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Resveratrol initiates differentiation of mouse skeletal muscle-derived C2C12 myoblasts
Abstract
Resveratrol is one of the most widely studied bio-active plant polyphenols. While its effect on endothelial blood vessel cells, cancer cells, inflammatory processes and neurodegenerative events is well documented, little is known about the implication of this phytophenol in differentiating processes, particularly in skeletal muscle cells. Here, we report the effects of resveratrol on mouse skeletal muscle-derived cells (C2C12) in either a nondifferentiated (myoblasts) or differentiated state (myotubes) by evaluating resveratrol uptake, cell proliferation, changes in cell shape, and the expression of genes encoding muscle-specific transcription factors or contractile proteins. Resveratrol: (1) rapidly accumulates within cells through passive and facilitated processes; (2) does not strongly affect cell viability, cell cycle and apoptosis; (3) behaves as a pro-differentiating agent as shown by the lengthening of cells, leading to a myotube phenotype; (4) upregulates muscular pro-differentiation markers and transcription factors (myogenin, Scrp3) starting after 12h of exposure and strongly increases heavy chain myosin content after 18h of exposure to resveratrol; (5) increases the Srf transcription factor's transcript level, a target mRNA of the miRNA-133b, which is itself downregulated by this polyphenol. These results put forward new pro-differentiating regulatory properties of resveratrol on skeletal muscles at least partly via modulation of specific miRNAs.
... A series of studies have considered the effect of resveratrol on the myogenic C2C12 cell line [11][12][13]. Resveratrol increased myoblast elongation and differentiation and this was associated with increased expression of the myogenic regulatory factors (MRFs) Myod and Myog [11]. ...
... A series of studies have considered the effect of resveratrol on the myogenic C2C12 cell line [11][12][13]. Resveratrol increased myoblast elongation and differentiation and this was associated with increased expression of the myogenic regulatory factors (MRFs) Myod and Myog [11]. In another study, resveratrol increased the expression of Myf5 and Myod during the proliferation phase and Myog and Myhc during the differentiation phase [12]. ...
... Whilst we have shown beneficial effects in mdx mice, we also investigated resveratrol's ability to affect healthy muscle. Previous in vitro studies demonstrate that resveratrol increases myoblast differentiation in C2C12 cells and in primary myoblasts [11,12,36]. Here we show a significant increase in the average myofibre diameter, particularly in fibres which are larger than average, in the quadriceps muscle of resveratrol treated wildtype mice. ...
Duchenne muscular dystrophy (DMD) is a progressive fatal neuromuscular disorder with no cure. Therapies to restore dystrophin deficiency have been approved in some jurisdictions but long-term effectiveness is yet to be established. There is a need to develop alternative strategies to treat DMD. Resveratrol is a nutraceutical with anti-inflammatory properties. Previous studies have shown high doses (100–400 mg/kg bodyweight/day) benefit mdx mice. We treated 4-week-old mdx and wildtype mice with a lower dose of resveratrol (5 mg/kg bodyweight/day) for 15 weeks. Voluntary exercise was used to test if a lower dosage than previously tested could reduce exercise-induced damage where a greater inflammatory infiltrate is present. We found resveratrol promoted skeletal muscle hypertrophy in wildtype mice. In dystrophic muscle, resveratrol reduced exercise-induced muscle necrosis. Gene expression of immune cell markers, CD86 and CD163 were reduced; however, signalling targets associated with resveratrol’s mechanism of action including Sirt1 and NF-κB were unchanged. In conclusion, a lower dose of resveratrol compared to the dosage used by other studies reduced necrosis and gene expression of inflammatory cell markers in dystrophic muscle suggesting it as a therapeutic candidate for treating DMD.
... Whether this growth promotion observed in fish fed the anthocyanin extract was due to a stimulatory effect on myogenesis needs to be further explored in fish species of aquaculture importance. In this regard, previous studies have reported that polyphenols including resveratrol and (−)-epicatechin promote myogenic differentiation in mammalian-derived C2C12 myoblasts by up-regulating the expression of several myogenic regulatory factors including myf5, myoD, myogenin and mef2 (Gutierrez-Salmean et al., 2014;Kaminski et al., 2012;Laçon;Montesano et al., 2013). In agreement with this statement Myburgh et al. (2012) observed an accelerated skeletal muscle recovery after in vivo administration of grape-derived proanthocyanidolic oligomers in rats with contusion-induced damage. ...
... Previous studies have demonstrated that polyphenols promote myogenic differentiation as well as muscle regeneration in mammalian models. (Bosutti and Degens, 2015;Gutierez-Salmean et al., 2014;Kaminski et al., 2012;Lançon et al., 2012;Montesano et al., 2013;Myburgh et al., 2012). However, whether polyphenols including anthocyanidins exert similar effects in myogenic cells from fish remains largely unknown. ...
... Previous studies have demonstrated that polyphenols promote myogenic differentiation as well as muscle regeneration in mammalian models. (Bosutti and Degens, 2015;Gutierez-Salmean et al., 2014;Kaminski et al., 2012;Lançon et al., 2012;Montesano et al., 2013;Myburgh et al., 2012). However, whether polyphenols including anthocyanidins exert similar effects in myogenic cells from fish remains largely unknown. ...
... Compound 1 was obtained as yellow amorphous powder and its molecular formula was determined as C 24 Table S1) signals corresponded to a backbone of 7,8-pyranoflavone with a methacryloyl group. In addition, the HMBC correlation (Figs. ...
... Four are classified as flavonoids (12)(13)(14)(15), two as moracins (21, 23) and one as a stilbene (30). Although previous research has demonstrated the molecular mechanisms of flavonoids and stilbenes on skeletal muscle health 23,24 , that of moracins has not been explored. Therefore, we focused on investigating the effects of moracins on skeletal muscle cell growth and the underlying mechanisms. ...
Twigs of Morus alba have been used in traditional medicine to treat muscle-related symptoms such as aches, numbness, and stiffness. Despite its clinical use in traditional medicine, its active compounds and mode of action have not yet been investigated. Therefore, we aimed to isolate the compounds from the twigs of M. alba and deduce active compounds, key gene targets, and mechanism of action against sarcopenia using network pharmacology analysis. Using various isolation techniques and spectroscopic methods, 43 phytochemicals, including 3 new flavonoids, were isolated and performed network pharmacology analysis. According to the computational-assistant analysis, 28 compounds, 9 genes, and the PI3K-Akt-mTOR signaling pathway were deduced as expected active compounds (EAC), key targets, and the main signaling pathway. To verify the predicted results, the cell proliferation activities of the EAC were evaluated. Especially, moracin E and M significantly increased by 130% ( p < 0.001) and 57% ( p < 0.05), respectively, which have more than 2- and 1.5-fold stronger effects compared to the control. Furthermore, both increased the expression level of proteins involved in the PI3K-Akt-mTOR signaling pathway and myogenic proteins, including myogenin and MyoD. This study demonstrated that moracin E and M exhibit cell proliferative effects on skeletal muscle cells through the PI3K-Akt-mTOR signaling pathway.
... Murine C2C12 myoblasts were purchased from the European Collection of Cell Cultures (ECACC ; Sigma-Aldrich, St Quentin Fallavier, France). C2C12 cells have the ability to differentiate in myotubes in particular culture conditions (Goto et al., 1999) (Kaminski et al., 2012) . In all experiments, C2C12 cells were seeded at 2,500 cells / cm 2 in Petri dishes (80 cm 2 ). ...
... In all experiments, C2C12 cells were seeded at 2,500 cells / cm 2 in Petri dishes (80 cm 2 ). In all experiments, C2C12 cells were previously cultured for 24 h and further treated for 24 h in the presence of 2,2′-azobis (2-amidinopropane) dihydrochloride (AAPH, 5 mM) with or without PRPE (10 and 50 µg / mL) as previously described (Kaminski et al., 2012) (Ghzaiel et al., 2021) . ROS were produced by the decomposition of AAPH. ...
Sarcopenia is a very disabling age-related disease which affects the mass and the strenght of skelettal muscles. This syndrome has no efficient treatment and is associated with an important oxidative stress which could play important roles in skelettal muscle degeneration. In this context, the cytoprotective activity and the antioxidant properties of a polyphenol-rich plant extract (PRPE) was evaluated in undifferentiated C2C12 murine skeletal muscle cells (myoblasts). PRPE is a potent antioxidant mixture as shown by its Reactive Oxygen Species (ROS) scavenging properties by using the KRL (Kit Radicaux Libres) method and the dihydroethidium (DHE) scavening assay. In addition, PRPE has significant protecting properties in C2C12 cells towards oxidative stress triggered by 2, 2′-azobis (2-amidinopropane) dihydrochloride (AAPH) which is a ROS generator, as measured by different complementary approaches. PRPE counteracts several AAPH-induced cytotoxic effects. PRPE prevents morphological changes evaluated by phase contrast microscopy, decreases the number of dying cells determined by counting in the presence of trypan blue and the intracellular ROS overproduction evaluated by flow cytometry after staining with dihydroethidium (DHE). In addition, PRPE tends to normalize the expression of genes (peroxiredoxin 1 (Pdrx1), nuclear factor erythroid 2-related factor 2 (Nrf2), peroxisome proliferator-activator receptor gamma coactivator 1-alpha (Pgc1α)) involved in the oxidant stress defense under ROS exposure. Altogether; our data show that PRPE has potent antioxidant properties and protects C2C12 skeletal muscle cells towards AAPH-induced oxidative stress. These cytoprotective properties of PRPE in skeletal muscle cells submitted to a pro-oxidant environment deserve further investigation in the context of sarcopenia.
... Furthermore, trans-resveratrol also has neurotrophic properties, defined as antioxidant and pro-differentiating activities on nerve cells (Uddin et al., 2020(Uddin et al., , 2021, which make it possible to envisage its use in the treatment of certain neurodegenerative diseases where oxidative stress is increased in association with a decrease in functional mature neurons as is the case in Parkinson's and Alzheimer's disease (Fukutomi et al., 2021;Namsi et al., 2018). Trans-resveratrol's differentiating activities have also been described on C2C12 murine myoblasts, suggesting that trans-resveratrol can also be of interest in the prevention and/or treatment of sarcopenia, which is a highly disabling muscle weakness in the elderly due to the loss of autonomy it causes (Kaminski et al., 2012). Despite its attractive physicochemical characteristics and biological activities, trans-resveratrol has the disadvantage of being rapidly metabolized both at the intestinal level under the influence of Classification of polyphenols. ...
... At lower concentrations, in the range of 1.5À6.25 μM, it will be interesting to evaluate the cytoprotective and neurotrophic activities of AZA-ST. Indeed, at low concentrations, some polyphenols (resveratrol, quercetin, apigenin) are known to induce cell differentiation on different cell types (N2a cells which are murine neuronal cells) (Namsi et al., 2018); C2C12 cells which are murine skeletal muscle cells (Kaminski et al., 2012)) and also to prevent 7-ketocholesterol-induced cell death by oxiapoptophagy involving OXIdative stress, APOPTOsis and autoPHAGY on N2a cells Yammine et al., 2020). Altogether, the data obtained support the hypothesis that AZA-ST could act on the cell signaling involved in the control of the RedOx status, mitochondrial activity, and cell death (apoptosis and autophagy). ...
Polyphenols are natural antioxidant molecules. They are present in several compounds, mainly fruits and vegetable, of the Mediterranean diet and have benefits on human health. Several studies support that they delay the aging process and prevent age-related diseases. However, as polyphenols are rapidly metabolized, their in vivo efficiency can be limited. Having more effective polyphenols in vivo is therefore a major challenge. In this context, aza-stilbenes deriving from resveratrol (trans-resveratrol) have been synthetized. In a first step, the cytotoxic activities of these molecules (AZA-ST I to VII) were compared with those of resveratrol (trans-resveratrol) in a range of concentrations from 1.5 to 100 µM after 48 h of culture on murine neuronal N2a cells with complementary micromethods: fluorescein diacetate assay, which evaluates cell viability, and crystal violet assay, which permits the measure of density of adherent cells. In a second step, the impact of these molecules at the mitochondrial level were determined: the transmembrane mitochondrial potential (ΔΨm) was measured by flow cytometry after staining with DiOC6(3) and the mitochondrial production of superoxide anions was quantified by flow cytometry after staining with MitoSOX. Altogether, comparatively to resveratrol, our data show different levels of cytotoxicity of AZA ST I to VII with 50% inhibiting concentrations (IC50) values in the range of 6.25–25 µM. In addition, as observed with resveratrol, these cytotoxic effects were associated with a dose-dependent loss of ΔΨm and an overproduction of superoxide anions at the mitochondrial level. In conclusion, the aza-stilbenes I to VII have varying toxicities. Like resveratrol, when they are used at high concentrations, they induce mitochondrial dysfunctions.
... Resveratrol displays a pro-apoptotic effect on SW480 cells [37]. Interestingly, resveratrol shows pro-differentiating effects on skeletal muscle stem cells [38]. ...
... Indeed, resveratrol initiates the early steps of skeletal muscle undifferentiated cells C 2 C 12 in a myoblast state into an ongoing differentiated stage as myotube. Essential tissue specific transcription factors are modulated by resveratrol through putative miRNAs (myf 5, myod1 by miR-20b and srf by miR-133) [38]. ...
In addition to the traditional treatments of cancer and cancer prevention, the use of natural compounds, especially those found in food, should be considered. To clarify if resveratrol has the potential for cancer prevention and the possibility of use in therapy, the following must be taken into account: data from epidemiology, clinical protocol (case and control), preclinical studies (lab animals), use of established cell lines as models of cancer cells, test tube assays (enzymes activities), and requirements of nanotechnologies in order to discover new drugs to fight cancer. From this perspective and future expected advances, more information is needed such as improved efficacy, methods of application, and the synergistic sensitization of resveratrol as an adjuvant. In addition, resveratrol nanoformulation is considered to overcome its weak bioavailability.
... Cell differentiation and myotube formation were evaluated through microscopic examination and immunofluorescence staining of MHC. The elongation index was calculated according to the formula [(L -S)/(L + S), where L and S represent cell length and cell width, respectively] [18]. A total of 50 cells from at least 10 random fields for each treatment group were included for calculation using Image-Pro Plus (Media Cybernetics). ...
... At the same time, cells grown on a differentiation medium were referred to differentiation control. The spindle and elongated shapes are morphological features of muscle cell differentiation, which can be reflected by an elongated index [18]. Like differentiation control, morphological elongation of C2C12 cells and immunofluorescence detection of increased MHC expression were found in AG-and teaghrelin-treated cells ( Fig. 2A). ...
... Both differentiating C2C12 myoblasts and mature myotubes were then investigated over time in GR conditions in the absence or presence of Sirtuin activator resveratrol and SIRT1 inhibitor, EX-527. These conditions were compared to conditions of 4.5 g/L (25 mM) high-glucose differentiation (DM) media that is universally and routinely used for both proliferating and differentiating skeletal muscle cells in vitro, with high-glucose DMEM used extensively in studies investigating the effect of resveratrol in C2C12 myoblasts [17,[31][32][33][34]. Overall, we hypothesised that (1) resveratrol treatment would help preserve differentiation and myotube hypertrophy in differentiating myoblasts under glucose restriction, and (2) resveratrol would improve myotube survival and prevent myotube atrophy in mature differentiated myotubes following glucose restriction (GR). ...
