Protective effects of resveratrol on calcium-induced oxidative stress in rat heart mitochondria

Trans-resveratrol is a nutraceutical with known antioxidant, anti-inflammatory, cardioprotective, and anti-apoptotic properties. The aim of this study was to evaluate the effects of resveratrol on heart mitochondria. Resveratrol significantly decreased Fe²⁺ + ascorbate oxidant system-induced lipid peroxide levels, preserved physiological levels of glutathione, and increased nitric oxide (NO) levels in mitochondria. Under calcium-mediated stress, there was a 2.7-fold increase in the NO levels, and a mild decoupling in the mitochondrial respiratory chain. These results provide a mechanism for and support the beneficial effects of resveratrol under pathological conditions induced by oxidative stress and calcium overload. In addition, these findings underscore the usefulness of resveratrol in the prevention of cardiovascular diseases.     

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.     

Resveratrol Ameliorates Imiquimod-Induced Psoriasis-Like Skin Inflammation in Mice

BackgroundThe polyphenol resveratrol has anti-inflammatory effects in various cells, tissues, animals and human settings of low-grade inflammation. Psoriasis is a disease of both localized and systemic low-grade inflammation. The Sirtuin1 enzyme thought to mediate the effects of resveratrol is present in skin and resveratrol is known to down regulate NF-κB; an important contributor in the development of psoriasis. Consequently we investigated whether resveratrol has an effect on an Imiquimod induced psoriasis-like skin inflammation in mice and sought to identify candidate genes, pathways and interleukins mediating the effects.MethodsThe study consisted of three treatment groups: A control group, an Imiquimod group and an Imiquimod+resveratrol group. Psoriasis severity was assessed using elements of the Psoriasis Area Severity Index, skin thickness measurements, and histological examination. We performed an RNA microarray from lesional skin and afterwards Ingenuity pathway analysis to identify affected signalling pathways. Our microarray was compared to a previously deposited microarray to determine if gene changes were psoriasis-like, and to a human microarray to determine if findings could be relevant in a human setting.ResultsImiquimod treatment induced a psoriasis-like skin inflammation. Resveratrol significantly diminished the severity of the psoriasis-like skin inflammation. The RNA microarray revealed a psoriasis-like gene expression-profile in the Imiquimod treated group, and highlighted several resveratrol dependent changes in relevant genes, such as increased expression of genes associated with retinoic acid stimulation and reduced expression of genes involved in IL-17 dependent pathways. Quantitative PCR confirmed a resveratrol dependent decrease in mRNA levels of IL-17A and IL-19; both central in developing psoriasis.ConclusionsResveratrol ameliorates psoriasis, and changes expression of retinoic acid stimulated genes, IL-17 signalling pathways, IL-17A and IL-19 mRNA levels in a beneficial manner, which suggests resveratrol, might have a role in the treatment of psoriasis and should be explored further in a human setting.     

Molecular mechanisms of resveratrol (3,4,5-trihydroxy-trans-stilbene) and its interaction with TNF-related apoptosis inducing ligand (TRAIL) in androgen-insensitive prostate cancer cells

Although resveratrol, an active ingredient derived from grapes and red wine, possesses chemopreventive properties against several cancers, the molecular mechanisms by which it inhibits cell growth and induces apoptosis have not been clearly understood. Here, we examined the molecular mechanisms of resveratrol and its interactive effects with TRAIL on apoptosis in prostate cancer PC-3 and DU-145 cells. Resveratrol inhibited cell viability and colony formation, and induced apoptosis in prostate cancer cells. Resveratrol downregulated the expression of Bcl-2, Bcl-X_L and survivin and upregulated the expression of Bax, Bak, PUMA, Noxa, and Bim, and death receptors (TRAIL-R1/DR4 and TRAIL-R2/DR5). Treatment of prostate cancer cells with resveratrol resulted in generation of reactive oxygen species (ROS), translocation of Bax to mitochondria and subsequent drop in mitochondrial membrane potential, release of mitochondrial proteins (cytochrome c, Smac/DIABLO, and AIF) to cytosol, activation of effector caspase-3 and caspase-9, and induction of apoptosis. Resveratrol-induced ROS production, caspase-3 activity and apoptosis were inhibited by N-acetylcysteine. Bax was a major proapoptotic gene mediating the effects of resveratrol as Bax siRNA inhibited resveratrol-induced apoptosis. Resveratrol enhanced the apoptosis-inducing potential of TRAIL, and these effects were inhibited by either dominant negative FADD or caspase-8 siRNA. The combination of resveratrol and TRAIL enhanced the mitochondrial dysfunctions during apoptosis. These properties of resveratrol strongly suggest that it could be used either alone or in combination with TRAIL for the prevention and/or treatment of prostate cancer.     

