an Ang II type 1 receptor (AT1R) antagonist but not by PD123319, Ang II type 2 receptor antagonist, indicating that AT1R is responsible for the induction of senescence. Resveratrol suppressed Ang II-induced senescence of VSMC in a dose-dependent manner. In addition, resveratrol suppressed Ang II-induced induction of p53 and its downstream target gene p21, both of which play an important role in the induction of senescence. Resveratrol suppressed senescence of VSMC possibly through inhibition of AT1R-dependent induction of p53/p21. Suppression of p53 induction may be involved in the longevity by resveratrol.     

Resveratrol modulates gene expression associated with apoptosis, proliferation and cell cycle in cells with mutated human c-Ha-Ras, but does not alter c-Ha-Ras mRNA or protein expression

An accumulating body of evidence suggests that resveratrol can inhibit carcinogenesis through antiproliferative and apoptotic effects. One proposed mechanism for this is the modulation of genes, for example, Ras and p53, frequently associated with human cancer. To test the effect of resveratrol on gene expression, we used the WR-21 cell line because it contains a mutated human c-Ha-ras gene. Cells at > or =70% confluency were incubated with media alone or with increasing concentrations of trans-resveratrol (0.1-1000 microM) for 24 h. Resveratrol (30-100 microM) decreased cellular proliferation by 80% (bromodeoxyuridine incorporation) and increased apoptosis by 60% (TUNEL). Cells were then treated with media alone or with 50-microM resveratrol for 24 h. RNA was isolated for nylon-based macroarray analyses and protein for immunoblotting. Resveratrol increased (+) and decreased (-) gene expression associated with apoptosis (Birc5+, Cash+, Mcl-1+, Mdm2+, Rpa-like+), cellular proliferation (Ctsd+, Mdm2+, Egr1+, ODC+) and cell cycle (cyclin D+, cyclin g+, Gadd45a-, Mad2l-, Mdm2+). Resveratrol consistently increased by > or =6-fold Mdm2 expression and other downstream p53 effectors, but not p53 itself at 24 h. Subsequent cell cycle analysis indicated a significant accumulation of cells in G2/M, and a decrease in G1/G0 suggesting a G2/M blockade. Further RT-PCR and Western blot analyses indicated no differential changes in Ras mRNA expression or p21(ras) protein levels, respectively. These results suggest that resveratrol potently inhibits cellular proliferation, increases apoptosis, alters cell cycle dynamics and modulates associated gene expression. Furthermore, these effects appear mediated, in part, by p53 without direct modulation of mutant c-Ha-ras expression.     

Resveratrol is a selective human cytochrome P450 1A1 inhibitor.

Resveratrol (trans-3,4',5-trihydroxystilbene) is a phytoalexin compound found in juice and wine produced from dark-skinned grape cultivars and reported to have anti-inflammatory and anticarcinogenic activities. To investigate the mechanism of anticarcinogenic activities of resveratrol, the effects on cytochrome P450 (P450) were determined in human liver microsomes and Escherichia coli membranes coexpressing human P450 1A1 or P450 1A2 with human NADPH-P450 reductase (bicistronic expression system). Resveratrol slightly inhibited ethoxyresorufin O-deethylation (EROD) activity in human liver microsomes with an IC(50) of 1.1 mM. Interestingly, resveratrol exhibited potent inhibition of human P450 1A1 in a dose-dependent manner with IC(50) of 23 microM for EROD and IC(50) of 11 microM for methoxyresorufin O-demethylation (MROD). However, the inhibition of human P450 1A2 by resveratrol was not so strong (IC(50) 1.2 mM for EROD and 580 microM for MROD). Resveratrol showed over 50-fold selectivity for P450 1A1 over P450 1A2. The activities of human NADPH-P450 reductase were not significantly changed by resveratrol. In a human P450 1A1/reductase bicistronic expression system, resveratrol inhibited human P450 1A1 activity in a mixed-type inhibition (competitive-noncompetitive) with a K(i) values of 9 and 89 microM. These results suggest that resveratrol is a selective human P450 1A1 inhibitor, and may be considered for use as a strong cancer chemopreventive agent in humans.     