... The excess gelatin was aspirated and cells were seeded at 8 × 10 4 cells/ml in 2 ml of GM per well, these were then incubated for 24 h until 80% confluency was attained. Experiments were initiated by removing GM, washing twice with phosphate buffered saline (PBS) followed by the addition of low serum differentiation media for a total period of 7 days, containing: 8.3 g/L of DMEM (D5030, Sigma-Aldrich, UK), 0.584 g/L l-Glutamine, 3.7 g/L Sodium Bicarbonate, 0.11 g/L Sodium Pyruvate, 0.0159 g/L Phenol red, 2% horse serum and 1% Pen-strep and either 0.6 g/L or 3.3 mM (glucose restricted/GR) or 4.5 g/L (25 mM) highglucose differentiation medium (DM) universally used for myoblast proliferation and differentiation and used extensively in previous studies assessing the role of resveratrol in C2C12 cells [17,[31][32][33][34], whilst also allowing relevant comparisons with the existing literature. The reduction in serum content causes C 2 C 12 myoblasts to undergo spontaneous differentiation without requiring the addition of growth factors to initiate the process [35]. ...
Glucose restriction (GR) impairs muscle cell differentiation and evokes myotube atrophy. Resveratrol treatment in skeletal muscle cells improves inflammatory-induced reductions in skeletal muscle cell differentiation. We therefore hypothesised that resveratrol treatment would improve muscle cell differentiation and myotube hypertrophy in differentiating C2C12 myoblasts and mature myotubes during GR. Glucose restriction at 0.6 g/L (3.3 mM) blocked differentiation and myotube hypertrophy versus high-glucose (4.5 g/L or 25 mM) differentiation media (DM) conditions universally used for myoblast culture. Resveratrol (10 µM) treatment increased SIRT1 phosphorylation in DM conditions, yet did not improve differentiation when administered to differentiating myoblasts in GR conditions. Resveratrol did evoke increases in hypertrophy of mature myotubes under DM conditions with corresponding elevated Igf-I and Myhc7 gene expression, coding for the ‘slow’ type I MYHC protein isoform. Inhibition of SIRT1 v
... Committed myoblast could remain responsive to environmental stimuli in order to determine cell fate between progressing toward terminal differentiation and keeping the cell poised to rapidly differentiate (Chapalamadugu et al., 2009;Olguín et al., 2004;Olguín and Pisconti, 2012). Previous studies have reported that polyphenols including resveratrol can either promote or inhibit myogenic differentiation in a dose-dependent manner in C2C12 myoblasts (Abdulla et al., 2013;Bosutti and Degens, 2015;Kaminski et al., 2012;Montesano et al., 2013). Low resveratrol concentrations (10 to 25 μM) have been reported to stimulate myoblast cell cycle arrest and favor further commitment toward terminal differentiation in C2C12 myoblasts (Bosutti and Degens, 2015;Kaminski et al., 2012;Montesano et al., 2013). ...
... Previous studies have reported that polyphenols including resveratrol can either promote or inhibit myogenic differentiation in a dose-dependent manner in C2C12 myoblasts (Abdulla et al., 2013;Bosutti and Degens, 2015;Kaminski et al., 2012;Montesano et al., 2013). Low resveratrol concentrations (10 to 25 μM) have been reported to stimulate myoblast cell cycle arrest and favor further commitment toward terminal differentiation in C2C12 myoblasts (Bosutti and Degens, 2015;Kaminski et al., 2012;Montesano et al., 2013). On the contrary, higher concentrations (above 40 μM) have been shown to inhibit myogenic differentiation in myoblast cells (Bosutti and Degens, 2015;Abdulla et al., 2013). ...
... In fact, the canonical Wnt signaling plays a vital role in CM differentiation of human iPSCs [5,16]. Besides, RSV functions as prodifferentiating agent promoting differentiation of mouse skeletal muscle-derived C2C12 myoblasts partly 2 Stem Cells International through enhancing serum response factor (SRF) transcription [17]. MiR-1, a SRF dependent miRNA, is highly expressed in cardiomyocytes and their precursors and regulates cardiomyogenesis [18][19][20]. ...
... Inhibiting SRF directly led to decreased miR-1 expression in the heart of mice [20]. RSV is a demonstrated agent promoting differentiation of mouse myoblasts partly through enhancing SRF transcription [17]. In the current study, we also found that RSV exerted strong regulation over SRF expression and subsequent miR-1 expression. ...
Resveratrol (trans-3,5,4′-trihydroxystilbene) (RSV) is a natural polyphenol with protective effects over cardiac tissues and can affect cell survival and differentiation in cardiac stem cells transplantation. However, whether this agent can affect cardiomyocytes (CMs) differentiation of induced pluripotent stem cells (iPSCs) is not yet clear. This study explored whether RSV can affect CMs differentiation of human iPSCs. Under embryoid bodies (EBs) condition, the effect of RSV on the change of pluripotent markers, endoderm markers, mesoderm markers, and ectoderm markers was measured using qRT-PCR. Under CM differentiation culture, the effect of RSV on CM specific markers was also measured. The regulative role of RSV over canonical Wnt signal pathway and serum response factor- (SRF-) miR-1 axis and the functions of these two axes were further studied. Results showed that RSV had no effect on the self-renewal of human iPSCs but could promote mesoderm differentiation. Under CM differentiation culture, RSV could promote CM differentiation of human iPSCs through suppressing canonical Wnt signal pathway and enhancing SRF-miR-1 axis.
... Myogenesis involves muscle stem cells and progenitor cells to proliferate as myoblasts and differentiate into myotubes [29]. Activated satellite cells migrate from an undamaged muscle to a damaged muscle when cells are injured and then fuse to form new muscle fibers or repair damaged fibers together with existing fibers [30,31]. ...
This study investigated the effects and active compounds of silkworm pupae, an edible insect, on C2C12 muscle differentiation. The protein of silkworm pupae was extracted using sonication after defatting with hexane. Subsequently, the extract was rehydrated using Alcalase to obtain a protein hydrolysate. The silkworm pupae protein hydrolysate effectively promoted C2C12 myogenic differentiation without cytotoxicity. Subsequently, the hydrolysate was fractionated into four subfractions using preparative high-performance liquid chromatography (Prep-HPLC). Subfraction 1 was the most effective in promoting C2C12 myogenic differentiation and significantly upregulated the expression of myoblast transcription factors, 1.5-fold of myoblast determination protein 1 (MyoD), 2-fold of myogenin, and 3-fold of myosin heavy chain (MyHC). Liquid chromatography–tandem mass spectrometry (LC-MS/MS) and multivariate statistical analysis were used to identify the active peptides in silkworm pupae responsible for the observed effects; then, dipeptides and essential amino acids, such as isoleucine (Ile), valine (Val), and methionine (Met), were identified. In addition, Val, Ile, and two dipeptides underwent quantification to determine the potential bioactive peptides that enhanced C2C12 myogenic differentiation. This study suggests that the peptides from silkworm pupae could be used as a nutraceutical to enhance muscle growth.
... This can have important applications in the context of regenerative medicine. In addition, as resveratrol has also been shown to induce the differentiation of murine myoblasts C2C12 in myotubes [39], there is also an interest to further evaluate the ability of aza-stilbenes to act on the differentiation of skeletal muscle cells. This can also have important applications in the context of aging for the treatment of sarcopenia, which is characterized by a decrease of the muscular mass in the elderly [40]. ...
Trans-resveratrol is a natural polyphenol showing numerous biological properties, especially anti-tumoral and antioxidant activity. Among numerous resveratrol derivatives, aza-stilbenes, which bear an imine bound, show interesting biological activities. In the present study, we synthesized a series of imine analogs of trans-resveratrol (seven aza-stilbenes) following an easy and low-cost procedure of green chemistry. The toxicity of synthesized aza-stilbenes, which is currently unknown, was evaluated on murine neuronal N2a cells, comparatively to trans-resveratrol, by considering: cell density evaluated by staining with sulforhodamine 101; esterase activity, which is a criteria of cell viability, by staining with fluorescein diacetate; and transmembrane mitochondrial potential, which is known to decrease during cell death, by staining with DiOC6(3) using flow cytometry. In addition, the antioxidant activity was quantified with the KRL (Kit Radicaux Libres) assay, the DPPH (2,2′-diphenyl-1-picrylhydrazyl radical) assay and the FRAP (ferric reducing antioxidant power) assay. The PAOT (Pouvoir Antioxidant Total) score was also used. The aza-stilbenes provide different cytotoxic and antioxidant activities, which are either higher or lower than those of trans-resveratrol. Based on their cytotoxic and antioxidant characteristics, all synthesized aza-stilbenes are distinguished from trans-resveratrol.
... Researchers have studied its health-promoting effects of neuroprotection (1) and cardioprotection (2) as well as its inhibiting actions to tumor cell proliferation (3) and microbial activity (4) and its diminishing effects on inflammation in humans and animals (5,6). Its pro-differentiation properties to human lung fibroblasts (7), embryonic cardiomyoblasts (8), and skeletal myoblast have also been studied (9). For example, Dirks Naylor (10) demonstrated the RSV effects on skeletal muscle metabolism, protein catabolism, and muscle-related ischemia and reperfusion injury disease in a review study (10). ...
Resveratrol (RSV) has been confirmed to benefit human health. Resveratrol supplemented in the feeds of animals improved pork, chicken, and duck meat qualities. In this study, we identified differentially expressed (DE) messenger RNAs (mRNAs) (n = 3,856) and microRNAs (miRNAs) (n = 93) for the weighted gene co-expression network analysis (WGCNA) to investigate the co-expressed DE mRNAs and DE miRNAs in the primary bovine myoblasts after RSV treatment. The mRNA results indicated that RSV treatments had high correlations with turquoise module (0.91, P-value = 0.01) and blue module (0.93, P-value < 0.01), while only the turquoise module (0.96, P-value < 0.01) was highly correlated with the treatment status using miRNA data. After biological enrichment analysis, the 2,579 DE genes in the turquoise module were significantly enriched in the Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. The top two GO terms were actin filament-based process (GO:0030029) and actin cytoskeleton organization (GO:0030036). The top two KEGG pathways were regulation of actin cytoskeleton (bta04810) and tight junction (bta04530). Then, we constructed the DE mRNA co-expression and DE miRNA co-expression networks in the turquoise module and the mRNA–miRNA targeting networks based on their co-expressions in the key module. In summary, the RSV-induced miRNAs participated in the co-expression networks that could affect mRNA expressions to regulate the primary myoblast differentiation. Our study provided a better understanding of the roles of RSV in inducing miRNA and of the characteristics of DE miRNAs in the key co-expressed module in regulation of mRNAs and revealed new candidate regulatory miRNAs and genes for the beef quality traits.
... These effects may be related to the expression of fatty acid translocase/CD36, carnitine palmitoyltransferase-1, and PGC-1 α [55] SkM regeneration in adult mammals is affected by mechanical damage or chronic diseases such as obesity and diabetes, resulting in the loss of muscle function [56] . RES can play a regulatory role in various chronic diseases, act as a prodifferentiator by up-regulating muscle pro-differentiation markers and transcription factors, myogenin and serum C-reactive protein 3, and even affect muscle-specific miRNA expression, promote myoblast differentiation, and increase myosin heavy chain expression ( Fig. 2 ) [57] . ...
Skeletal muscle (SkM) is essential for body movement, energy metabolism, and material metabolism, and directly influences the quality of human life. Aging, chronic diseases, and strenuous exercise often lead to various health problems associated with SkM, including muscle atrophy, loss of muscle mass and strength, and metabolic disorders. Various natural products(NaPs), mainly resveratrol (RES), quercetin (QUE), ursolic acid (UA), ecdysone (ECD; mainly 20-OH ECD, 20-HE), and vitamin D, have been reported to protect or regulate SkM health. Some of the products are functionally equivalent to sex hormones, and some are even referred to as "plant exercise pills." However, controversy persists regarding the role of NaPs in SkM health. Therefore, this review objectively summarizes the in vivo and vitro biological activities, molecular mechanisms, and clinical research results of studies on NaPs applied in the regulation of SkM health over the past decade. The present review could advance further research on NaPs and SkM health, and facilitate the revelation of new evidence that could facilitate the application of NaPs in ensuring SkM health.
... Importantly, our results indicate that Si-ions at a concentration of 1.0 mM accelerate muscle wound healing by increasing the cell migration rate protecting it from the toxic oxidative damage compared to a positive control. Previous studies had indicated that antioxidants can improve muscle mass recovery [36], enhance viability by reducing muscle cell death [55], and promote the early stages of differentiation by inducing the expression of differentiation markers [28,56,57]. In line with this, our results indicate that Si-ions at a concentration of 0.1 mM can enhance C2C12 cell viability, proliferation, differentiation, and myogenic marker expression, such as MyoG and MyoD, analogous to the antioxidant effect of Si-ions on osteogenesis [41,47,48]. ...
Volumetric muscle loss injuries overwhelm the endogenous regenerative capacity of skeletal muscle, and the associated oxidative damage can delay regeneration and prolong recovery. This study aimed to investigate the effect of silicon-ions on C2C12 skeletal muscle cells under normal and excessive oxidative stress conditions to gain insights into its role on myogenesis during the early stages of muscle regeneration. In vitro studies indicated that 0.1 mM Si-ions into cell culture media significantly increased cell viability, proliferation, migration, and myotube formation compared to control. Additionally, MyoG, MyoD, Neurturin, and GABA expression were significantly increased with addition of 0.1, 0.5, and 1.0 mM of Si-ion for 1 and 5 days of C2C12 myoblast differentiation. Furthermore, 0.1–2.0 mM Si-ions attenuated the toxic effects of H2O2 within 24 h resulting in increased cell viability and differentiation. Addition of 1.0 mM of Si-ions significantly aid cell recovery and protected from the toxic effect of 0.4 mM H2O2 on cell migration. These results suggest that ionic silicon may have a potential effect in unfavorable situations where reactive oxygen species is predominant affecting cell viability, proliferation, migration, and differentiation. Furthermore, this study provides a guide for designing Si-containing biomaterials with desirable Si-ion release for skeletal muscle regeneration.
... For example, while SIRT1 activates vascular endothelial growth factor (VEGF) expression [210,211], treatment with RSV rather downregulates it [212,213]. Likewise, activation of SIRT1 inhibits muscle differentiation and mitochondrial biogenesis [214][215][216][217], whereas RSV rather potentiates muscle differentiation [218,219] and acts as an exercise mimetic [220]. Activation of SIRT1 sensitizes neurons to oxidative stress [221,222] and inhibits neurogenesis [223] while RSV protects neurons against oxidative stress [224,225] and enhances neurogenesis [226,227]. ...
