Resveratrol protects against 4-hydroxynonenal-induced apoptosis by blocking JNK and c-JUN/AP-1 signaling.

In the present study we have studied the effect of resveratrol in signal transduction mechanisms leading to apoptosis in 3T3 fibroblasts when exposed to 4-hydroxynonenal (HNE). In order to gain insight into the mechanisms of apoptotic response by HNE, we followed MAP kinase and caspase activation pathways; HNE induced early activation of JNK and p38 proteins but downregulated the basal activity of ERK (1/2). We were also able to demonstrate HNE-induced release of cytochrome c from mitochondria, caspase-9, and caspase-3 activation. Resveratrol effectively prevented HNE-induced JNK and caspase activation, and hence apoptosis. Activation of AP-1 along with increased c-Jun and phospho-c-Jun levels could be inhibited by pretreatment of cells with resveratrol. Moreover, Nrf2 downregulation by HNE could also be blocked by resveratrol. Overexpression of dominant negative c-Jun and JNK1 in 3T3 fibroblasts prevented HNE-induced apoptosis, which indicates a role for JNK-c-Jun/AP-1 pathway. In light of the JNK-dependent induction of c-Jun/AP-1 activation and the protective role of resveratrol, these data may show a critical potential role for JNK in the cellular response against toxic products of lipid peroxidation. In this respect, resveratrol acting through MAP kinase pathways and specifically on JNK could have a role other than acting as an antioxidant-quenching reactive oxygen intermediate.     

Resveratrol inhibits phenotypic and functional maturation of murine bone marrow-derived dendritic cells

This study investigated the effects of resveratrol, a natural polyphenol found in grapes and grape products such as wine and having a wide range of biological and pharmacological activities effecting on the phenotypic and functional maturation of bone marrow (BM)-derived dendritic cells (DC). Resveratrol inhibited the expression of costimulatory molecules (CD80 and CD86), and major histocompatibility complex (MHC) classes I and II significantly, and had the same effect dose-dependently on DC. Resveratrol also significantly suppressed the ability of BM-DC to produce intracellular IL-12 p40/p70 and secretory IL-12 p70 in response to lipopolysaccharides (LPS) stimulation. Resveratrol-treated DC were highly efficient in antigen capture via mannose receptor-mediated endocytosis. Also, they were poor stimulators of na?ve allogeneic T-cell proliferation and induced lower levels of IL-2 in responding T cells. These results indicate the immunosuppressive properties of resveratrol, which may be therapeutically useful in controlling chronic immune and/or inflammatory diseases through the down-regulation of DC differentiation and maturation.     

Effect of resveratrol, a natural polyphenolic compound, on reactive oxygen species and prostaglandin production

Resveratrol is a natural molecule with antioxidant action. Moreover, resveratrol is also considered to be a molecule with anti-inflammatory action, an effect attributed to suppression of prostaglandin (PG) biosynthesis. The aim of the present study was to investigate the effects of resveratrol, a polyphenol present in most red wines, on reactive oxygen species formation as well as on arachidonic acid (AA) release, cyclooxygenase expression, and PG synthesis in murine resident peritoneal macrophages. Results show that resveratrol exerted a strong inhibitory effect on superoxide radical (O2-) and hydrogen peroxide (H2O2) produced by macrophages stimulated by lipopolysaccharides (LPS) or phorbol esters (PMA). Resveratrol also significantly decreased [3H]AA release induced by LPS and PMA or by exposure to O2- or H2O2. Resveratrol treatment caused a significant impairment of cyclooxygenase-2 (COX-2) induction stimulated by LPS and PMA or by O2- or H2O2 exposure. These effects of resveratrol on [3H]AA release and COX-2 overexpression were correlated with a marked reduction of PG synthesis. Our results indicate that the antioxidant action of resveratrol affects AA mobilization and COX-2 induction.     