Resveratrol inhibits AGEs-induced proliferation and collagen synthesis activity in vascular smooth muscle cells from stroke-prone spontaneously hypertensive rats

Advanced glycation end-products (AGEs) of plasma proteins and/or matrix proteins are candidate mediators for various vascular complications such as atherosclerosis. We previously reported a significantly larger accumulation of AGEs of the aorta in stroke-prone spontaneously hypertensive rats (SHRSP) than in age-matched Wistar-Kyoto rats (WKY). In this study, we examined the effects of AGEs on vascular smooth muscle cells (VSMC) from SHRSP and WKY rats. We also studied the in vitro effects of resveratrol (3, 4',5-trihydroxystilbene), a natural phytestrogen, on VSMC proliferation, DNA synthesis, and collagen synthesis activity in SHRSP-VSMC. AGEs accelerated the proliferation of SHRSP- or WKY-VSMC in a time- and dose-dependent manner. VSMC from SHRSP were more sensitive to AGEs than VSMC from normotensive WKY. AGEs also significantly increased DNA synthesis and prolyl hydroxylase activity, a marker for collagen synthesis, in SHRSP-VSMC. AGEs-induced increases in TGF-beta1 mRNA in SHRSP-VSMC were significantly greater than in WKY-VSMC. Resveratrol inhibited AGEs-stimulated proliferation, DNA synthesis, and prolyl hydroxylase activity in SHRSP-VSMC in a dose-dependent manner. ICI 182780, a specific estrogen receptor antagonist, partly blocked the inhibitory effects of resveratrol on AGEs-stimulated proliferation, DNA synthesis, and prolyl hydroxylase activity. Resveratrol significantly inhibited AGEs-induced TGF-beta1 mRNA increases in a dose-dependent manner. Thus, resveratrol may confer protective effects on the cardiovascular system by attenuating vascular remodeling and may be clinically useful as a safer substitute for feminizing estrogens in preventing cardiovascular disease.     

Lovastatin inhibits G1/S transition of normal human B-lymphocytes independent of apoptosis.

Lovastatin is a potent inhibitor of protein prenylation, and it has been reported to have pleiotropic cellular effects. In the present study we have elucidated the effects of lovastatin on cell cycle progression and apoptosis of normal human B-lymphocytes. When added to B-lymphocytes stimulated with anti-immunoglobulin (anti-mu) and SAC, lovastatin (20 microM) inhibited the cells in the late G1 phase of the cell cycle. Thus, no early activation parameters such as Ca(2+) flux or MYC induction were affected by lovastatin, whereas progression of cells into the second cell cycle as well as DNA synthesis was markedly reduced. We therefore examined the effects of lovastatin on components of the cell cycle machinery responsible for regulating the G1/S transition. We demonstrated that pRB phosphorylation, cdk2 activity needed for this phosphorylation, and the levels of cyclin A, D, and E were inhibited after 24 h of lovastatin treatment, while the levels of p27(Kip1) were elevated. There was no effect on p21(Cip1), cyclin D2, cdk4, and cdk6. These data are consistent with the cells being inhibited by lovastatin between 24 and 32 h into G1. Lovastatin added to stimulated B-cells in late G1 still inhibited the DNA synthesis by 60%, but at this point only minor effects were noted on the cell cycle machinery. We therefore looked for induced apoptosis as an explanation for reduced S-phase entry of the cells. However, despite the ability to enhance the apoptosis of unstimulated B-cells from 48 to 61% as judged by the TUNEL method, lovastatin only marginally affected apoptosis when administered to stimulated B-cells. Thus, it appears that accelerated apoptosis cannot account for the effect of lovastatin on cell cycle progression.     

Magnolol suppresses proliferation of cultured human colon and liver cancer cells by inhibiting DNA synthesis and activating apoptosis

Magnolol, a hydroxylated biphenyl compound isolated from the Chinese herb Hou p'u of Magnolia officinalis, has been reported to have anti-cancer activity. In the present study, magnolol at very low concentrations of 3-10 microM inhibited DNA synthesis and decreased cell number in cultured human cancer cells (COLO-205 and Hep-G2) in a dose-dependent manner, but not in human untransformed cells such as keratinocytes, fibroblasts, and human umbilical vein endothelial cells (HUVEC). Magnolol was not cytotoxic at these concentrations and this indicates that it may have an inhibitory effect on cell proliferation in the subcultured cancer cell lines. [(3)H] thymidine incorporation and flow cytometry analyses revealed that magnolol treatment decreased DNA synthesis and arrested the cells at the G0/G1 phase of the cell cycle. Moreover, the magnolol-induced cell cycle arrest occurred when the cyclin-CDK system was inhibited, just as p21 protein expression was augmented. When magnolol concentration was increased to 100 microM, apoptosis was observed in COLO-205 and Hep-G2 cells, but not in cultured human fibroblasts and HUVEC. COLO-205 cells implanted subcutaneously in nude mice formed solid tumors; subsequent daily i.p.-injections of magnolol led to profound regression of these tumors of up to 85%. In these tumors, an increase in the expression of p21 protein level and the occurrence of apoptosis were observed. These findings demonstrate for the first time that magnolol can inhibit the proliferation of tumor cells in vitro and in vivo.     