Unlike widely perceived, resveratrol (RSV) decreased the average lifespan and extended only the replicative lifespan in yeast. Similarly, although not widely discussed, RSV is also known to evoke neurite degeneration, kidney toxicity, atherosclerosis, premature senescence, and genotoxicity through yet unknown mechanisms. Nevertheless, in vivo animal models of diseases and human clinical trials demonstrate inconsistent protective and beneficial effects. Therefore, the mechanism of action of RSV that elicits beneficial effects remains an enigma. In a previously published work, we demonstrated structural similarities between RSV and tyrosine amino acid. RSV acts as a tyrosine antagonist and competes with it to bind to human tyrosyl-tRNA synthetase (TyrRS). Interestingly, although both isomers of RSV bind to TyrRS, only the cis-isomer evokes a unique structural change at the active site to promote its interaction with poly-ADP-ribose polymerase 1 (PARP1), a major determinant of cellular NAD+-dependent stress response. However, retention of trans-RSV in the active site of TyrRS mimics its tyrosine-bound conformation that inhibits the auto-poly-ADP-ribos(PAR)ylation of PARP1. Therefore, we proposed that cis-RSV-induced TyrRS-regulated auto-PARylation of PARP1 would contribute, at least in part, to the reported health benefits of RSV through the induction of protective stress response. This observation suggested that trans-RSV would inhibit TyrRS/PARP1-mediated protective stress response and would instead elicit an opposite effect compared to cis-RSV. Interestingly, most recent studies also confirmed the conversion of trans-RSV and its metabolites to cis-RSV in the physiological context. Therefore, the finding that cis-RSV and trans-RSV induce two distinct conformations of TyrRS with opposite effects on the auto-PARylation of PARP1 provides a potential molecular basis for the observed dichotomic effects of RSV under different experimental paradigms. However, the fact that natural RSV exists as a diastereomeric mixture of its cis and trans isomers and cis-RSV is also a physiologically relevant isoform has not yet gained much scientific attention.
... Interestingly, it has been demonstrated that aging is a multifactorial phenomenon linked mainly to the production of free radicals [3,4]. Indeed, emerging studies suggest that peroxisomal function may also be altered with aging and contribute to the pathogenesis of a variety of diseases, including Recently, the effects of RSV on mouse skeletal muscle derived cells (C2C12 cells) in either undifferentiated (myoblasts) or differentiated state (myotubes) have been reported [27][28][29][30]. RSV inhibits protein degradation and attenuates atrophy of skeletal muscle fibers [26,31]. ...
The increased risk of illness and disability is related to the age inevitable biological changes. Oxidative stress is a proposed mechanism for many age-related diseases. The crucial importance of polyphenol pharmacophore for aging process is largely described thanks to its effects on concentrations of reactive oxygen species. Resveratrol (3,5,4′-trihydroxy-trans-stilbene, RSV) plays a critical role in slowing the aging process but has a poor bioavailabity after oral intake. In this present work, a series of RSV derivatives was designed, synthesized, and evaluated as potential antioxidant agents. These derivatives contain substituents with different electronic and steric properties in different positions of aromatic rings. This kind of substituents affects the activity and the bioavailability of these compounds compared with RSV used as reference compound. Studies of Log P values demonstrated that the introduction of halogens gives the optimum lipophilicity to be considered promising active agents. Among them, compound 6 showed the higher antioxidant activity than RSV. The presence of trifluoromethyl group together with a chlorine atom increased the antioxidant activity compared to RSV.
... In vitro studies using C2C12 myoblast cells exposed to different concentrations of resveratrol have shown that the cell cycle is affected, with consequent reduction of cell proliferation [17]. Moreover, studies have shown that resveratrol also stimulates osteogenic differentiation [36]; it inhibits the activity of osteoclasts [27] and protects against bone loss in ovariectomized rats, immobilized rats, and aged rats [35]. ...
Purpose
We aimed at verifying whether resveratrol can decrease cell proliferation and change osteogenic differentiation of cells obtained from patients with type 1 neurofibromatosis (NF1).
Methods
Deciduous dental pulp derived stem cells were isolated from NF1 patient and healthy volunteer. These cells were subjected to increasing concentrations of resveratrol and evaluated for proliferation and mineralization of osteogenic differentiation.
Results
The results showed that resveratrol reduced the difference in proliferation between CNT and NF1 cells in a dose-dependent manner and this property was more prominent in affected cells than in healthy cells. Resveratrol showed no statistically significant changes in mineralization in osteogenic differentiation of NF1 cells, at low doses tested.
Conclusions
In conclusion, in a dose-dependent manner, resveratrol displays interesting properties that could be applied in a possible treatment aimed at decreasing cellular proliferation in neurofibromatosis. Furthermore, it is selective concerning healthy cells and not affecting cell differentiation. Further research to cell selectivity, differentiation to other tissue types, and cell cytotoxicity are needed.
... In addition, polyphenols such as resveratrol and apigenin are now considered as neurotrophines since, on nerve cells, they have both anti-oxidant and differentiating properties [15][16][17]. In addition, resveratrol has been shown to trigger muscle differentiation on mouse-skeletal muscle-derived C2C12 myoblasts [18]. Altogether, these data support that polyphenols have gerontoprotective activities and could be also of interest in preventing frailty syndrome: this is a clinical syndrome in older adults that carries an increased risk for poor health outcomes including falls, incident disability, hospitalization, and mortality [19]. ...
The Mediterranean diet is associated with health benefits due to bioactive compounds such as polyphenols. The biological activities of three polyphenols (quercetin (QCT), resveratrol (RSV), apigenin (API)) were evaluated in mouse neuronal N2a cells in the presence of 7-ketocholesterol (7KC), a major cholesterol oxidation product increased in patients with age-related diseases, including neurodegenerative disorders. In N2a cells, 7KC (50 µM; 48 h) induces cytotoxic effects characterized by an induction of cell death. When associated with RSV, QCT and API (3.125; 6.25 µM), 7KC-induced toxicity was reduced. The ability of QCT, RSV and API to prevent 7KC-induced oxidative stress was characterized by a decrease in reactive oxygen species (ROS) production in whole cells and at the mitochondrial level; by an attenuation of the increase in the level and activity of catalase; by attenuating the decrease in the expression, level and activity of glutathione peroxidase 1 (GPx1); by normalizing the expression, level and activity of superoxide dismutases 1 and 2 (SOD1, SOD2); and by reducing the decrease in the expression of nuclear erythroid 2-like factor 2 (Nrf2) which regulates antioxidant genes. QCT, RSV and API also prevented mitochondrial dysfunction in 7KC-treated cells by counteracting the loss of mitochondrial membrane potential (ΨΔm) and attenuating the decreased gene expression and/or protein level of AMP-activated protein kinase α (AMPKα), sirtuin 1 (SIRT1) and peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) implicated in mitochondrial biogenesis. At the peroxisomal level, QCT, RSV and API prevented the impact of 7KC by counteracting the decrease in ATP binding cassette subfamily D member (ABCD)3 (a peroxisomal mass marker) at the protein and mRNA levels, as well as the decreased expresssion of genes associated with peroxisomal biogenesis (Pex13, Pex14) and peroxisomal β-oxidation (Abcd1, Acox1, Mfp2, Thiolase A). The 7KC-induced decrease in ABCD1 and multifunctional enzyme type 2 (MFP2), two proteins involved in peroxisomal β-oxidation, was also attenuated by RSV, QCT and API. 7KC-induced cell death, which has characteristics of apoptosis (cells with fragmented and/or condensed nuclei; cleaved caspase-3; Poly(ADP-ribose) polymerase (PARP) fragmentation) and autophagy (cells with monodansyl cadaverine positive vacuoles; activation of microtubule associated protein 1 light chain 3–I (LC3-I) to LC3-II, was also strongly attenuated by RSV, QCT and API. Thus, in N2a cells, 7KC induces a mode of cell death by oxiapoptophagy, including criteria of OXIdative stress, APOPTOsis and autoPHAGY, associated with mitochondrial and peroxisomal dysfunction, which is counteracted by RSV, QCT, and API reinforcing the interest for these polyphenols in prevention of diseases associated with increased 7KC levels.
... Dietary compounds have been shown to influence miRNAs levels in skeletal muscle and, given the importance of miRNAs as crucial regulators of skeletal muscle mass, composition and function, they may represent diagnostic or prognostic biomarkers of age-related muscle dysfunctions (Table 1). Modulation of skeletal muscle differentiation [198,226,227] Palmitic Acid miR-29a Insulin resistance and diabetes [228] Vitamin D miR-26a Skeletal muscle regeneration and differentiation [229] Fructose miR-101, miR-30a miR-128a ...
Sarcopenia is a progressive age-related loss of skeletal muscle mass and strength, which may result in increased physical frailty and a higher risk of adverse events. Low-grade systemic inflammation, loss of muscle protein homeostasis, mitochondrial dysfunction, and reduced number and function of satellite cells seem to be the key points for the induction of muscle wasting, contributing to the pathophysiological mechanisms of sarcopenia. While a range of genetic, hormonal, and environmental factors has been reported to contribute to the onset of sarcopenia, dietary interventions targeting protein or antioxidant intake may have a positive effect in increasing muscle mass and strength, regulating protein homeostasis, oxidative reaction, and cell autophagy, thus providing a cellular lifespan extension. MicroRNAs (miRNAs) are endogenous small non-coding RNAs, which control gene expression in different tissues. In skeletal muscle, a range of miRNAs, named myomiRNAs, are involved in many physiological processes, such as growth, development, and maintenance of muscle mass and function. This review aims to present and to discuss some of the most relevant molecular mechanisms related to the pathophysiological effect of sarcopenia. Besides, we explored the role of nutrition as a possible way to counteract the loss of muscle mass and function associated with ageing, with special attention paid to nutrient-dependent miRNAs regulation. This review will provide important information to better understand sarcopenia and, thus, to facilitate research and therapeutic strategies to counteract the pathophysiological effect of ageing.
... Oxidative stress is known to adversely influence myoblast survival and ability to repair damaged muscle [11]. Indeed, in vitro myogenesis is impaired by oxidative stress and supported by a reducing environment [12][13][14][15][16]. Although it is clear that signaling via reactive oxygen species (ROS) regulates myogenesis at numerous points [17], it is imperative to identify specific protein redox sensors that are targets of cellular oxidation and serve to propagate redox signals throughout metabolic and transcriptional networks, orchestrating changes in muscle physiology that ultimately define human healthspan. ...
Muscle aging is accompanied by blunted muscle regeneration in response to injury and disuse. Oxidative stress likely underlies this diminished response, but muscle redox sensors that act in regeneration have not yet been characterized. Calmodulin contains multiple redox sensitive methionines whose oxidation alters the regulation of numerous cellular targets. We have used the CRISPR-Cas9 system to introduce a single amino acid substitution M109Q that mimics oxidation of methionine to methionine sulfoxide in one or both alleles of the CALM1 gene, one of three genes encoding the muscle regulatory protein calmodulin, in C2C12 mouse myoblasts. When signaled to undergo myogenesis, mutated myoblasts failed to differentiate into myotubes. Although early myogenic regulatory factors were present, cells with the CALM1 M109Q mutation in one or both alleles were unable to withdraw from the cell cycle and failed to express late myogenic factors. We have shown that a single oxidative modification to a redox-sensitive muscle regulatory protein can halt myogenesis, suggesting a molecular target for mitigating the impact of oxidative stress in age-related muscle degeneration.
... RSV is widely studied since the 90s. Because of its numerous biological activities, such as anti-oxidant [7], antitumoral [8], antiviral [9], and anti-inflammatory activities [10] and more recently due to its differentiating properties [11,12]. In addition, trans-resveratrol is a neuroprotective agent [13], and acts against platelet aggregation [14]. ...
Several series of natural polyphenols are described for their biological and therapeutic potential. Natural stilbenoid polyphenols, such as trans-resveratrol, pterostilbene and piceatannol are well-known for their numerous biological activities. However, their moderate bio-availabilities, especially for trans-resveratrol, prompted numerous research groups to investigate innovative and relevant synthetic resveratrol derivatives. This review is focused on isosteric resveratrol analogs aza-stilbenes and azo-stilbenes in which the C=C bond between both aromatic rings was replaced with C=N or N=N bonds, respectively. In each series, synthetic ways will be displayed, and structural sights will be highlighted and compared with those of resveratrol. The biological activities of some of these molecules will be presented as well as their potential therapeutic applications. In some cases, structure-activity relationships will be discussed.
... On the other hand, resveratrol, which is known as an anti-inflammatory factor, could not change PAX-7 level by itself. In two other studies consistent with the present study, resveratrol did not significantly change PAX-7 levels (28,29). These researches have been conducted on elderly people, and although old age is different from cancer, only papers on this subject were found. ...
Introduction: Although resistance training and resveratrol is recommended
as a stimulant for muscle regeneration in healthy subjects their effects on
cancer-related cachexia indices are unclear. This study aimed to investigate
the effect of a course of resistance training with resveratrol supplement on
PAX-7 and eMHC in Gastrocnemius muscle tissue of mice bearing colon
cancer induced cachexia.
Methods: This was an experimental study conducted on 20 six-week-old BALB/c mice implanted with CT-26 tumor. The mice were randomly divided into four groups: resistance training, resveratrol, resveratrol with resistance training, and control. The resistance training group performed 6 weeks of progressive resistance training, the resveratrol group received 100 mg/kg resveratrol per day. 48 hours after the last experimental session, the mice were sacrificed and PAX-7 and eMHC protein levels were measured
using ELISA test through specimen taken from gastrocnemius muscles.
Statistical test of one way ANOVA was used for data analysis.
Results: PAX-7 level was significantly lower in resveratrol with resistance
training group compared to control (P=0.009), resveratrol (P=0.005), and
resistance training groups (P=0.002). eMHC was significantly lower in the
resistance training group compared to the control (P=0.08), resistance
training, and resveratrol groups (P=0.002).
Conclusion: Resistance training was not an appropriate intervention for
treating reduced muscular regeneration due to cachexia. However, regarding
reducing the effect of resveratrol supplementation with resistance training on
PAX-7 level, it may be recommended to improve muscular regeneration
... In muscular atrophy, sarcopenia, and other musclerelated diseases, the differentiation of muscle-derived stem cells is inhibited, and with time, the body gradually loses weight. Phytochemicals that have been reported to promote the differentiation of muscle stem cells include catechins (Gutierrez-Salmean et al. 2014;, resveratrol (Kaminski et al. 2012;Dugdale et al. 2017), apigenin (Jang et al. 2017), and canadine . In addition to the induction of differentiation of muscle stem cells, catechins also protect against nuclear apoptosis. ...