Inhibition of Na⁺-H⁺ exchange as a mechanism of rapid cardioprotection by resveratrol

BACKGROUND AND PURPOSE Resveratrol is a polyphenol abundantly found in grape skin and red wine. In the present study, we investigated whether resveratrol exerts protective effects against cardiac ischaemia/reperfusion and also explored its mechanisms. EXPERIMENTAL APPROACH Infarct size and functional recovery in rat isolated perfused hearts subjected to no-flow global ischaemia followed by reperfusion were measured. Cultured neonatal rat cardiomyocytes were exposed to H(2)O(2) (100 μmol·L(-1)) to induce cell injury. Intracellular ion concentrations were measured using specific dyes. Western blotting was used to quantify protein expression levels. KEY RESULTS In rat isolated perfused hearts, treatment with resveratrol (20 and 100 μmol·L(-1)) 15 min before ischaemia considerably improved left ventricular functional recovery and infarct size. In cultured neonatal rat cardiomyocytes, resveratrol significantly attenuated the increase in reactive oxygen species (ROS) and loss of mitochondrial inner membrane potential. Resveratrol also suppressed the increase in intracellular concentrations of Na(+) ([Na(+)](i)) and Ca(2+) ([Ca(2+)](i)) after H(2)O(2) application; however, it did not suppress the ouabain-induced [Ca(2+) ](i) increase. By measuring changes in intracellular pH recovery after acidification, we also confirmed that acid-induced activation of the Na(+)-H(+) exchanger (NHE) was prevented by pretreatment with resveratrol. Furthermore, resveratrol inhibited the H(2)O(2)-induced translocation of PKC-α from the cytosol to the cell membrane; this translocation is believed to activate NHE. CONCLUSION AND IMPLICATIONS Resveratrol exerts cardioprotection by reducing ROS and preserving mitochondrial function. The PKC-α-dependent inhibition of NHE and subsequent attenuation of [Ca(2+)](i) overload may be a cardioprotective mechanism.     

白藜芦醇对U937细胞基质金属蛋白酶-9转录的抑制作用

目的:观察白藜芦醇对佛波酯诱导的U937细胞中基质金属蛋白酶-9活性的影响,并从蛋白、mRNA及核转录因子激活蛋白-1(AP-1)水平对其影响进行分析。方法:酶谱法测定U937细胞培养上清中MMP-9的活性;Western blot法考察MMP-9蛋白的生成;RT-PCR法检测MMP-9 mRNA的表达;电泳迁移率变动分析法(EMSA)研究核转录因子SP-1的活性。结果:PMA 10nmol/L 可显著诱导无血清培养的U937细胞中MMP-9活性(P<0.01);白藜芦醇在1和10 μmol/L浓度下可抑制 PMA 10 nmol/L诱导的MMP-9活性(P<0.05 和P<0.01);PMA 10 nmol/L可显著诱导U937细胞中MMP-9蛋白生成(P<0.01)和MMP-9 mRNA的表达(P<0.01),白藜芦醇在1、10μmol/L浓度下可抑制PMA 10 nmol/L诱导的MMP-9蛋白生成和MMP-9 mRNA的表达(P<0.05);白藜芦醇在10、1和0.1μmol/L浓度下可抑制PMA诱导的U937细胞中AP-1的活化。结论:白藜芦醇可有效地抑制PMA诱导的U937细胞中MMP-9的活性,其作用可能是通过抑制PMA诱导的U937细胞核转录因子AP-1活化,进而降低MMP-9 mRNA表达,减少MMP-9蛋白生成而实现的。     

Anti-inflammatory mechanisms of resveratrol in activated HMC-1 cells: Pivotal roles of NF-κB and MAPK