Cellular Ras and Cyclin D1 Are Required during Different Cell Cycle Periods in Cycling NIH 3T3 Cells

Novel techniques were used to determine when in the cell cycle of proliferating NIH 3T3 cells cellular Ras and cyclin D1 are required. For comparison, in quiescent cells, all four of the inhibitors of cell cycle progression tested (anti-Ras, anti-cyclin D1, serum removal, and cycloheximide) became ineffective at essentially the same point in G1 phase, approximately 4 h prior to the beginning of DNA synthesis. To extend these studies to cycling cells, a time-lapse approach was used to determine the approximate cell cycle position of individual cells in an asynchronous culture at the time of inhibitor treatment and then to determine the effects of the inhibitor upon recipient cells. With this approach, anti-Ras antibody efficiently inhibited entry into S phase only when introduced into cells prior to the preceding mitosis, several hours before the beginning of S phase. Anti-cyclin D1, on the other hand, was an efficient inhibitor when introduced up until just before the initiation of DNA synthesis. Cycloheximide treatment, like anti-cyclin D1 microinjection, was inhibitory throughout G1 phase (which lasts a total of 4 to 5 h in these cells). Finally, serum removal blocked entry into S phase only during the first hour following mitosis. Kinetic analysis and a novel dual-labeling technique were used to confirm the differences in cell cycle requirements for Ras, cyclin D1, and cycloheximide. These studies demonstrate a fundamental difference in mitogenic signal transduction between quiescent and cycling NIH 3T3 cells and reveal a sequence of signaling events required for cell cycle progression in proliferating NIH 3T3 cells.     

Butyrate inhibits proliferation-induced Proliferating Cell Nuclear Antigen expression (PCNA) in rat vascular smooth muscle cells

Arterial injury-induced vascular smooth muscle cell (VSMC) proliferation in intima is the important etiologic factor in vascular proliferative disorders such as atherosclerosis, hypertension and restenosis after balloon angioplasty. Butyrate, a naturally occurring short chain fatty acid, is produced by bacterial fermentation of dietary fiber and by mammary glands of certain mammals. Studies have shown that butyrate at millimolar concentrations, which are physiological, induces growth arrest, differentiation and apoptosis. We examined the effect of physiological concentrations of butyrate on rat VSMC proliferation and proliferation-induced PCNA expression to determine anti-atherogenic potential of butyrate. Butyrate concentrations, closer to physiological range, exhibited antiproliferative effects on both serum-induced proliferation of serum-starved quiescent VSMCs and actively proliferating non-confluent VSMCs. Treatment of serum-starved quiescent VSMCs with 1-8 mmol/l concentration of butyrate caused a concentration-dependent decrease in serum-induced VSMC proliferation and cell proliferation-associated increase in total cellular proteins and RNA levels. Similarly, exposure of actively growing VSMCs to 5 mmol/l butyrate resulted in the inhibition of cell proliferation and proliferation-induced increase in cellular proteins and RNA levels. Furthermore, cellular morphology was significantly altered. Analysis of cell cycle regulatory proteins indicated that levels of PCNA, an excellent marker for cell proliferation, was significantly altered by butyrate both in actively proliferating and serum-induced quiescent VSMCs. These observations suggest that butyrate exhibits potential antiatherogenic capability by inhibiting VSMC proliferation and proliferation-associated increase in PCNA expression and thus merits further investigations regarding therapeutic significance of butyrate in vascular proliferative disorders.     