Muscle-related disorders, such as sarcopenia and cachexia, caused by aging and chronic diseases can lead to the loss of muscle mass and strength to different degrees, severely affecting human health. Globally, tea is one of the three most popular beverages, and its major active ingredient catechins have been reported to delay muscular atrophy and enhance movement. However, currently, there is no systematic review to elaborate its roles and the associated mechanisms. This article reviews the (1) functions and mechanisms of catechins in the differentiation of myogenic stem cells, biogenesis of mitochondria, synthesis and degradation of proteins, regulation of glucose level, and metabolism of lipids in muscle cells; and (2) effect of catechins on the blood vessels, bones, and nerves that are closely related to the skeletal muscles. Catechins could prevent, mitigate, delay, and even treat muscle-related disorders caused by aging and diseases.
... Accordingly, trans-resveratrol is part of our daily diet and this is a precious chance for our health because this molecule provides numerous biological activities, such as anti-oxidant [11], antitumoral [12], antiviral [13], and anti-inflammatory activities [14]. In addition, trans-resveratrol extents longevity [15], induces cell pro-differentiation [16,17], is a neuroprotective agent [18], and acts against platelet aggregation [19]. Cell targets have already been identified, such as membrane receptors, tyrosine kinases, phosphatases, sirtuins, and p53 anti-oncogene [20]. ...
Trans-resveratrol, the most well-known polyphenolic stilbenoid, is found in grapes and accordingly in wine and it is considered to be beneficial for human health, especially towards the aging-linked cell alterations by providing numerous biological activities, such as anti-oxidant, antitumoral, antiviral, anti-inflammatory, neuroprotective, and platelet anti-aggregation properties. Although trans-resveratrol is a promising molecule, it cannot be considered as a drug, due to its weak bio-availability and fast metabolism. To overcome these weaknesses, several research teams have undertaken the synthesis of innovative trans-resveratrol derivatives, with the aim to increase its solubility in water and pharmacological activities towards cell targets. The aim of this review is to show the chronological evolution over the last 25 years of different strategies to develop more efficient trans-resveratrol derivatives towards organism physiology and, therefore, to enhance various pharmacological activities. While the literature on the development of new synthetic derivatives is impressive, this review will focus on selected strategies regarding the substitution of trans-resveratrol phenyl rings, first with hydroxy, methoxy, and halogen groups, and next with functionalized substituents. The effects on cell functions and dysfunctions of interesting resveratrol analogs will be addressed in this review.
... There is also lot of evidence that several polyphenols have anti-tumor properties (cell cycle delay, apoptosis induction, metastasis prevention) [11,12]. Interestingly, there is now recent evidence that polyphenols (especially resveratrol, a polyphenol of the stilbene family, found in grapes, blackberries, or peanuts for example) have pro-differentiating properties on different cell types: adipocytes, hematopoietic cells, human umbilical cord mesenchymal stem cells, cancer cells (thyroid, glioblastoma, colon), human lung fibroblasts, keratinocytes, embryonic cardiomyoblasts, and myoblasts [13][14][15][16][17][18]. There is also now evidence that many dietary components of the Mediterranean diet (curcumin, resveratrol, and polyunsaturated fatty acids (PUFAs)), and diets enriched with polyphenols and PUFAs, as well as caloric restriction, physical exercise, and learning, are able to induce neurogenesis in the adult brain. ...
In the prevention of neurodegeneration associated with aging and neurodegenerative diseases (Alzheimer’s disease, Parkinson’s disease), neuronal differentiation is of interest. In this context, neurotrophic factors are a family of peptides capable of promoting the growth, survival, and/or differentiation of both developing and immature neurons. In contrast to these peptidyl compounds, polyphenols are not degraded in the intestinal tract and are able to cross the blood–brain barrier. Consequently, they could potentially be used as therapeutic agents in neurodegenerative pathologies associated with neuronal loss, thus requiring the stimulation of neurogenesis. We therefore studied the ability to induce neuronal differentiation of two major polyphenols present in the Mediterranean diet: resveratrol (RSV), a major compound found in grapes and red wine, and apigenin (API), present in parsley, rosemary, olive oil, and honey. The effects of these compounds (RSV and API: 6.25–50 µM) were studied on murine neuro-2a (N2a) cells after 48 h of treatment without or with 10% fetal bovine serum (FBS). Retinoic acid (RA: 6.25–50 µM) was used as positive control. Neuronal differentiation was morphologically evaluated through the presence of dendrites and axons. Cell growth was determined by cell counting and cell viability by staining with fluorescein diacetate (FDA). Neuronal differentiation was more efficient in the absence of serum than with 10% FBS or 10% delipidized FBS. At concentrations inducing neuronal differentiation, no or slight cytotoxicity was observed with RSV and API, whereas RA was cytotoxic. Without FBS, RSV and API, as well as RA, trigger the neuronal differentiation of N2a cells via signaling pathways simultaneously involving protein kinase A (PKA)/phospholipase C (PLC)/protein kinase C (PKC) and MEK/ERK. With 10% FBS, RSV and RA induce neuronal differentiation via PLC/PKC and PKA/PLC/PKC, respectively. With 10% FBS, PKA and PLC/PKC as well as MEK/ERK signaling pathways were not activated in API-induced neuronal differentiation. In addition, the differentiating effects of RSV and API were not inhibited by cyclo[DLeu5] OP, an antagonist of octadecaneuropeptide (ODN) which is a neurotrophic factor. Moreover, RSV and API do not stimulate the expression of the diazepam-binding inhibitor (DBI), the precursor of ODN. Thus, RSV and API are able to induce neuronal differentiation, ODN and its receptor are not involved in this process, and the activation of the (PLC/PKC) signaling pathway is required, except with apigenin in the presence of 10% FBS. These data show that RSV and API are able to induce neuronal differentiation and therefore mimic neurotrophin activity. Thus, RSV and API could be of interest in regenerative medicine to favor neurogenesis.
... It is up-regulated in the differentiation process and has been reported to confer protection against oxidative stress in neural cells (Looyenga et al., 2013). Importantly, resveratrol, a stilbene and one of the main polyphenols present in grape pomace, has exhibited the ability to promote the differentiation of C2C12 cells (Kaminski et al., 2012). ...
Coffee and grape contain various bioactive compounds like polyphenols that may exert beneficial effects, especially antioxidant activity, on human health upon consumption. However, the molecular mechanisms through which these effects are achieved are not fully elucidated. Thus, in the present study in order to investigate these mechanisms, a whole genome expression DNA microarray analysis was carried out in myoblasts treated with polyphenols of coffee and grape pomace at concentrations that improved the redox status. Grape was composed of catechin, epicatechin, cyanidin, malvidin, delphinidin, petunidin, myrtillin, kuromanin, oenin, peonidin, quercetin, gallic acid and caftaric acid as LC-MS revealed, with a total polyphenolic content (TPC) of 648mg of gallic acid equivalents/g of dry matter. Coffee had a TPC of 42.61mg GAE/g coffee and was composed of 3-chlorogenic acid (16.61mg/g), 4- and 5-chlorogenic acids (13.62mg/g), as UHPLC-HRMS revealed. According to the results, grape polyphenols altered mainly the expression of cytoskeleton and differentiation-associated genes, while coffee compounds had a more profound effect, on the expression levels of many metabolic and antioxidant genes possibly through the nuclear factor (erythroid-derived 2) like-2 (Nrf2) pathway.
... Similarly, in vitro experiments showed that MuSCs cultured with dithiothreitol (a reductant agent) display a higher expression of the contractile protein myosin heavy chain as compared with those cultured under oxidized conditions (H 2 O 2 in medium) (341). Similar results were observed on myogenic differentiation in vitro by using the antioxidant resveratrol (160,224). ...
Aims:
A kidney-brain interaction has been described in acute kidney injury but the mechanisms are uncertain. Since we recently described a reno-cerebral reflex, we tested the hypothesis that renal ischemia-reperfusion injury activates a sympathetic reflex that interlinks the renal and cerebral renin-angiotensin axis to promote oxidative stress and progression of the injury.
Results:
Bilateral ischemia-reperfusion activated the intrarenal and cerebral, but not the circulating, renin-angiotensin system, increased sympathetic activity in the kidney and the cerebral sympathetic regulatory regions, and induced brain inflammation and kidney injury. Selective renal afferent denervation with capsaicin or renal denervation significantly attenuated IRI-induced activation of central renin-angiotensin system and brain inflammation. Central blockade of renin-angiotensin system or oxidative stress by intracerebroventricular losartan or tempol reduced the renal ischemic injury score by 65% or 58%, respectively, and selective renal afferent denervation or reduction of sympathetic tone by intracerebroventricular clonidine decreased the score by 42% or 52%, respectively (all P<0.05). Ischemia-reperfusion-induced renal damage and dysfunction persisted after controlling blood pressure with hydralazine.
Innovation:
This study uncovered a novel reflex pathway between ischemic kidney and the brain that sustains renal oxidative stress and local renin-angiotensin system activation to promote ongoing renal damage.
Conclusions:
These data suggest that the renal and cerebral renin-angiotensin axes are interlinked by a reno-cerebral sympathetic reflex that is activated by ischemia-reperfusion, which contributes to ischemia-reperfusion-induced brain inflammation and worsening of the acute renal injury.
... Similarly, in vitro experiments showed that MuSCs cultured with dithiothreitol (a reductant agent) display a higher expression of the contractile protein myosin heavy chain as compared with those cultured under oxidized conditions (H 2 O 2 in medium) (341). Similar results were observed on myogenic differentiation in vitro by using the antioxidant resveratrol (160,224). ...
Skeletal muscle shows high plasticity in response to external demand. Moreover, adult skeletal muscle is capable of complete regeneration after injury, due to the properties of Muscle Stem Cells (MuSCs), the satellite cells, which follow a tightly regulated myogenic program to generate both new myofibers and new MuSCs for further needs. Whereas reactive oxygen species (ROS) and reactive nitrogen species (RNS) have long been associated with skeletal muscle physiology, their implication in the cell and molecular processes at work during muscle regeneration is more recent. This review focuses on redox regulation during skeletal muscle regeneration. An overview of the basics of ROS/RNS and antioxidant chemistry and biology occurring in skeletal muscle is first provided. Then, the comprehensive knowledge on redox regulation of MuSCs and their surrounding cell partners (macrophages, endothelial cells) during skeletal muscle regeneration is presented in normal muscle and in specific physiological (exercise-induced muscle damage, aging) and pathological (muscular dystrophies) contexts. Recent advances in the comprehension of these processes has led to the development of therapeutic assays using antioxidant supplementation, which result in inconsistent efficiency, underlying the need for new tools aimed at precisely deciphering and targeting ROS networks. This review should provide an overall insight of the redox regulation of skeletal muscle regeneration while highlighting the limits of the use of non-specific antioxidants to improve muscle function.
... In C2C12 muscle myoblast cells, resveratrol increases myoblast differentiation by upregulating myogenic regulatory factors such as MyoD and myogenin [210], and increases glucose uptake and glucose transport through activation of AMP-activated protein kinase (AMPK) [211]. In addition, resveratrol treatment induced myoblast apoptosis [212] and enhanced cell survival [213,214]. ...
In recent years, complementary and alternative medicine has become increasingly popular. This trend has not escaped the Duchenne Muscular Dystrophy community with one study showing that 80% of caregivers have provided their Duchenne patients with complementary and alternative medicine in conjunction with their traditional treatments. These statistics are concerning given that many supplements are taken based on purely “anecdotal” evidence. Many nutraceuticals are thought to have anti-inflammatory or anti-oxidant effects. Given that dystrophic pathology is exacerbated by inflammation and oxidative stress these nutraceuticals could have some therapeutic benefit for Duchenne Muscular Dystrophy (DMD). This review gathers and evaluates the peer-reviewed scientific studies that have used nutraceuticals in clinical or pre-clinical trials for DMD and thus separates the credible from the conjecture.
... Even the exact molecular mechanisms are still unknown, dietary composition appears implicated in miRNAs' expression in several tissues including skeletal muscle [105]. Furthermore, there is evidence that polyphenols and other natural compounds, as well as resveratrol, modulate miRNAs expression promoting skeletal muscle differentiation [106]. As recognised, essential amino acids (EAA) stimulate muscle protein synthesis and the ingestion of 10g EAA results in eliciting decreased myostatin and increased expression of miR-1, miR-23a, miR-208b, miR-499 and pri-miR-206 in skeletal muscle within 3 hours of ingestion [105]. ...
Micro-RNAs (miRNAs) are responsible for important and evolutionary-conserved regulatory functions in several cellular processes such as apoptosis, signalling, differentiation and proliferation. There is a growing interest in understanding more clearly the mechanisms regulating activation and suppression of miRNAs expression in benefit of health prevention advancement. It is now acknowledged that physical activity represents one of the most effective preventive agents in chronic-degenerative diseases. Indeed, a regular exercise exerts a great influence on several parameters and biological pathways, both at genomic and post-genomic levels. Recent works have highlighted the effects of structured physical activity on miRNAs modulation. Modulation of MiRNAs, regulated by exercise in human skeletal muscle, depends on typology type, duration and intensity of an exercise performed. The aim of this review is to provide a comprehensive overview of scientific evidence concerning the effects of physical activity on miRNAs and its relevance for chronic-degenerative diseases prevention.
Over the past three decades, studies have shown that consuming polyunsaturated fatty acids (PUFAs) can enhance animal and human health and welfare through biological, biochemical, pathological, and pharmacological impacts. Furthermore, omega-6 plays key roles in the cardiopulmonary system, including promoting airway relaxation and inhibiting atherosclerosis and hypertension. However, findings from investigations of the effects of omega-6 fatty acids on molecular and cellular activity and discussions on their influence on biomarkers are still unclear. Therefore, the present study aimed to evaluate omega-6 fatty acids, the arachidonic acid (AA), and linoleic acid (LA) effects on C2C12 proliferation, myogenesis morphology, and relative myogenic biomarker expression through the Wnt pathway. C2C12 cells were cultured with and without 25, 50, 100, and 150 µM of LA and AA and then subjected to CCK8, Giemsa staining, RT qPCR, Western blotting, and RNA Sequencing. The CCK8 Assay results showed that 25, 50, 100, and 150 µM LA significantly decreased the viability after 72 h for 25, 50, 100, and 150 µM concentrations. Also, AA supplementation decreased cell viability after 24 h for 150 µM, 48 h for 150 µM, and 72 h for 50, 100, and 150 µM concentrations. Moreover, the LA and AA inhibitory effects noticed through Gimesa staining were morphological changes during myoblast differentiation. Both LA and AA showed inhibiting IGF1, Cola1, Col6a2, Col6a1, Itga10, Itga11, SFRP2, DAAM2, and NKD2 effects; however, the depressing effect was higher for AA compared to LA. The previous results were confirmed through Western blotting, which showed that 50 µM LA and AA significantly reduced DAAM2 and SFRP2 protein levels compared to the control. Regarding RNA sequencing results, LA and AA increased the number of differentially expressed (DE) Mt-rRNA and snoRNA; however, the numbers of lncRNA detected decreased compared to the control. Our findings demonstrate that high and moderate LA and AA concentrations reduce primary myoblast proliferation and differentiation. Also, they highlight novel biomarkers and regulatory factors to improve our understanding of how the nutrition of fatty acids can control and modulate the myogenesis and differentiation process through different biomarker families.