Resveratrol treatment increases mitochondrial biogenesis and improves viability of porcine germinal-vesicle stage vitrified-warmed oocytes

Vitrification induces mitochondrial dysfunction in warmed oocytes, and degeneration and biogenesis of mitochondria are crucial for maintaining oocyte quality. The present study addresses a hypothesis that treatment of vitrified-warmed oocytes with resveratrol could improve the viability of oocytes by activating mitochondrial biosynthesis. Cumulus oocyte complexes (COCs) collected from gilt ovaries were vitrified, warmed, and cultured in a medium containing vehicle or 1 μM resveratrol. Resveratrol treatment improved survival, maturation, and mitochondrial membrane potential of vitrified-warmed oocytes, but did not improve the development into blastocysts after parthenogenetic activation. Resveratrol treatment increased mitochondrial synthesis, as determined by the expression levels of TOMM20 and mitochondrial DNA copy number, in vitrified-warmed oocytes, but not in non-vitrified oocytes. In addition, the effect of resveratrol treatment on mitochondrial synthesis was reduced by EX527, a SIRT1 inhibitor. Resveratrol treatment of vitrified-warmed oocytes increased the expression levels of genes involved in mitochondrial synthesis (TFAM, POLG, and PGC1α) and increased nuclear translocation of NRF2. Furthermore, vitrification induced mitophagy, as confirmed by a reduction in TOMM20 expression and decreased p62 aggregation, whereas resveratrol treatment did not affect these mitophagic features. In conclusion, vitrification induced mitochondrial clearance in oocytes, whereas activation of SIRT1 by resveratrol treatment facilitated the recovery of vitrified-warmed oocytes through the activation of mitochondrial synthesis.     

Resveratrol Modulates Mitochondria Dynamics in Replicative Senescent Yeast Cells

Mitochondria form a reticulum network dynamically fuse and divide in the cell. The balance between mitochondria fusion and fission is correlated to the shape, activity and integrity of these pivotal organelles. Resveratrol is a polyphenol antioxidant that can extend life span in yeast and worm. This study examined mitochondria dynamics in replicative senescent yeast cells as well as the effects of resveratrol on mitochondria fusion and fission. Collecting cells by biotin-streptavidin sorting method revealed that majority of the replicative senescent cells bear fragmented mitochondrial network, indicating mitochondria dynamics favors fission. Resveratrol treatment resulted in a reduction in the ratio of senescent yeast cells with fragmented mitochondria. The readjustment of mitochondria dynamics induced by resveratrol likely derives from altered expression profiles of fusion and fission genes. Our results demonstrate that resveratrol serves not only as an antioxidant, but also a compound that can mitigate mitochondria fragmentation in replicative senescent yeast cells.     

Zebrafish Tg(hb9:MTS-Kaede): a new in vivo tool for studying the axonal movement of mitochondria

ObjectivesDeregulation of axonal transport in neurons is emerging as the major cause of many neurodegenerative diseases in human, such as Charcot-Marie-Tooth (CMT) neuropathy. However, little is known about how mitochondria move in vivo and whether cell culture systems truly represent what happens in living animals. Here we describe the generation of a new zebrafish transgenic line that specifically allows to study mitochondrial dynamics in motor neurons and its application to analyse mitochondrial movement in zebrafish models expressing CMT2A causing mutations.MethodsThe Tol2 transposon system was used to generate a transgenic zebrafish line expressing the photoconvertible fluorescent protein Kaede in mitochondria of motor neurons. Mitochondrial shape and movement were monitored by time-lapse confocal live imaging and measured by kymograph analysis. The effects of two well-known CMT causing mutations, L76P and R94Q substitutions in MFN2, were then investigated with the same methods.ResultsWe generated the transgenic zebrafish Tg(hb9:MTS-Kaede) line with genetically labelled mitochondria in motor neurons. Kaede protein was correctly and stably targeted to mitochondrial matrix while retaining its photoconvertibility, thus qualifying this model for in vivo studies. Expression of the L76P and R94Q mutations reduced mitochondrial movement in axons and altered mitochondrial distribution in distinct ways.Conclusions and general significanceThese findings confirm previously published data obtained in cell cultures and strengthen the hypothesis of different mechanism of action of the two MFN2 mutations. Considering the number of neurodegenerative diseases associated to mitochondrial dynamics, the Tg(hb9:MTS-Kaede) zebrafish line is a promising model to study in vivo alterations of mitochondrial transport underlying human diseases.     