Hesperetin, a bioflavonoid, inhibits rat aortic vascular smooth muscle cells proliferation by arresting cell cycle

Diet can be one of the most important factors that influence risks for cardiovascular diseases. Hesperetin, a flavonoid present in grapefruits and oranges, is one candidate that may benefit the cardiovascular system. In this study, we have investigated the effect of hesperetin on the platelet-derived growth factor (PDGF)-BB-induced proliferation of primary cultured rat aortic vascular smooth muscle cells (VSMCs). Hesperetin significantly inhibited 50 ng/ml PDGF-BB-induced rat aortic VSMCs proliferation and [(3)H]-thymidine incorporation into DNA at concentrations of 5, 25, 50, and 100 microM. In accordance with these findings, hesperetin revealed blocking of the PDGF-BB-inducible progression through G(0)/G(1) to S phase of the cell cycle in synchronized cells. Western blot showed that hesperetin inhibited not only phosphorylation of retinoblastoma protein (pRb) and expressions of cyclin A, cyclin D, cyclin E, cyclin-dependent kinase 2 (CDK2) as well as proliferating cell nuclear antigen (PCNA) protein, but also downregulation of cyclin-dependent kinase inhibitor (CKI) p27(kip1), while did not affect CKI p21(cip1), p16(INK4), p53, and CDK4 expressions as well as early signaling transductions such as PDGF beta-receptor, extracellular signal-regulated kinase (ERK) 1/2, Akt, p38, and JNK phosphorylation. These results suggest that hesperetin inhibits PDGF-BB-induced rat aortic VSMCs proliferation via G(0)/G(1) arrest in association with modulation of the expression or activation of cell-cycle regulatory proteins, which may contribute to the beneficial effect of grapefruits and oranges on cardiovascular system.     

Time- and concentration-dependent effects of resveratrol in HL-60 and HepG2 cells

Resveratrol, a phytochemical present in grapes, has been demonstrated to inhibit tumourigenesis in animal models. However, the specific mechanism by which resveratrol exerts its anticarcinogenic effect has yet to be elucidated. In the present study, the inhibitory effects of resveratrol on cell proliferation and apoptosis were evaluated in the human leukaemia cell line HL-60 and the human hepatoma derived cell line HepG2. We found that after a 2 h incubation period, resveratrol inhibited DNA synthesis in a concentration-dependent manner. The IC50 value was 15 microm in both HL-60 and HepG2 cells. When the time of treatment was extended, an increase in IC50 value was observed; for example, at 24 h the IC50 value was 30 microm for HL-60 cells and 60 microm for HepG2 cells. Flow cytometry revealed that cells accumulated in different phases of the cell cycle depending on the resveratrol concentration. Furthermore, an increase in nuclear size and granularity was observed in the G1 and S phases of HL-60 treated and HepG2-treated cells. Apoptosis was also stimulated by resveratrol in a concentration-dependent manner in HL-60 and HepG2 cells. In conclusion, resveratrol inhibits cell proliferation in a concentration- and time-dependent manner by interfering with different stages of the cell cycle. Furthermore, resveratrol treatment causes stimulation of apoptosis as well as an increase in nuclear size and granularity.     

Arachidonic acid-derived oxidation products initiate apoptosis in vascular smooth muscle cells

The mechanism of arachidonic acid (AA)-induced apoptosis in vascular smooth muscle cells (VSMCs) was studied in the A-10 rat aortic smooth muscle cell line. Treatment of serum-deprived VSMCs with 50 microM AA for 24 h resulted in a loss of cell viability. The apoptotic effect of AA was characterized by annexin V binding, sub-G1 population of cells, cell shrinkage and chromatin condensation. AA-induced VSMC death was attenuated by antioxidants alpha-tocopherol and glutathione, the hydrogen peroxide (H2O2) scavenger catalase and by serum proteins, albumin and gamma globulins. Moreover, the AA peroxidation products, 12(S)-hydroperoxyeicosatetraenoic acid (HPETE), 15(S)-HPETE, 4-hydroxy-2-nonenal (HNE) and malondialdehyde (MDA) caused VSMC apoptosis. These data suggest an oxidative mechanism of AA-induced VSMC death. The apoptotic effect of AA was pH-dependent, being inhibited by extracellular alkalinization to pH 8.0. AA inhibited serum-stimulated cell cycle progression in quiescent cells, but not in proliferating cells. In conclusion, AA, through its oxidation products causes VSMC apoptosis. Antioxidants, by inhibiting VSMC apoptosis, may prevent consequent pathological events such as atherosclerotic plaque rupture.     