Aging is a complex biological process which can be associated with skeletal muscle degradation leading to sarcopenia. The aim of this study consisted i) to determine the oxidative and inflammatory status of sarcopenic patients and ii) to clarify the impact of oxidative stress on myoblasts and myotubes. To this end, various biomarkers of inflammation (C-reactive protein (CRP), TNF-α, IL-6, IL-8, leukotriene B4 (LTB4)) and oxidative stress (malondialdehyde, conjugated dienes, carbonylated proteins and antioxidant enzymes: catalase, superoxide dismutase, glutathione peroxidase) as well as oxidized derivatives of cholesterol formed by cholesterol autoxidation (7-ketocholesterol, 7β-hydroxycholesterol), were analyzed. Apelin, a myokine which contributes to muscle strength, was also quantified. To this end, a case-control study was conducted to evaluate the RedOx and inflammatory status in 45 elderly subjects (23 non-sarcopenic; 22 sarcopenic) from 65 years old and higher. SARCopenia-Formular (SARC-F) and Timed Up and Go (TUG) tests were used to distinguish between sarcopenic and non-sarcopenic subjects. By using red blood cells, plasma and/or serum, we observed in sarcopenic patients an increased activity of major antioxidant enzymes (superoxide dismutase, glutathione peroxidase, catalase) associated with lipid peroxidation and protein carbonylation (increased level of malondialdehyde, conjugated dienes and carbonylated proteins). Higher levels of 7-ketocholesterol and 7β-hydroxycholesterol were also observed in the plasma of sarcopenic patients. Significant differences were only observed with 7β-hydroxycholesterol. In sarcopenic patients comparatively to non-sarcopenic subjects, significant increase of CRP, LTB4 and apelin were observed whereas similar levels of TNF-α, IL-6 and IL-8 were found. The increased plasma level of 7-ketocholesterol and 7β-hydroxycholesterol in sarcopenic patients led us to study the cytotoxic effect of these oxysterols on undifferentiated (myoblasts) and differentiated (myotubes) murine C2C12 cells. With the fluorescein diacetate and sulforhodamine 101 assays, an induction of cell death was observed both on undifferentiated and differentiated cells: the cytotoxic effects were less pronounced with 7-ketocholesterol. In addition, IL-6 secretion was never detected whatever the culture conditions, TNF-α secretion was significantly increased on undifferentiated and differentiated C2C12 cells treated with 7-ketocholesterol- and 7β-hydroxycholesterol, and IL-8 secretion was increased on differentiated cells. 7-ketocholesterol- and 7β-hydroxycholesterol-induced cell death was strongly attenuated by α-tocopherol and Pistacia lentiscus L. seed oil both on myoblasts and/or myotubes. TNF-α and/or IL-8 secretions were reduced by α-tocopherol and Pistacia lentiscus L. seed oil. Our data support the hypothesis that the enhancement of oxidative stress observed in sarcopenic patients could contribute, especially via 7β-hydroxycholesterol, to skeletal muscle atrophy and inflammation via cytotoxic effects on myoblasts and myotubes. These data bring new elements to understand the pathophysiology of sarcopenia and open new perspectives for the treatment of this frequent age-related disease.
Skeletal muscle differentiation is an essential process in embryonic development as well as regeneration and repair throughout the lifespan. It is well-known that dietary fat intake impacts biological and physiological function in skeletal muscle, however, understanding of the contribution of nutritional factors in skeletal muscle differentiation is limited. Therefore, the objective of the current study was to evaluate the effects of free fatty acids (FFAs) on skeletal muscle differentiation in vitro. We used C2C12 murine myoblasts and treated them with various FFAs, which revealed a unique response of angiopoietin-like protein-4 (ANGPTL4) with linoleic acid (LA) treatment that was associated with reduced differentiation. LA significantly inhibited myotube formation and lowered the protein expression of myogenic regulatory factors, including MyoD and MyoG and increased Pax7 during cell differentiation. Next, recombinant ANGPTL4 protein or siRNA knockdown of ANGPTL4 was employed to examine its role in skeletal muscle differentiation, and we confirmed that ANGPTL4 knockdown at day 2 and -6 of differentiation restored myotube formation in the presence of LA. RNA-sequencing analysis revealed that ANGPTL4-mediated inhibition of skeletal muscle differentiation at day 2 as well as LA at day 2 or -6 led to a reduction in Wnt/β-catenin signaling pathways. We confirmed that LA reduced Wnt11 and Axin2 while increasing expression of the Wnt inhibitor, Dkk2. ANGPTL4 knockdown increased β-catenin protein in the nucleus in response to LA and increased Axin2 and Wnt11 expression. Taken together, these results demonstrate that LA induced ANGPTL4 inhibits C2C12 differentiation by suppressing Wnt/β-catenin signaling.
Ripretinib is a multikinase inhibitor drug approved in 2020 by the FDA and in 2021 by EMA for use in the treatment of advanced gastrointestinal stromal tumors (GIST) which have not adequately responded to previous treatments with kinase inhibitors. The most common side effects of the drug are myalgia and fatigue, which likely causes interruption of the treatment or reduction of the dose. Skeletal muscle cells highly depend on ATP to perform their functions and mitochondrial damage may play a role in skeletal muscle toxicity induced by kinase inhibitors. However, the molecular mechanism has not been clearly identified in the literature yet. In this study, it has been aimed to elucidate the role of mitochondria in the toxic effect of ripretinib on skeletal muscle using the mouse C2C12 myoblast-derived myotubes. The myotubes were exposed to ripretinib at the range of 1-20μM concentrations for 24h. To determine the potential role of mitochondrial impairment in ripretinib-induced skeletal muscle toxicity, intracellular ATP level, mitochondrial membrane potential (MMP), mitochondrial ROS production (mtROS), mitochondrial DNA (mtDNA) copy number, and mitochondrial mass were examined after ripretinib treatment. Furthermore, changes in PGC 1α/NRF 1/NRF 2 expression levels that play a role in mitochondrial biogenesis and mitophagy were investigated. Additionally, the mitochondrial electron transport chain (ETC) enzyme activities were evaluated. Lastly, a molecular docking study was done to see ripretinib's possible interaction with DNA polymerase gamma (POLG) which is important for DNA replication in the mitochondria. According to the findings, ripretinib decreases the ATP level and mtDNA copy number, induces loss of MMP, and reduces mitochondrial mass. The activities of the ETC complexes were inhibited with ripretinib exposure which is in line with the observed ATP depletion and MMP loss. The molecular docking study revealed that ripretinib has inhibitory potential against POLG which supports the observed inhibition of mtDNA. The expression of PGC 1α was reduced in the nuclear fraction indicating that PGC-1α was not activated since the NRF 1 expression was reduced and NRF 2 level did not show significant change. Consequently, mtROS production increased in all treatment groups and mitophagy-related gene expressions and Parkin protein expression level were up-regulated at high doses. In conclusion, mitochondrial damage/loss can be one of the underlying causes of ripretinib-induced skeletal muscle toxicity. However, further studies are needed to confirm the results in vivo.
Most diseases are preventable, and they are related to what we eat. Moreover, most doctor visits are for lifestyle-based diseases, which means they can be prevented by adopting a healthy lifestyle. Treating the causes of illness rather than symptoms of the disease is not only safer and cheaper, but it can work better. In this context, the Special Issue on “Nutraceuticals in Immune System” for the journal Molecules was launched in January 2020. Soon after that, the world was ravaged by the COVID-19 pandemic causing a grave health threat. Paradoxically, the immune system can be both friend and foe of COVID-19. The COVID-19 manifests only mild to moderate symptoms for most infected people who recover without hospitalisation, demonstrating the proper functioning of the immune system in fighting such an infection. For some, however, the overactivation of the immune response causing “cytokine storm” has dire consequences, with severe respiratory failure leading to multiple organ failure, which could be fatal. In fact, most deaths from COVID-19 came from organ inflammation due to undesirable immune system responses. As such, the COVID-19 is a good case in point, demonstrating the importance of a healthy immune system.
Nutraceuticals are products derived from food sources with health benefits in addition to the basic nutritional values. Many of them can positively affect and enhance the immune system, which is particularly pertinent in the current turbulent times of COVID-19. Not surprisingly, nutraceutical sales rose dramatically during the pandemic period. However, much research is still needed to understand how natural products interact with the immune system to clarify their chemical compositions, mechanisms of action and effects on health and illnesses.
This Special Issue provided an open forum for researchers to share their research findings in the growing interest of nutraceuticals. We received an overwhelming response with a total of 33 submissions, of which only nine original research papers and ten reviews were accepted after rigorous peer-review. The included articles research into natural substances of interest in nutraceuticals ranging from herbal medicine to vitamins to microbiota-derived metabolites. The investigated immune-related responses include cancer, neurological diseases, gastroenterological disorders, inflammatory conditions, and infections.
We thank the publisher for this excellent opportunity to serve the research community. As academic editors for this Special Issue, it is our pleasure to review these insightful manuscripts first-hand. We thank all the authors for their contributions. The collected works represent our current understanding and latest findings of nutraceuticals in immune system, which we hope will continue to inspire knowledge quests into the field of nutraceuticals.
Postnatal skeletal muscle differentiation from quiescent satellite cells is a highly regulated process, although our understanding of the contribution of nutritional factors in myogenesis is limited. Free fatty acids (FFAs) are known to cause detrimental effects to differentiated skeletal muscle cells by increasing oxidative stress which leads to muscle wasting and insulin resistance in skeletal muscle. In addition, FFAs are thought to act as inhibitors of skeletal muscle differentiation. However, the precise molecular mechanisms underlying the effects of FFAs on skeletal muscle differentiation remains to be elucidated. There is a clear relationship between dietary FFAs and their ability to suppress myogenesis and we propose the hypothesis that the FFA-mediated increase in angiopoietin-like protein 4 (ANGPTL4) may play a role in the inhibition of differentiation. This review discusses the role of FFAs in skeletal muscle differentiation to-date and proposes potential mechanisms of FFA-induced ANGPTL4 mediated inhibition of skeletal muscle differentiation.
The skeletal muscle (SkM) is the largest organ, which plays a vital role in controlling musculature, locomotion, body heat regulation, physical strength, and metabolism of the body. A sedentary lifestyle, aging, cachexia, denervation, immobilization, etc. Can lead to an imbalance between protein synthesis and degradation, which is further responsible for SkM atrophy (SmA). To date, the understanding of the mechanism of SkM mass loss is limited which also restricted the number of drugs to treat SmA. Thus, there is an urgent need to develop novel approaches to regulate muscle homeostasis. Presently, some natural products attained immense attraction to regulate SkM homeostasis. The natural products, i.e., polyphenols (resveratrol, curcumin), terpenoids (ursolic acid, tanshinone IIA, celastrol), flavonoids, alkaloids (tomatidine, magnoflorine), vitamin D, etc. exhibit strong potential against SmA. Some of these natural products have been reported to have equivalent potential to standard treatments to prevent body lean mass loss. Indeed, owing to the large complexity, diversity, and slow absorption rate of bioactive compounds made their usage quite challenging. Moreover, the use of natural products is controversial due to their partially known or elusive mechanism of action. Therefore, the present review summarizes various experimental and clinical evidence of some important bioactive compounds that shall help in the development of novel strategies to counteract SmA elicited by various causes.
Obesity is increasing rapidly worldwide and is accompanied by many complications, including impaired muscle regeneration. Obesity is known to inhibit AMP-activated protein kinase (AMPK) activity, which impedes mitochondrial biogenesis, myogenic differentiation and muscle regeneration. Resveratrol has an effective anti-obesity effect, but its effect on regeneration of muscle in obese mice remains to be tested. We hypothesized that resveratrol activates AMPK and mitochondrial biogenesis to improve muscle regeneration. Male C57BL/6J mice were fed a control diet or a 60% high-fat diet with or without resveratrol supplementation for 8 weeks and, then, the tibialis anterior muscle was subjected to cardiotoxin-induced muscle injury. Muscle tissue was collected at 3 and 7days after injury. We found that resveratrol enhanced both proliferation and differentiation of satellite cells following injury in obese mice. Markers of mitochondrial biogenesis were upregulated in resveratrol-treated mice. In C2C12 myogenic cells, resveratrol activated AMPK and stimulated the expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), which were associated with enhanced myogenic differentiation. Such effects of resveratrol were abolished by AMPKα1 ablation, showing the mediatory roles of AMPK. In summary, dietary resveratrol activates AMPK/PGC-1α axis to facilitate mitochondrial biogenesis and muscle regeneration impaired due to obesity.
Background: Resveratrol (RSV), a phenolic compound, is present in many human dietary sources, such as peanuts, peanut butter, grapes skin, and grape wine. RSV has been widely known for its benefits on human health. Beef from cattle skeletal muscle is one of the main sources of protein for human consumption. Previous studies have also found that pork and chicken qualities are influenced by the feed supplementation with RSV. In addition, our previous study demonstrated the RSV effects on bovine myoblast differentiation using messenger RNA (mRNA) data. In this study, we mainly focused on the influences of RSV on microRNA (miRNA) expression. Method: We used 20 μM RSV to treat primary bovine myoblasts and extracted RNA for miRNA sequencing. After quality control and alignment for clean reads, we conducted quantification and analysis of differentially expressed (DE) miRNAs in the case (RSV-treated) group versus control (non-RSV treated) group. Next, we predicted the target genes for the DE miRNAs and analyzed them for the enrichments of Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. Results: Finally, we identified 93 DE miRNAs (adjusted P-value < 0.05), of them 44 were upregulated and 49 were downregulated. Bta-miR-34c was the most significantly upregulated miRNA. In silico, prediction results indicated 1,869 target genes for the 93 DE miRNAs. GO enrichment analysis for the genes targeted by DE miRNAs revealed two significant GO terms (adjusted P-value < 0.05), in which the most significant one was stereocilium (GO:0032420). KEGG enrichment analysis showed five significant pathways, and the top significant KEGG pathway was the insulin signaling pathway (bta04910) (adjusted P-value < 0.05). Conclusions: This study provided an improved understanding of effects of RSV on primary bovine myoblast differentiation through the miRNA modulations. The results suggested that RSV could promote differentiation of primary bovine myoblast by stimulating the miRNA expressions. The target genes of DE miRNAs were significantly enriched in the insulin signaling pathway, thus potentially contributing to improving muscle leanness by increasing the energy metabolism.
Aging and muscle diseases often lead to a decline in the differentiation capacity of myoblasts, which in turn results in the deterioration of skeletal muscles (SkM) function and impairment of regeneration ability after injury. Theaflavins, the "gold molecules" found in black tea, have been reported to possess various biological activities and have a positive effect on maintaining human health. In this study, we found that among the four theaflavins (TF1, TF2A, TF2B, and TF3), TF1 (20 μM) significantly promoted the fusion index of myoblasts, number of mature myotubes, and degree of myotube development. Combining transcriptomics, bioinformatics and molecular biology experiments, we showed that TF1 may promote myoblast differentiation by 1) regulating withdrawal of myoblasts from the cell cycle, inducing the release of myogenic factors (MyoD, MyoG, and MyHC) and accelerating myogenic differentiation, and 2) regulating the adhesion force of myoblasts and mechanical properties of mature myotubes and promoting the migration, fusion, and development of myoblasts. In conclusion, our study outcomes show TF1 can promote myoblast differentiation and regulates mayotube mechanical properties. It is a potential dietary supplement for the elderly. Our findings provide a new scientific basis for the relationship between tea drinking and aging.