白藜芦醇对6-羟基多巴所致SN4741细胞损伤的保护作用和机制研究

目的:探讨白藜芦醇对6-羟基多巴引起的细胞损伤的内在保护机制。方法:以SN4741细胞系为实验对象,分为对照组、6-羟基多巴处理组和白藜芦醇预处理、6-羟基多巴处理组组。MTT法测定细胞活性。Western blot检测细胞内DJ-1表达水平。ROS检测反映细胞的氧化应激水平和线粒体损伤情况。线粒体膜电位检测反映细胞线粒体功能。结果:白藜芦醇可以剂量依赖性方式提高6-OHDA诱导的SN4741细胞的存活率。白藜芦醇预处理显著逆转6-OHDA诱导的SN4741细胞DJ-1水平的下降,降低6-OHDA引起的氧化应激水平和线粒体损伤。结论:白藜芦醇预处理能够保护6-羟基多巴所致的SN4741细胞损伤,可能与提高DJ-1的表达,减轻细胞内的氧化应激水平,改善线粒体功能有关。     

HS-1793, a recently developed resveratrol analogue protects rat heart against hypoxia/reoxygenation injury via attenuating mitochondrial damage

Resveratrol is known to exert a cardioprotective effect against hypoxia/reoxygenation (H/R) injury. HS-1793 is a novel, more stable resveratrol analog, but its cardioprotective effects were unknown. The present study aimed to test the cardioprotective effect of HS-1793 against H/R injury and investigate the role of mitochondria in Sprague Dawley rat heart damage using an ex vivo Langendorff system. HS-1793 ameliorated H/R-induced mitochondrial dysfunction by reducing mitochondrial reactive oxygen species production, improving mitochondrial oxygen consumption and suppressing mitochondrial calcium (Ca²⁺) overload during reperfusion. Moreover, HS-1793-treated rat heart showed reduced infarct size. Our data suggest that HS-1793 can protect cardiac against mitochondrial damage following H/R, thereby suppressing injury.     

Resveratrol inhibits collagen-induced platelet stimulation through suppressing NADPH oxidase and oxidative inactivation of SH2 domain-containing protein tyrosine phosphatase-2

Reactive oxygen species (ROS) produced upon collagen stimulation are implicated in propagating various platelet-activating pathways. Among ROS-producing enzymes, NADPH oxidase (NOX) is largely responsible for collagen receptor-dependent ROS production. Therefore, NOX has been proposed as a novel target for the development of antiplatelet agent. We here investigate whether resveratrol inhibits collagen-induced NOX activation and further examine the effects of resveratrol on ROS-dependent signaling pathways in collagen-stimulated platelets. Collagen-induced superoxide anion production in platelets was inhibited by resveratrol. Resveratrol suppressed collagen-induced phosphorylation of p47^phox, a major regulatory subunit of NOX. Correlated with the inhibitory effects on NOX, resveratrol protected SH2 domain-containing protein tyrosine phosphatase-2 (SHP-2) from ROS-mediated inactivation and subsequently attenuated the specific tyrosine phosphorylation of key components (spleen tyrosine kinase, Vav1, Bruton’s tyrosine kinase, and phospholipase Cγ2) for collagen receptor signaling cascades. Resveratrol also inhibited downstream responses such as cytosolic calcium elevation, P-selectin surface exposure, and integrin-αIIbβ3 activation. Furthermore, resveratrol inhibited platelet aggregation and adhesion in response to collagen. The antiplatelet effects of resveratrol through the inhibition of NOX-derived ROS production and subsequent oxidative inactivation of SHP-2 suggest that resveratrol is a potential compound for prevention and treatment of thrombovascular diseases.     