Resveratrol is a phytoalexin polyphenolic compound found in various plants, including grapes, berries, and peanuts. Recently, studies have documented various health benefits of resveratrol including cardiovascular and cancer-chemopreventive properties. The aim of the present study was to demonstrate the effects of resveratrol on the expression of pro-inflammatory cytokines, as well as to elucidate its mechanism of action in the human mast cell line (HMC-1). Cells were stimulated with phorbol 12-myristate 13-acetate (PMA) plus A23187 in the presence or absence of resveratrol. To study the possible effects of resveratrol, ELISA, RT-PCR, real-time RT-PCR, Western blot analysis, fluorescence, and luciferase activity assays were used in this study. Resveratrol significantly inhibited the PMA plus A23187-induction of inflammatory cytokines such as tumour necrosis factor (TNF)-α, interleukin (IL)-6 and IL-8. Moreover, resveratrol attenuated cyclooxygenase (COX)-2 expression and intracellular Ca²⁺ levels. In activated HMC-1 cells, phosphorylation of extra-signal response kinase (ERK) 1/2 decreased after treatment with resveratrol. Resveratrol inhibited PMA plus A23187-induced nuclear factor (NF)-κB activation, IκB degradation, and luciferase activity. Resveratrol suppressed the expression of TNF-α, IL-6, IL-8 and COX-2 through a decrease in the intracellular levels of Ca²⁺and ERK 1/2, as well as activation of NF-κB. These results indicated that resveratrol exerted a regulatory effect on inflammatory reactions mediated by mast cells.     

Effect of arsenic on transcription factor AP-1 and NF-kappaB DNA binding activity and related gene expression

Both acute (24 h) and chronic (10-20 week) exposure of human fibroblast cells to low dose sodium arsenite (As(III)) significantly affects activating protein-1 (AP-1) and nuclear factor kappa B (NF-kappa B) DNA binding activity. Short-term treatment with 0.1-5 microM As(III) up-regulates expression of c-Fos and c-Jun and the redox regulators, thioredoxin (Trx) and Redox factor-1 (Ref-1) and activates both AP-1 and NF-kappa B binding. Chronic exposure to 0.1 or 0.5 microM As(III) decreased c-Jun, c-Fos and Ref-1 protein levels and AP-1 and NF-kappa B binding activity, but increased Trx expression. Short term exposure to phorbol 12-myristate 13-acetate (TPA), a phorbol ester tumour promoter, or hydrogen peroxide (H(2)O(2)) also activates AP-1 and NF-kappa B binding. However, pre-treatment with As(III) prevents this increase. These results suggest that As(III) may alter AP-1 and NF-kappa B activity, in part, by up-regulating Trx and Ref-1. The different effects of short- versus long-term As(III) treatment on acute-phase response to oxidative stress reflect changes in the expression of Ref-1, c-Fos and c-Jun, but not Trx.     

The effect of resveratrol and its methylthio-derivatives on NF-κB and AP-1 signaling pathways in HaCaT keratinocytes

BackgroundResveratrol is a natural stilbene derivative whose chemopreventive activity has been well established. Our previous studies have shown that modification of the stilbene backbone with the methylthio group may influence selectivity and inhibitory potency toward P450 isozymes. The aim of this study was to further investigate the mechanism of their potential chemopreventive activity by evaluating the effect of two 4′-methylthio-trans-stilbene derivatives possessing one (3-M-4′-MTS; S2) and two (3,5-DM-4′-MTS; S5) additional methoxy groups on constitutive nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) activation in immortalized human HaCaT keratinocytes.MethodsThe synthesis of MTS was performed as described earlier. Translocation of NF-κB and AP-1 was evaluated by Western blot analysis. Binding of p65 (NF-κB) and c-Jun and c-Fos subunits (AP-1) to consensus oligonucleotide was assessed by ELISA. Real-time PCR and Western blot were used to evaluate COX-2 and iNOS expression.ResultsWe found differential modulation of signaling pathways depending on the stilbene structure after 24 h of cells treatment. The S2 compound, in contrast to S5 and resveratrol, significantly reduced NF-κB activation by blocking the translocation of the p65 subunit to the nucleus, and decreasing IκB kinase activity. All compounds, but particularly S5, increased c-Jun binding to the AP-1 consensus sequence, while c-Fos binding was not affected.ConclusionsWe conclude that methylthiostilbenes differently modulate constitutive signal transduction pathways in HaCaT cells. These observations should be taken into account in designing new stilbene derivatives with potential chemopreventive activity.     