Background and Objective: Physical exercise and nutrition supplements are recommended interventions to reduce cachexia. This study was conducted to determine the effect of resistance training and resveratrol supplementation on muscle regeneration indices of MyoD (myoblast determination protein) and eMHC (embryonic Myosin Heavy Chain) in CT-26 colon cancer mice.
Methods: This experimental study was performed on 20 six-week-old BALB/c mice to which CT-26 tumor was implanted. The mice were divided into four groups of resistance training, resveratrol, combination of resveratrol with resistance training, and control. The resistance training group performed six weeks of progressive resistance training. The resveratrol group received 100 mg/kg resveratrol per day, and the control and resistance training group received the solution of Methyl cellulose through gavage. In gastrocnemius muscle MyoD protein and eMHC level were measured using western blot and ELISA methods, respectively.
Results: eMHC protein in combination of resveratrol with exercise group (4.66±0.25) increased significantly compared to the exercise group (3.46±0.64) (P<0.05). Body weight of mice without tumor (21.50±1.30) in the resistance training group increased significantly compared to other groups (P<0.05). There was no significant difference in the tumor weight of mice and MyoD protein in expermental groups.
Conclusion: Regarding the increasing effect of combination of resveratrol with exercise group compared to resistance training group on eMHC, resveratrol supplementation at higher doses may be recommended along with resistance training to improve muscle regeneration.
Skeletal muscles constitute a high proportion of the cellular mass that is essential for the growth traits in cattle. Resveratrol (RSV) is a natural polyphenol compound involved in pleiotropic biological activities of muscle. Therefore, the aim of our study was to investigate the transcriptome-level effects of RSV on bovine primary myoblast to reveal differentially expressed genes (DEGs). We treated three replicates of primary myoblasts with 20 μM mother solution containing RSV, whereas three other replicates without RSV were used as control group. Then, we conducted genome-wide transcriptome analysis for the two groups. The results of expression analysis identified 3856 DEGs of which 1805 genes were up-regulated and 2051 genes were down-regulated (adjusted P < 0.05). In addition, qRT-PCR analysis of 19 selected DEGs were consistent with the expression levels observed in the transcriptome data. Gene Ontology (GO) and pathway enrichment analysis showed 72 and 66 significant GO terms and KEGG pathways, respectively (adjusted P < 0.05). The most significant GO term was actin cytoskeleton organization (GO:0030036). The top significant KEGG pathway was focal adhesion (bta04510). Predicted protein-protein interactions (PPIs) showed that CDKN1A encoding cyclindependent kinase inhibitor 1A connects several larger protein complexes. In conclusion, our results found a list of DEGs, significant GO terms and pathways, and provided an improved and expanded understanding of the impact of RSV on cattle muscle cells at the transcriptomic level. The study elucidates the potential of using the genes enriched in pathways mediating resveratrol effects as targets in genomic selection for muscle development and growth in beef cattle.
Scope:
Data on resveratrol-(trans-3,5,4'-trihydroxystilbene)-induced caloric-restriction-(CR)-mimicking effects in mice receiving a high-fat diet (HFD) are contradictory. We hypothesized that this could possibly stem from different bioactivities of resveratrol (RSV) microbial metabolites.
Methods and results:
We fed C57BL/6Rj mice an ad-libitum HFD supplemented with RSV or its metabolites, dihydroresveratrol (DHR) and lunularin (LUN) (approximately 28 mg (dihydro)stilbene/kg mouse*day) and included a 40% CR group in our study. While the CR mice showed robust changes in bodyweight and composition, hormone levels and mRNA expression, we found slight changes (more muscle, less adipose tissue) in body composition, leptin and insulin levels in RSV-supplemented mice compared to the ad-libitum controls. LUN hardly and DHR did not change the hormone levels measured. Metabolome analysis of serum showed changes in CR mice but only slight, if any, changes in RSV-, DHR- or LUN-supplemented mice compared to the controls. Evaluating the capability of RSV and its metabolites to inhibit carbohydrate-hydrolyzing enzymes in vitro, we found that RSV reduced α-glucosidase activity to a stronger extent than DHR and LUN.
Conclusion:
Decelerated carbohydrate breakdown by RSV might have contributed to the moderate impact of dietary RSV supplementation on mouse insulin sensitivity (lowered fasting and post-glucose-bolus insulin levels). This article is protected by copyright. All rights reserved.
This study investigated the effects of resveratrol and miR-22-3p on muscle fiber type conversion in mouse C2C12 myotubes. Here we showed that resveratrol significantly increased the protein level of slow myosin heavy chain (MyHC) and the activities of succinic dehydrogenase and malate dehydrogenase, as well as markedly decreased the protein level of fast MyHC and the activity of lactate dehydrogenase. Immunofluorescence staining showed that resveratrol remarkably upregulated the number of slow MyHC-positive myotubes and downregulated the number of fast MyHC-positive myotubes, suggesting that resveratrol promoted muscle fiber type conversion from fast-twitch to slow-twitch in C2C12 myotubes. We also showed that miR-22-3p had an opposite function on muscle fiber type conversion and resveratrol was able to repress the expression of miR-22-3p. Furthermore, AMP-activated protein kinase (AMPK) inhibitor Compound C and miR-22-3p mimics could attenuate and eliminate muscle fiber type conversion from fast-twitch to slow-twitch cause by resveratrol, respectively. Together, we provided the first evidence that resveratrol promotes muscle fiber type conversion from fast-twitch to slow-twitch via miR-22-3p and AMPK/SIRT1/PGC-1α pathway in C2C12 myotubes.
Skeletal muscle wasting is highly correlated with not only the reduced quality of life but also higher morbidity and mortality. Although increasing patients are suffering from various kinds of muscle atrophy and weakness, there’s still no effective therapy available, and skeletal muscle is considered as an under-medicated organ. Food provided not only essential macronutrients but functional substances involved in modulation of physiological systems of our body. Natural constituents from commonly-consumed dietary plants, either extracts or compounds, have attracted more and more attention to be developed as agents for preventing and treating muscle wasting due to their safety and effectiveness, as well as structural diversity. This review provides an overview of the mechanistic aspects of muscle wasting, and summarizes the extracts and compounds from food sources as potential therapies against muscle wasting.
Phytochemicals have multifunctional bioactivities, such as anti-inflammatory and anti-obesity activities. However, there are still relatively few studies that have investigated the possible anti-fatigue of functional phytochemical compounds (such as resveratrol, triterpenoids, and catechins) in animal models. Therefore, the aim of this study was to develop of in vitro models as tools for cellular and molecular studies of anti-fatigue. C2C12 myotubes were used for evaluating in vitro the anti-fatigue of functional compounds. The results indicated that treatment with some functional compounds (including resveratrol, quercetin, epigallocatechin gallate, and epicatechin) significantly increased the expression of mitochondrial DNA genes (including cytochrome b and cytochrome c oxidase subunit II) and the activities of mitochondrial enzymes (including citrate synthase and cytochrome c oxidase) in C2C12 myotubes. These results demonstrated that resveratrol, quercetin, epigallocatechin gallate, and epicatechin have great potential as novel mitochondrial-activating agents that may play an important role in anti-fatigue.
Aims:
Here, we decided to investigate whether HK activity and its interaction with mitochondria could be a target of 4-PBA action.
Main methods:
L6 myotubes were treated with 1mM 4-PBA for 24, 48 or 72h. We evaluated HK activity, glucose and oxygen consumption, gene and protein expression.
Key findings:
We found that L6 myotubes treated with 4-PBA presented more HK activity in the particulate fraction, increased glucose consumption and augmented Glut4, Hk2 and Vdac1 mRNA expression. Moreover, 4-PBA prevented the deleterious effect of antimycin-A on HK particulate activity.
Significance:
Together, these results suggest a new role of 4-PBA in glucose metabolism that includes HK as a potential target of beneficial effect of 4-PBA.
Epigenetics is an area of research that has recently gained much attention from scientists. Epigenetic processes can induce changes within an organism without altering its genetic makeup. More interestingly, epigenetic mechanisms have the strong ability to modulate gene expression without directly altering the sequences of DNA bases. Dietary compounds consist of several bioactive constituents, which actively regulate different molecular targets involved in tumorigenesis. Keeping these facts in view, we provide evidence that these dietary components (e.g. resveratrol (RES), curcumin, genistein, polyphenols and sulforaphane) might interact with various epigenetic targets in cancer therapeutics. These bioactive compounds can modulate normal DNA methylation and histone acetylation patterns, which are essential for the activation of cancer fighting genes. Compounds, such as the ones listed above, induce epigenetic changes associated with the expression of tumor suppressor genes, such as p53, and inhibition of tumor promoting genes such as telomerase reverse transcriptase during tumor progression. Therefore, in this chapter, we present considerable evidence that bioactive compounds and their epigenetic targets are linked with cancer therapeutics, which may open the door to novel drug discovery and development. Remarkable improvements in our understanding of basic epigenetic mechanisms coupled with the rapid progress in the development of powerful new technologies hold great promise for the advancement of cancer treatment.
Un concept qui a fait son apparition ces dernières années est l’utilisation de micronutriments comme agents thérapeutiques connus pour leurs propriétés anti-inflammatoires, antioxydantes, cardio-protectrices ou anticancéreuses. Dans notre travail, nous nous sommes intéressés à l’étude des mécanismes d’actions in vivo et in vitro du resvératrol et de deux formulations naturelles, l’une impliquée dans les effets antioxydants (produit « X ») et l’autre (produit « Y ») destinée à corriger les désordres de nature lipidique de sujets présentant un syndrome métabolique. Nous avons caractérisé les effets pro-différenciateurs d’un polyphénol naturel, le resvératrol, sur cellules squelettiques musculaires C2C12 en culture. L’étude a montré que ce polyphénol permet d’induire l’expression de facteurs de la différenciation musculaire (Myogénine, Scrp3) et d’augmenter le taux de la myosine, protéine impliquée dans la contraction musculaire. Enfin, le resvératrol permet de moduler l’expression de microRNA, comme par exemple miR-133b impliqué dans les mécanismes de différenciation musculaire. Les résultats sur le stress oxydant montrent que le produit « X » est capable, à la fois, de moduler de façon modérée l’expression des gènes des défenses anti-radicalaires au niveau du muscle gastrocnémiens (Sirt1, Sod1, Ucp2, Nrf1) mais aussi au niveau des cellules musculaires squelettiques murines C2C12 (Prdx1, Nrf2, Pgc1α) en culture. Les résultats au niveau des gastrocnémiens ont montré également une élévation du potentiel global des défenses anti-radicalaires par dosage KRL. Une diminution du taux de ROS intracellulaires a été observée avec sonde DHE dans les cellules C2C12 en culture. L’étude des mécanismes d’action de « Y » a montré qu’il est capable de moduler de façon modérée l’expression des gènes du métabolisme énergétique (Gs1, Srebp1c, Fabp3, Vlcad, Pparβ) dans les cellules C2C12. Nos résultats montrent également que « Y » induit un effet anti-inflammatoire par la diminution du nombre de lymphocytes sous un régime riche en calorie. Toutefois, « Y » n’est pas un agoniste de PPARα et de PPARγ dans nos conditions. L’ensemble des résultats obtenus au cours de ce travail confirme que les micronutriments sont capables d’induire des effets au niveau génique quoique de façon modérée. Des études à long terme sont nécessaires pour comprendre et confirmer réellement leur impact sur l’organisme
Resveratrol is one of the best known polyphenol. While its effect on endothelial blood vessel cells, cancer cells, inflammatory processes and neurodegenerative events is well documented, only little is established on the metabolic implication of this phytophenol, particularly on skeletal muscle cells. Here, we report the effect of resveratrol on mouse skeletal muscle cells (C2C12) by measurement of cell proliferation, expression of metabolic genes and their transcription factor dependency, and modulation of non-coding microRNA expression. Resveratrol slightly decreases cell proliferation, while it up regulates PGC-1 a PPAR transcription factor co-activator involved in the control of metabolic genes. It also modulates microRNAs which are expressed in skeletal muscle cells and are involved in differentiation or contraction-relaxation. Results put forward new regulatory properties of resveratrol on skeletal muscles and highlight new potential applications of this molecule in muscle performances.
Resveratrol, a polyphenol in red wine, has been reported as a calorie restriction mimetic with potential antiaging and antidiabetogenic properties. It is widely consumed as a nutritional supplement, but its mechanism of action remains a mystery. Here, we report that the metabolic effects of resveratrol result from competitive inhibition of cAMP-degrading phosphodiesterases, leading to elevated cAMP levels. The resulting activation of Epac1, a cAMP effector protein, increases intracellular Ca(2+) levels and activates the CamKKβ-AMPK pathway via phospholipase C and the ryanodine receptor Ca(2+)-release channel. As a consequence, resveratrol increases NAD(+) and the activity of Sirt1. Inhibiting PDE4 with rolipram reproduces all of the metabolic benefits of resveratrol, including prevention of diet-induced obesity and an increase in mitochondrial function, physical stamina, and glucose tolerance in mice. Therefore, administration of PDE4 inhibitors may also protect against and ameliorate the symptoms of metabolic diseases associated with aging.
Piceatannol, a natural stilbene, is an analog and a metabolite of resveratrol. Despite a well documented health benefit of resveratrol in intervention of the development of obesity, the role of piceatannol in the development of adipose tissue and related diseases is unknown. Here, we sought to determine the function of piceatannol in adipogenesis and elucidate the underlying mechanism. We show that piceatannol inhibits adipogenesis of 3T3-L1 preadipocytes in a dose-dependent manner at noncytotoxic concentrations. This anti-adipogenic property of piceatannol was largely limited to the early event of adipogenesis. In the early phase of adipogenesis, piceatannol-treated preadipocytes displayed a delayed cell cycle entry into G(2)/M phase at 24 h after initiation of adipogenesis. Furthermore, the piceatannol-suppressed mitotic clonal expansion was accompanied by reduced activation of the insulin-signaling pathway. Piceatannol dose-dependently inhibited differentiation mixture-induced phosphorylation of insulin receptor (IR)/insulin receptor substrate-1 (IRS-1)/Akt pathway in the early phase of adipogenesis. Moreover, we showed that piceatannol is an inhibitor of IR kinase activity and phosphatidylinositol 3-kinase (PI3K). Our kinetics study of IR further identified a K(m) value for ATP of 57.8 μm and a K(i) value for piceatannol of 28.9 μm. We also showed that piceatannol directly binds to IR and inhibits IR kinase activity in a mixed noncompetitive manner to ATP, through which piceatannol appears to inhibit adipogenesis. Taken together, our study reveals an anti-adipogenic function of piceatannol and highlights IR and its downstream insulin signaling as novel targets for piceatannol in the early phase of adipogenesis.