Signaling pathways of prohibitin and its role in diseases

Abstract

Prohibitin (PHB), appearing to be a negative regulator of cell proliferation and to be a tumor suppressor, has been connected to diverse cellular functions including cell cycle control, senescence, apoptosis and the regulation of mitochondrial activities. It is a growth regulatory gene that has pleiotropic functions in the nucleus, mitochondria and cytoplasmic compartments. However, in different tissues/cells, the expression of PHB was different, such as that it was increased in most of the cancers, but its expression was reduced in kidney diseases. Signaling pathways might be very important in the pathogenesis of diseases. This review was performed to provide a relatively complete signaling pathways flowchart for PHB to the investigators who were interested in the roles of PHB in the pathogenesis of diseases. Here, we review the signal transduction pathways of PHB and its role in the pathogenesis of diseases.     

Closer association of mitochondria with lipid droplets in hepatocytes and activation of Kupffer cells in resveratrol-treated senescence-accelerated mice

Resveratrol has been extensively investigated because of its beneficial effects in delaying age-related diseases, thus extending the lifespan, possibly by mimicking calorie restriction. For this study, cell biological techniques were used to examine how resveratrol influenced hepatocytes in a senescence-accelerated mouse P10 (SAMP10), treated from 35 to 55 weeks of age, with special emphasis on the relationship between mitochondria and lipid droplets. Survival ratio, body weight and food intake of SAMP10 did not differ significantly between the control and resveratrol-treated groups. Compared with the control, the treated livers were altered significantly, as follows. Lipid droplets were reduced and mitochondria were increased in number in hepatocytes. Phosphorylation of acetyl-CoA carboxylase and the expression of both the mitochondrial ATP synthase β subunit and Mn superoxide dismutase (SOD2) were increased. Mitochondria, expressing more SOD2, were more tightly associated with lipid droplets, suggesting the enhancement of lipolysis through the activation of mitochondrial functions. Cathepsin D expression was less in hepatocytes but enhanced in Kupffer cells, which were increased in number and size with more numerous lysosome-related profiles. Together, resveratrol may activate mitochondria resulting in consuming lipids, and may also activate Kupffer cells by which a beneficial milieu for hepatocytes may be created. Both might be related to improvement in the functioning of the liver, which is the organ that is central to metabolic regulation.     

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.     

Resveratrol disrupts peroxynitrite-triggered mitochondrial apoptotic pathway: a role for Bcl-2

Resveratrol (3,4′,5-trihydroxystilbene) is a phytochemical believed to be partly responsible for the cardioprotective effects of red wine due to its numerous biological activities. Here, we studied biochemical pathways underlying peroxynitrite-mediated apoptosis in endothelial cells and potential mechanisms responsible for resveratrol cytoprotective action. Peroxynitrite triggered endothelial cell apoptosis through caspases-8, -9 and -3 activation implying both mitochondrial and death receptor apoptotic pathways. Resveratrol was able to prevent peroxynitrite-induced caspases-3 and -9 activation, but not caspase-8 activation. Additionally, peroxynitrite increased intracellular levels of Bax without affecting those of Bcl-2, increasing consequently the Bax/Bcl-2 ratio. This ratio decreased when cells where pre-incubated with 10 and 50 μM resveratrol, mainly due to resveratrol ability per se to increase Bcl-2 intracellular levels without affecting Bax intracellular levels. These results propose an additional mechanism whereby resveratrol may exert its cardioprotective effects and suggest a key role for Bcl-2 in the resveratrol anti-apoptotic action, especially in disrupting peroxynitrite-triggered mitochondrial pathway.     