Resveratrol (trans-3,5,4'-trihydroxystilbene) ameliorates experimental allergic encephalomyelitis, primarily via induction of apoptosis in T cells involving activation of aryl hydrocarbon receptor and estrogen receptor

Resveratrol (trans-3,5,4'-trihydroxystilbene), a polyphenolic compound found in plant products, including red grapes, exhibits anticancer, antioxidant, and anti-inflammatory properties. Using an animal model of multiple sclerosis (MS), we investigated the use of resveratrol for the treatment of autoimmune diseases. We observed that resveratrol treatment decreased the clinical symptoms and inflammatory responses in experimental allergic encephalomyelitis (EAE)-induced mice. Furthermore, we observed significant apoptosis in inflammatory cells in spinal cord of EAE-induced mice treated with resveratrol compared with the control mice. Resveratrol administration also led to significant down-regulation of certain cytokines and chemokines in EAE-induced mice including tumor necrosis factor-alpha, interferon-gamma, interleukin (IL)-2, IL-9, IL-12, IL-17, macrophage inflammatory protein-1alpha (MIP-1alpha), monocyte chemoattractant protein-1 (MCP-1), regulated on activation normal T-cell expressed and secreted (RANTES), and Eotaxin. In vitro studies on the mechanism of action revealed that resveratrol triggered high levels of apoptosis in activated T cells and to a lesser extent in unactivated T cells. Moreover, resveratrol-induced apoptosis was mediated through activation of aryl hydrocarbon receptor (AhR) and estrogen receptor (ER) and correlated with up-regulation of AhR, Fas, and FasL expression. In addition, resveratrol-induced apoptosis in primary T cells correlated with cleavage of caspase-8, caspase-9, caspase-3, poly(ADP-ribose) polymerase, and release of cytochrome c. Data from the present study demonstrate, for the first time, the ability of resveratrol to trigger apoptosis in activated T cells and its potential use in the treatment of inflammatory and autoimmune diseases including, MS.     

Modulation of AP-1 by Natural Chemopreventive Compounds in Human Colon HT-29 Cancer Cell Line

PURPOSE: Activator protein-1 (AP-1) has been implicated as playing important roles in apoptosis and cancer development. In this work, we studied several natural chemopreventive compounds for their potential chemopreventive properties in the modulation of AP-1 signaling pathway in HT-29 colon cancer cells. METHODS: The HT-29 cells were transfected with AP-1-luciferase reporter gene, and one of the stable clones (C-4) was used for subsequent experiments. The HT-29 C-4 cells were treated for 1 h with various natural chemopreventive agents and challenged with AP-1 stimulators such as 12-O-tetradecanoylphorbol-13-acetate (TPA) or hydrogen peroxide (H2O2) for 6 h. The c-Jun N-terminal kinase (JNK) was examined to understand the effect of these compounds on the upstream signaling activator of AP-1. The protein expression level of endogenous cyclin D1, a gene that is under the control of AP-1, was also analyzed after treatments with the agents. In addition, cell death induced by these compounds was evaluated by MTS assay [3-(4.5-dimethylthiazol-2-yl)-5-(3-arboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt]. RESULTS: TPA and H2O2 treatments strongly induced AP-1-luciferase activity as expected. Phenethyl isothiocyanate, sulforaphane, curcumin, and resveratrol increased AP-1-luciferase activity dose-dependently and then decreased at higher doses in the presence or absence of TPA. Allyl isothiocyanate and (-)-epigallocatechin-3-gallate (EGCG) increased AP-1-luciferase activity dose-dependently up to 50 and 100 microM. Other tea catechins and procyanidin dimers, however, had little or no effect on AP-1-luciferase activity. The JNK activity was induced by the isothiocyanates and EGCG. Most of the chemopreventive compounds induced cell death in a dose-dependent manner, with the exception of epicatechin (EC) and the procyanidins, which had little effect. The expression of endogenous cyclin D1 protein was well correlated with those of AP-1-luciferase assay. CONCLUSION: Taken together, these results suggest that natural chemopreventive compounds may have differential biological functions on the signal transduction pathways such as AP-1 in the intervention of colon cancer progression and carcinogenesis.