Background:
Polyphenols are the most abundant antioxidants in the human diet and are widespread constituents of fruits and beverages, such as tea, coffee or wine. Epidemiological, clinical and animal studies support a role of polyphenols in the prevention of various diseases, such as cardiovascular diseases, cancers or neurodegenerative diseases. Recent findings suggest that polyphenols could interact with cellular signaling cascades regulating the activity of transcription factors and consequently affecting the expression of genes. However, the impact of polyphenol on the expression of microRNA, small non-coding RNAs, has not yet been studied. The aim of this study was to investigate the impact of dietary supplementation with polyphenols at nutritional doses on miRNA expression in the livers of apolipoprotein E-deficient mice (apoE⁻/⁻) jointly with mRNA expression profiling.
Methodology/principal findings:
Using microarrays, we measured the global miRNA expression in the livers of wild-type (C57B6/J) mice or apoE⁻/⁻ mice fed diets supplemented with one of nine different polyphenols or a control diet. This analysis revealed that knock-out of the apoE gene induced significant modulation in the expression of miRNA. Moreover, changes in miRNA expression were observed after polyphenol supplementation, and five miRNAs (mmu-miR-291b-5p, mmu-miR-296-5p, mmu-miR-30c-1*, mmu-miR-467b* and mmu-miR-374*) were identified as being commonly modulated by these polyphenols. We also observed that these polyphenols counteracted the modulation of miRNA expression induced by apoE mutation. Pathway analyses on these five miRNA-target genes revealed common pathways, some of which were also identified from a pathway analysis on mRNA profiles.
Conclusion:
This in vivo study demonstrated for the first time that polyphenols at nutritional doses modulate the expression of miRNA in the liver. Even if structurally different, all polyphenols induced a similar miRNA expression profile. Common pathways were identified from both miRNA-target and mRNA analysis, revealing cellular functions that could be regulated by polyphenols at both the miRNA and mRNA level.
Sirtuin 1 also known as NAD-dependent deacetylase sirtuin 1, is a protein that in humans is encoded by the Sirt1 gene. Sirt1 is an enzyme that deacetylates proteins that contribute to cellular regulation and is a key regulator of cell defenses and survival in response to stress. Deletion of Sirt1 abolishes the increase in lifespan induced by calorie restriction or sublethal cytokine stress, indicating that Sirt1 promotes longevity and survival. We have demonstrated that administration of a sublethal dose of tumour necrosis factor-α (TNF-α; 1.25 ng ml(-1)) inhibits myotube formation, and co-incubation with insulin-like growth factor I (IGF-I; 1.5 ng ml(-1)) facilitates C2 myoblast death rather than rescuing differentiation. A higher dose of TNF-α (10 ng ml(-1)) resulted in significant apoptosis, which was rescued by IGF-I (1.5 ng ml(-1); 50% rescue; P < 0.05). We aimed to investigate the role of Sirt1 in the conflicting roles of IGF-I. Quantitative real-time PCR revealed that Sirt1 expression was elevated in myoblasts following incubation of 10 ng ml(-1) TNF-α or 1.25 ng ml(-1) TNF-α plus IGF-I (fivefold and 7.2-fold increases versus control, respectively; P < 0.05). A dose of 10 ng ml(-1) TNF-α induced ∼21 ± 0.7% apoptosis, which was reduced (∼50%; P < 0.05) when administered with IGF-I. Likewise, Sirt1 expression was elevated following 10 ng ml(-1) TNF-α administration, but was reduced (∼30%; P < 0.05) in the presence of IGF-I. C2C12 myoblasts, a subclone of the C2 cell line produced for their differentiation potential and used to examine intrinsic ageing, unlike C2 cells, do not die in the presence of TNF-α and do not upregulate Sirt1. As conditions that induced the greatest myoblast stress/damage resulted in elevated Sirt1 expression, we investigated the effects of Sirt1 gene silencing. Treatment with 10 ng ml(-1) TNF-α or co-incubation with 1.25 ng ml(-1) TNF-α and 1.5 ng ml(-1) IGF-I resulted in apoptosis (20.33 ± 2.08 and 19 ± 2.65%, respectively), which was increased when myoblasts were pretreated with Sirt1 small interfering RNA (31 ± 2.65 and 27.33 ± 2.52%, respectively; P < 0.05) and was reduced (14.33 ± 3.05%, P < 0.05 and 12.78 ± 4.52%, P = 0.054) by resveratrol, which also significantly rescued the block on differentiation. In conclusion, Sirt1 expression increases in conditons of stress, potentially serving to reduce or dampen myoblast death.
Resveratrol is a natural compound that affects energy metabolism and mitochondrial function and serves as a calorie restriction mimetic, at least in animal models of obesity. Here, we treated 11 healthy, obese men with placebo and 150 mg/day resveratrol (resVida) in a randomized double-blind crossover study for 30 days. Resveratrol significantly reduced sleeping and resting metabolic rate. In muscle, resveratrol activated AMPK, increased SIRT1 and PGC-1α protein levels, increased citrate synthase activity without change in mitochondrial content, and improved muscle mitochondrial respiration on a fatty acid-derived substrate. Furthermore, resveratrol elevated intramyocellular lipid levels and decreased intrahepatic lipid content, circulating glucose, triglycerides, alanine-aminotransferase, and inflammation markers. Systolic blood pressure dropped and HOMA index improved after resveratrol. In the postprandial state, adipose tissue lipolysis and plasma fatty acid and glycerol decreased. In conclusion, we demonstrate that 30 days of resveratrol supplementation induces metabolic changes in obese humans, mimicking the effects of calorie restriction.
Resveratrol (Res) is a natural polyphenolic compound with anti-inflammatory and antioxidant properties. Also, Res can inhibit lipogenesis and adipocyte differentiation. However, the underlying mechanisms of Res's functions remain largely unknown. AMP-activated protein kinase (AMPK) is a key player in adipocyte differentiation. Therefore, the purpose of our study was to determine the role played by AMPK in the Res-mediated regulation of adipocyte differentiation. Incubation of 3T3-L1 cells with Res confirmed that Res inhibited adipocyte differentiation. The phosphorylation of AMPKα was increased by Res in a dose-dependent manner, while total AMPKα levels were unchanged, and peroxisome proliferator-activated receptor γ (PPARγ), CCAAT-enhancer-binding protein α (C/EBPα), and sterol regulatory element-binding protein 1c (SREBP-1c) levels were decreased. Interestingly, pretreatment with AMPKα siRNA and Res promoted adipocyte differentiation, while the decrease of p-AMPKα increased PPARγ, C/EBPα, and SREBP-1c protein expression. Our study shows that Res is capable of inhibiting lipogenesis and differentiation of 3T3-L1 adipocytes via activation of AMPK, suggesting its potential therapeutic application in the treatment or prevention of obesity.
Inflammatory cell recruitment and intimal neovascularization contribute to atherosclerotic plaque destabilization. The anti-inflammatory red wine polyphenol, resveratrol, has been implicated in cardiovascular protection. In this study, we investigated the effects of resveratrol on endothelial and monocytic cell migration.
Human umbilical vein endothelial cell (EC) migration was assessed in a modified barrier assay. Chemotaxis of THP-1 monocytic cells towards monocyte chemoattractant protein (MCP)-1 was determined using a Boyden chamber. Erk phosphorylation downstream of MCP-1 receptor and activation of myosin phosphatase targeting subunit 1 (pMYPT1) downstream of Rho kinase were determined by Western blotting.
In resveratrol-treated cells, progressive shape elongation was observed, evident after 6h of treatment. Treatment with resveratrol (1-20 µmol/L) dose-dependently inhibited EC migration. This effect of resveratrol was independent of nuclear factor (NF)-kappaB and sirtuin 1, but was abrogated in the presence of Rho kinase inhibitors. Moreover, resveratrol induced pMYPT1 activation, indicating a novel mechanism of resveratrol activity in EC. In monocytic cells, treatment with resveratrol significantly inhibited chemotaxis towards MCP-1 already at 1 µmol/L. At a resveratrol concentration of 10 µmol/L, chemotaxis was reduced nearly to the negative control (unstimulated with MCP-1) levels. This effect was independent of NF-kappaB and RhoA signaling. In resveratrol treated monocytic cells, MCP-1-induced Erk phosphorylation downstream of CCR2 receptor was dose-dependently inhibited, as observed by Western blot analysis.
Resveratrol dose-dependently inhibited endothelial cell migration and MCP-1-induced monocytic cell chemotaxis. This activity may contribute to the cardioprotective effects of resveratrol by inhibition of intimal neovascularization and monocyte recruitment into the artery wall.
MicroRNAs (miRNAs or miRs) participate in the regulation of several biological processes, including cell differentiation. Recently, miR-34a has been implicated in the differentiation of monocyte-derived dendritic cells, human erythroleukemia cells, and mouse embryonic stem cells. In addition, members of the miR-34 family have been identified as direct p53 targets. However, the function of miR-34a in the control of the differentiation program of specific neural cell types remains largely unknown. Here, we investigated the role of miR-34a in regulating mouse neural stem (NS) cell differentiation.
miR-34a overexpression increased postmitotic neurons and neurite elongation of mouse NS cells, whereas anti-miR-34a had the opposite effect. SIRT1 was identified as a target of miR-34a, which may mediate the effect of miR-34a on neurite elongation. In addition, acetylation of p53 (Lys 379) and p53-DNA binding activity were increased and cell death unchanged after miR-34a overexpression, thus reinforcing the role of p53 during neural differentiation. Interestingly, in conditions where SIRT1 was activated by pharmacologic treatment with resveratrol, miR-34a promoted astrocytic differentiation, through a SIRT1-independent mechanism.
Our results provide new insight into the molecular mechanisms by which miR-34a modulates neural differentiation, suggesting that miR-34a is required for proper neuronal differentiation, in part, by targeting SIRT1 and modulating p53 activity.
MicroRNAs are short noncoding RNAs that regulate the expression of many target genes posttranscriptionally and are thus implicated in a wide array of cellular and developmental processes. The expression of miR-155 or miR-21 is upregulated during the course of the inflammatory response, but these microRNAs are also considered oncogenes due to their upregulation of expression in several types of tumors. Furthermore, it is now well established that inflammation is associated with the induction or the aggravation of nearly 25% of cancers. Therefore, the above microRNAs are thought to link inflammation and cancer. Recently, resveratrol (trans-3,4',5-trihydroxystilbene), a natural polyphenol with antioxidant, anti-inflammatory, and anticancer properties, currently at the stage of preclinical studies for human cancer prevention, has been shown to induce the expression of miR-663, a tumor-suppressor and anti-inflammatory microRNA, while downregulating miR-155 and miR-21. In this paper we will discuss how the use of resveratrol in therapeutics may benefit from the preanalyses on the status of expression of miR-155 or miR-21 as well as of TGFβ1. In addition, we will discuss how resveratrol activity might possibly be enhanced by simultaneously manipulating the levels of its key target microRNAs, such as miR-663.
Thyroid carcinogenesis is accompanied by loss of thyroid-specific functions and refractory to radioiodine and thyroid stimulating hormone (TSH) suppression therapy. Redifferentiating agents have been shown to inhibit tumor growth and improve the response to conventional therapy. Polyphenol phytochemicals (PPs) in fruits and vegetables have been reported to inhibit cancer initiation, promotion, progression and induce redifferentiation in selected types. In this study we examined PPs induce redifferentiation in thyroid cancer cell lines. We investigated the effects of genistein, resveratrol, quercetin, kaempferol, and resorcinol on the F9 embryonal carcinoma cell differentiation model. The thyroid cancer cell lines, TPC-1, FTC-133, NPA, FRO, and ARO, displayed growth inhibition in response to genistein, resveratrol, quercetin. We further demonstrated that genistein decreased the dedifferention marker CD97 in NPA cells and resveratrol decreased CD97 in FTC-133, NPA, FRO cells and quercetin decreased CD97 in all cell lines. We observed increased expression of differentiation marker NIS in FTC-133 cells in response to genistein, and resveratrol but no change in NPA, FRO, ARO cells. Quercetin increased or induced NIS in FTC-133, NPA, FRO cells. These findings suggest that PPs may provide a useful therapeutic intervention in thyroid cancer redifferentiation therapy.
Muscular dystrophies are inherited myogenic disorders accompanied by progressive skeletal muscle weakness and degeneration. We previously showed that resveratrol (3,5,4'-trihydroxy-trans-stilbene), an antioxidant and activator of the NAD(+)-dependent protein deacetylase SIRT1, delays the progression of heart failure and prolongs the lifespan of δ-sarcoglycan-deficient hamsters. Because a defect of dystroglycan complex causes muscular dystrophies, and δ-sarcoglycan is a component of this complex, we hypothesized that resveratrol might be a new therapeutic tool for muscular dystrophies. Here, we examined resveratrol's effect in mdx mice, an animal model of Duchenne muscular dystrophy. mdx mice that received resveratrol in the diet for 32 weeks (4 g/kg diet) showed significantly less muscle mass loss and nonmuscle interstitial tissue in the biceps femoris compared with mdx mice fed a control diet. In the muscles of these mice, resveratrol significantly decreased oxidative damage shown by the immunostaining of nitrotyrosine and 8-hydroxy-2'-deoxyguanosine and suppressed the up-regulation of NADPH oxidase subunits Nox4, Duox1, and p47(phox). Resveratrol also reduced the number of α-smooth muscle actin (α-SMA)(+) myofibroblast cells and endomysial fibrosis in the biceps femoris, although the infiltration of CD45(+) inflammatory cells and increase in transforming growth factor-β1 (TGF-β1) were still observed. In C2C12 myoblast cells, resveratrol pretreatment suppressed the TGF-β1-induced increase in reactive oxygen species, fibronectin production, and expression of α-SMA, and SIRT1 knockdown blocked these inhibitory effects. SIRT1 small interfering RNA also increased the expression of Nox4, p47(phox), and α-SMA in C2C12 cells. Taken together, these findings indicate that SIRT1 activation may be a useful strategy for treating muscular dystrophies.
trans-Resveratrol has been proposed to prevent tumor growth and to sensitize cancer cells to anticancer agents. Polyphenol entry into the cells has remained poorly understood. Here, we show that [(3)H]-resveratrol enters colon cancer cells (SW480, SW620, HT29) and leukemia U937 cells through a monensin (5-20 μmol/L) -sensitive process that suggests clathrin-independent endocytosis. Uptake of the molecule can be prevented by methyl-β-cyclodextrin (2-12 mg/mL), nystatin (12 ng/mL), and filipin (1 μg/mL), which all disrupt plasma membrane lipid rafts. Accordingly, radiolabeled resveratrol accumulates in sphingomyelin- and cholesterol-enriched cell fractions. Interestingly, extracellular signal-regulated kinases (ERK), c-Jun NH(2)-terminal kinases (JNK), and Akt also accumulate in lipid rafts on resveratrol exposure (IC(50) at 48 h ≈ 30 μmol/L in SW480 and U937 cells). In these rafts also, resveratrol promotes the recruitment, by the integrin α(V)β(3) (revealed by coimmunoprecipitation with an anti-integrin α(V)β(3) antibody), of signaling molecules that include the FAK (focal adhesion kinase), Fyn, Grb2, Ras, and SOS proteins. Resveratrol-induced activation of downstream signaling pathways and caspase-dependent apoptosis is prevented by endocytosis inhibitors, lipid raft-disrupting molecules, and the integrin antagonist peptide arginine-glycine-aspartate (500 nmol/L). Altogether, these data show the role played by lipid rafts in resveratrol endocytosis and activation of downstream pathways leading to cell death.