Resveratrol with antioxidant activity inhibits matrix metalloproteinase via modulation of SIRT1 in human fibrosarcoma cells

AimsResveratrol, a silent information regulator 1 (SIRT1) activator, has been reported to act as an antioxidant contained in red wine and prevent the development of cardiovascular diseases. Histone deacetylase such as SIRT1 is involved in the regulation of lifespan extension. In this study, the effect of resveratrol on matrix metalloproteinases (MMPs) that play an important role in metastasis was examined in human fibrosarcoma cell line, HT1080.Main methodsThe effect of resveratrol on MMPs' activity was evaluated using gelatin zymography. Western blot analysis and RT-PCR assay were used to determine the effect of resveratrol on the expression level of MMP-9, MAPK and SIRT1 proteins and genes, respectively.Key findingsIt was observed that resveratrol exhibited not only antioxidant effects on lipid peroxidation and DNA oxidation but also inhibitory effects on the expression of MMP-2 and 9 in HT1080 cells stimulated with either phorbol myristate acetate or phenazine methosulfate. Furthermore, it was found that treatment with resveratrol decreased the level of MMP-9 expression via down-regulation of p-ERK, c-fos and p65. In addition, the level of SIRT1 gene expression was also enhanced by treatment of resveratrol alone but the level of MMP-9 gene expression was decreased.SignificanceThese results suggest that the activation of SIRT1 in the presence of resveratrol especially inhibits the expression of MMP-9 in HT1080 cells, providing evidence that resveratrol can be a potential candidate for chemoprevention of cancer.     

What Is New for an Old Molecule? Systematic Review and Recommendations on the Use of Resveratrol

Background

Resveratrol is a natural compound suggested to have beneficial health effects. However, people are consuming resveratrol for this reason without having the adequate scientific evidence for its effects in humans. Therefore, scientific valid recommendations concerning the human intake of resveratrol based on available published scientific data are necessary. Such recommendations were formulated after the Resveratrol 2010 conference, held in September 2010 in Helsingør, Denmark.

Methodology

Literature search in databases as PubMed and ISI Web of Science in combination with manual search was used to answer the following five questions: ¹Can resveratrol be recommended in the prevention or treatment of human diseases?; ²Are there observed “side effects” caused by the intake of resveratrol in humans?; ³What is the relevant dose of resveratrol?; ⁴What valid data are available regarding an effect in various species of experimental animals?; ⁵Which relevant (overall) mechanisms of action of resveratrol have been documented?

Conclusions/Significance

The overall conclusion is that the published evidence is not sufficiently strong to justify a recommendation for the administration of resveratrol to humans, beyond the dose which can be obtained from dietary sources. On the other hand, animal data are promising in prevention of various cancer types, coronary heart diseases and diabetes which strongly indicate the need for human clinical trials. Finally, we suggest directions for future research in resveratrol regarding its mechanism of action and its safety and toxicology in human subjects.     

Resveratrol-induced mitochondrial dysfunction and apoptosis are associated with Ca2+ and mCICR-mediated MPT activation in HepG2 cells

Resveratrol, a natural polyphenolic antioxidant, has been reported to possess the cancer chemopreventive potential in wide range by means of triggering tumor cells apoptosis through various pathways. It induced apoptosis through the activation of the mitochondrial pathway in some kinds of cells. In the present reports, we showed that resveratrol-induced HepG2 cell apoptosis and mitochondrial dysfunction was dependent on the induction of the mitochondrial permeability transition (MPT), because resveratrol caused the collapse of the mitochondrial membrane potential (ΔΨ_m) with the concomitant release of cytochrome c (Cyt.c). In addition, resveratrol induced a rapid and sustained elevation of intracellular [Ca²⁺], which compromised the mitochondrial ΔΨ_m and triggered the process of HepG2 cell apoptosis. In permeabilized HepG2 cells, we further demonstrated that the effect of the resveratrol was indeed synergistic with that of Ca²⁺ and Ca²⁺ is necessary for resveratrol-induced MPT opening. Calcium-induced calcium release from mitochondria (mCICR) played a key role in mitochondrial dysfunction and cell apoptosis: (1) mCICR inhibitor, ruthenium red (RR), prevent MPT opening and Cyt.c release; and (2) RR attenuated resveratrol-induced HepG2 cell apoptotic death. Furthermore, resveratrol promotes MPT opening by lowering Ca²⁺-threshold. These data suggest modifying mCICR and Ca²⁺ threshold to modulate MPT opening may be a potential target to control cell apoptosis induced by resveratrol. Xuemei Tian—Foundation item: Chinese National Natural Science Foundation (No.30300455).