Resveratrol-induced limitation of dysfunction of mitochondria isolated from rat brain in an anoxia-reoxygenation model

Resveratrol protection on the main functions of purified rat brain mitochondria submitted to anoxia-reoxygenation was investigated. Resveratrol (<0.1 microM) reversed partly (23.3%) the respiratory control ratio (RCR) decrease by protecting both states 3 and 4. This effect was both observed when resveratrol was added before anoxia or reoxygenation. Resveratrol fully inhibited the release of cytochrome c in a concentration-dependent manner and significantly decreased the superoxide anion (O2(0-)) production at a concentration of 1 nM. The mitochondrial membranes damaged after the anoxia-reoxygenation were partly protected (about 70%) by resveratrol at 0.1 microM. The oxygen consumption of mitochondria in presence of NADH and cytochrome c was significantly inhibited by resveratrol with a low EC50 of 18.34 pM. Resveratrol inhibited the CCCP-induced uncoupling from about 20%. The effects of resveratrol on oxidative phosphorylation parameters were also investigated in rats after pretreatment (0.4, 2 and 10 mg/kg/day) for one week. After the isolation of brain mitochondria, the RCR was significantly less decreased in the resveratrol group compared to the control group. These results showed that resveratrol could preserve the mitochondrial functions with at least three mechanisms: antioxidant properties, action on complex III and a membrane stabilizing effect.     

Dysregulation of CREB binding protein triggers thrombin-induced proliferation of vascular smooth muscle cells

Thrombin is a potent mitogen for vascular smooth muscle cells (VSMCs). CBP has been regarded as a potential therapeutic target on the basis of its ability to affect cell growth. Therefore we hypothesized that CBP mediates thrombin-induced proliferation of VSMCs. We constructed recombinant adenoviral vector that expresses four short hairpin RNA (shRNA) targeting rat CBP mRNA (CBP-shRNA/Ad). VSMCs were infected with CBP-shRNA/Ad and treated with thrombin. CBP level were analyzed by quantitative real-time PCR and Western blot. To evaluate VSMC proliferation, the cell cycle and DNA synthesis were analyzed by flow cytometry and (3)H-thymidine incorporation, respectively. CBP-shRNA/Ad infection inhibited thrombin-induced CBP expression in a dose-dependent manner concomitant with a decrease in the percentage of cells in the S phase and in DNA synthesis. These findings suggest that CBP plays a pivotal role in the S phase progression of VSMCs.     

Resveratrol depletes mitochondrial DNA and inhibition of autophagy enhances resveratrol-induced caspase activation

We recently demonstrated that resveratrol induces caspase-dependent apoptosis in multiple cancer cell types. Whether apoptosis is also regulated by other cell death mechanisms such as autophagy is not clearly defined. Here we show that inhibition of autophagy enhanced resveratrol-induced caspase activation and apoptosis. Resveratrol inhibited colony formation and cell proliferation in multiple cancer cell types. Resveratrol treatment induced accumulation of LC3-II, which is a key marker for autophagy. Pretreatment with 3-methyladenine (3-MA), an autophagy inhibitor, increased resveratrol-mediated caspase activation and cell death in breast and colon cancer cells. Inhibition of autophagy by silencing key autophagy regulators such as ATG5 and Beclin-1 enhanced resveratrol-induced caspase activation. Mechanistic analysis revealed that Beclin-1 did not interact with proapoptotic proteins Bax and Bak; however, Beclin-1 was found to interact with p53 in the cytosol and mitochondria upon resveratrol treatment. Importantly, resveratrol depleted ATPase 8 gene, and thus, reduced mitochondrial DNA (mtDNA) content, suggesting that resveratrol induces damage to mtDNA causing accumulation of dysfunctional mitochondria triggering autophagy induction. Together, our findings indicate that induction of autophagy during resveratrol-induced apoptosis is an adaptive response.     

Effects of resveratrol and 4-hexylresorcinol on hydrogen peroxide-induced oxidative DNA damage in human lymphocytes

The protective effects of resveratrol and 4-hexylresorcinol against oxidative DNA damage in human lymphocytes induced by hydrogen peroxide were investigated. Resveratrol and 4-hexylresorcinol showed no cytotoxicity to human lymphocytes at the tested concentration (10-100 μM). In addition, DNA damage in human lymphocytes induced by H 2 O 2 was inhibited by resveratrol and 4-hexylresorcinol. Resveratrol and 4-hexylresorcinol at concentrations of 10-100 μM induced an increase in glutathione (GSH) levels in a concentration-dependent manner. Moreover, these two compounds also induced activity of glutathione peroxidase (GPX) and glutathione reductase (GR). The activity of glutathione-S-transferase (GST) in human lymphocytes was induced by resveratrol. Resveratrol and 4-hexylresorcinol inhibited the activity of catalase (CAT). These data indicate that the inhibition of resveratrol and 4-hexylresorcinol on oxidative DNA damage in human lymphocytes induced by H 2 O 2 might be attributed to increase levels of GSH and modulation of antioxidant enzymes (GPX, GR and GST).