Carnitine palmitoyl transferase 2 (CPT2) and very-long-chain Acyl-CoA dehydrogenase (VLCAD) deficiencies are among the most common inborn mitochondrial fatty acid β-oxidation (FAO) disorders. Despite advances in their clinical and molecular characterizations, few therapeutic approaches exist for these diseases. Resveratrol (RSV) is a natural polyphenol extensively studied for its potential health benefits. Indeed, it is presently thought that RSV could delay the onset of some cancers, and have protective effects against common aging disorders such as type II diabetes, cardiovascular or neurodegenerative diseases. Here, we show that exposure to RSV induces a dose- and time-dependant increase in FAO flux in human fibroblasts, and can restore normal FAO capacities in a panel of patients' fibroblasts with the mild forms (harboring various genotypes) of CPT2 or VLCAD deficiency. The correction of FAO flux correlated with a marked increase in mutant CPT2 or VLCAD protein level, in cells treated by RSV. Inhibition of sirtuin 1 (SIRT1) by Sirtinol and the use of peroxisome proliferator-activated receptor gamma co-activator-1-alpha (PGC-1α) small interfering RNAs demonstrate that the RSV-induced stimulation of FAO requires the presence of PGC-1α and SIRT1. These results show, for the first time, that RSV markedly induces mitochondrial FAO capacities in human fibroblasts, and provides the initial proof-of-concept that RSV might be efficient for correction of inherited FAO disorders.
The authors investigated the role of resveratrol (RV), a natural poliphenolic molecule with several biological activities, in transforming growth factor-β (TGF-β)-induced proliferation and differentiation of ex vivo human pulmonary fibroblasts into myofibroblasts. The effects of RV treatment were evaluated by analyzing TGF-β-induced α-smooth muscle actin (α-SMA) expression and collagen production, as well as cell proliferation of both normal and idiopathic pulmonary fibrosis (IPF) lung fibroblasts. Results demonstrate that RV inhibits TGF-β-induced cell proliferation of both normal and pathological lung fibroblasts, attenuates α-SMA expression at both the mRNA and protein levels, and also inhibits intracellular collagen deposition. In order to understand the molecular mechanisms, the authors also investigated the effects of RV treatment on signaling pathways involved in TGF-β-induced fibrosis. The authors show that RV inhibited TGF-β-induced phosphorylation of both extracellular signal-regulated kinases (ERK1/2) and the serine/threonine kinase, Akt. Moreover, RV treatment blocked the TGF-β-induced decrease in phosphatase and tensin homolog (PTEN) expression levels.
Resveratrol is a phytochemical with chemopreventive activity in preclinical rodent models of colorectal carcinogenesis. Antiproliferation is one of the many chemopreventive modes of action it has been shown to engage in. Concentrations of resveratrol, which can be achieved in human tissues after p.o. administration, have not yet been defined. The purpose of this study was to measure concentrations of resveratrol and its metabolites in the colorectal tissue of humans who ingested resveratrol. Twenty patients with histologically confirmed colorectal cancer consumed eight daily doses of resveratrol at 0.5 or 1.0 g before surgical resection. Resveratrol was found to be well tolerated. Normal and malignant biopsy tissue samples were obtained before dosing. Parent compound plus its metabolites resveratrol-3-O-glucuronide, resveratrol-4'-O-glucuronide, resveratrol-3-O-sulfate, resveratrol-4'-O-sulfate, resveratrol sulfate glucuronide, and resveratrol disulfate were identified by high-performance liquid chromatography (HPLC) with UV or mass spectrometric detection in colorectal resection tissue. Quantitation was achieved by HPLC/UV. Cell proliferation, as reflected by Ki-67 staining, was compared in preintervention and postintervention tissue samples. Resveratrol and resveratrol-3-O-glucuronide were recovered from tissues at maximal mean concentrations of 674 and 86.0 nmol/g, respectively. Levels of resveratrol and its metabolites were consistently higher in tissues originating in the right side of the colon compared with the left. Consumption of resveratrol reduced tumor cell proliferation by 5% (P = 0.05). The results suggest that daily p.o. doses of resveratrol at 0.5 or 1.0 g produce levels in the human gastrointestinal tract of an order of magnitude sufficient to elicit anticarcinogenic effects. Resveratrol merits further clinical evaluation as a potential colorectal cancer chemopreventive agent.
An inflammatory component is present in the microenvironment of most neoplastic tissues, including those not causally related to an obvious inflammatory process. Several microRNAs, and especially miR-155, play an essential role in both the innate and adaptative immune response. Resveratrol (trans-3,4',5-trihydroxystilbene) is a natural antioxidant with anti-inflammatory properties that is currently at the stage of preclinical studies for human cancer prevention. Here, we establish that, in human THP-1 monocytic cells as well as in human blood monocytes, resveratrol upregulates miR-663, a microRNA potentially targeting multiple genes implicated in the immune response. In THP-1 cells, miR-663 decreases endogenous activator protein-1 (AP-1) activity and impairs its upregulation by lipopolysaccharides (LPS), at least in part by directly targeting JunB and JunD transcripts. We further establish that the downregulation of AP-1 activity by resveratrol is miR-663 dependent and that the effects of resveratrol on both AP-1 activity and JunB levels are dose dependent. Finally, we show that resveratrol impairs the upregulation of miR-155 by LPS in a miR-663-dependent manner. Given the role of miR-155 in the innate immune response and the fact that it is upregulated in many cancers, our results suggest that manipulating miR-663 levels may help to optimize the use of resveratrol as both an anti-inflammatory and anticancer agent against malignancies associated with high levels of miR-155.
Several global transcriptomic and proteomic approaches have been applied in order to obtain new molecular insights on skeletal myogenesis, but none has generated any specific data on glycogenome expression, and thus on the role of glycan structures in this process, despite the involvement of glycoconjugates in various biological events including differentiation and development. In the present study, a quantitative real-time RT-PCR technology was used to profile the dynamic expression of 375 glycogenes during the differentiation of C2C12 myoblasts into myotubes.
Of the 276 genes expressed, 95 exhibited altered mRNA expression when C2C12 cells differentiated and 37 displayed more than 4-fold up- or down-regulations. Principal Component Analysis and Hierarchical Component Analysis of the expression dynamics identified three groups of coordinately and sequentially regulated genes. The first group included 12 down-regulated genes, the second group four genes with an expression peak at 24 h of differentiation, and the last 21 up-regulated genes. These genes mainly encode cell adhesion molecules and key enzymes involved in the biosynthesis of glycosaminoglycans and glycolipids (neolactoseries, lactoseries and ganglioseries), providing a clearer indication of how the plasma membrane and extracellular matrix may be modified prior to cell fusion. In particular, an increase in the quantity of ganglioside GM3 at the cell surface of myoblasts is suggestive of its potential role during the initial steps of myogenic differentiation.
For the first time, these results provide a broad description of the expression dynamics of glycogenes during C2C12 differentiation. Among the 37 highly deregulated glycogenes, 29 had never been associated with myogenesis. Their biological functions suggest new roles for glycans in skeletal myogenesis.
Resveratrol is a natural polyphenol antioxidant that has been shown to facilitate osteogenic differentiation. A recent breakthrough has demonstrated that ectopic expression of four genes is sufficient to reprogram murine and human fibroblasts into induced pluripotent stem (iPS) cells. However, the roles of resveratrol in the differentiation and cytoprotection of iPS cells have never been studied. In this study, we showed that, in addition to cardiac cells, neuron-like cells, and adipocytes, mouse iPS cells could differentiate into osteocyte-like cells. Using atomic force microscopy that provided nanoscale resolution, we monitored mechanical properties of living iPS cells during osteogenic differentiation. The intensity of mineralization and stiffness in differentiating iPS significantly increased after 14 days of osteogenic induction. Furthermore, resveratrol was found to facilitate osteogenic differentiation in both iPS and embryonic stem cells, as shown by increased mineralization, up-regulation of osteogenic markers, and decreased elastic modulus. Dexamethasone-induced apoptosis in iPS cell-derived osteocyte-like cells was effectively prevented by pretreatment with resveratrol. Furthermore, resveratrol significantly increased manganese superoxide dismutase expression and intracellular glutathione level, thereby efficiently decreasing dexamethasone-induced reactive oxygen species (ROS) production and cytotoxicity. Transplantation experiments using iPS cell-derived osteocyte-like cells further demonstrated that oral intake of resveratrol could up-regulate osteopontin expression and inhibit teratoma formation in vivo. In sum, resveratrol can facilitate differentiation of iPS cells into osteocyte-like cells, protect these iPS cell-derived osteocyte-like cells from glucocorticoid-induced oxidative damage, and decrease tumorigenicity of iPS cells. These findings implicate roles of resveratrol and iPS cells in the stem cell therapy of orthopedic diseases.
In diverse organisms, calorie restriction slows the pace of ageing and increases maximum lifespan. In the budding yeast Saccharomyces cerevisiae, calorie restriction extends lifespan by increasing the activity of Sir2 (ref. 1), a member of the conserved sirtuin family of NAD(+)-dependent protein deacetylases. Included in this family are SIR-2.1, a Caenorhabditis elegans enzyme that regulates lifespan, and SIRT1, a human deacetylase that promotes cell survival by negatively regulating the p53 tumour suppressor. Here we report the discovery of three classes of small molecules that activate sirtuins. We show that the potent activator resveratrol, a polyphenol found in red wine, lowers the Michaelis constant of SIRT1 for both the acetylated substrate and NAD(+), and increases cell survival by stimulating SIRT1-dependent deacetylation of p53. In yeast, resveratrol mimics calorie restriction by stimulating Sir2, increasing DNA stability and extending lifespan by 70%. We discuss possible evolutionary origins of this phenomenon and suggest new lines of research into the therapeutic use of sirtuin activators.
Understanding the molecular mechanisms that regulate cellular proliferation and differentiation is a central theme of developmental biology. MicroRNAs (miRNAs) are a class of regulatory RNAs of approximately 22 nucleotides that post-transcriptionally regulate gene expression. Increasing evidence points to the potential role of miRNAs in various biological processes. Here we show that miRNA-1 (miR-1) and miRNA-133 (miR-133), which are clustered on the same chromosomal loci, are transcribed together in a tissue-specific manner during development. miR-1 and miR-133 have distinct roles in modulating skeletal muscle proliferation and differentiation in cultured myoblasts in vitro and in Xenopus laevis embryos in vivo. miR-1 promotes myogenesis by targeting histone deacetylase 4 (HDAC4), a transcriptional repressor of muscle gene expression. By contrast, miR-133 enhances myoblast proliferation by repressing serum response factor (SRF). Our results show that two mature miRNAs, derived from the same miRNA polycistron and transcribed together, can carry out distinct biological functions. Together, our studies suggest a molecular mechanism in which miRNAs participate in transcriptional circuits that control skeletal muscle gene expression and embryonic development.
Sir2 regulates lifespan in model organisms, which has stimulated interest in understanding human Sir2 homolog functions. The human Sir2 gene family comprises seven members (SIRT1-SIRT7). SIRT1, the human ortholog of the yeast Sir2 by closest sequence similarity, is a nicotinamide adenine dinucleotide (NAD(+))-dependent deacetylase with enzymatic properties indistinguishable from the yeast enzyme. We studied the involvement of SIRT1 in normal human keratinocyte physiology by a transcriptional microarray analysis of primary keratinocytes either overexpressing or underexpressing SIRT1. Using a systems biology analytical approach, we predicted that SIRT1 induces keratinocyte differentiation through a pathway integral to or overlapping with that of calcium-induced differentiation. We experimentally assayed this prediction and found that the SIRT1 inhibitor nicotinamide inhibited expression of keratinocyte differentiation markers, whereas a SIRT1 activator, resveratrol, enhanced expression of keratinocyte differentiation markers. Similar results were obtained in keratinocytes manipulated to overexpress or underexpress SIRT1, and modulating SIRT1 significantly affected keratinocyte proliferation rates. We conclude that SIRT1 functions in normal human keratinocytes to inhibit proliferation and to promote differentiation.
Real-time reverse transcription followed by polymerase chain reaction (RT–PCR) is the most suitable method for the detection
and quantification of mRNA. It offers high sensitivity, good reproducibility and a wide quantification range. Today, relative
expression is increasingly used, where the expression of a target gene is standardised by a non-regulated reference gene.
Several mathematical algorithms have been developed to compute an expression ratio, based on real-time PCR efficiency and
the crossing point deviation of an unknown sample versus a control. But all published equations and available models for the
calculation of relative expression ratio allow only for the determination of a single transcription difference between one
control and one sample. Therefore a new software tool was established, named REST© (relative expression software tool), which
compares two groups, with up to 16 data points in a sample and 16 in a control group, for reference and up to four target
genes. The mathematical model used is based on the PCR efficiencies and the mean crossing point deviation between the sample
and control group. Subsequently, the expression ratio results of the four investigated transcripts are tested for significance
by a randomisation test. Herein, development and application of REST© is explained and the usefulness of relative expression
in real-time PCR using REST© is discussed. The latest software version of REST© and examples for the correct use can be downloaded
at http://www.wzw.tum.de/gene-quantification/.
Cited By (since 1996):11, Export Date: 27 November 2013, Source: Scopus
Grapes produce large amounts of polyphenols. Many of them accumulate in the skin, pulp, and seeds and are consequently found in wine. The health benefits of a moderate consumption of wine have been attributed at least in part to grape's polyphenols. Among them, resveratrol (3,5,4'-trihydroxystilbene) is a phytoalexin that stimulates plant cell defenses against infections and also plays protective roles in humans, where it delays cardiovascular alterations and exerts anticancer and anti-inflammatory effects. Despite numerous studies, the molecular mechanisms of resveratrol action are only partially understood. Given its pleiotropic effects, it was previously suggested that resveratrol protective properties may arise from its modulation of the expression of microRNAs. Therefore, this review will focus on the effects of resveratrol on microRNA populations in humans and human cell lines, especially emphasizing the microRNAs that have been implicated in resveratrol effects on inflammation, cancer, metabolism, and muscle differentiation.
Resveratrol induces mitochondrial biogenesis and protects against metabolic decline, but whether SIRT1 mediates these benefits is the subject of debate. To circumvent the developmental defects of germline SIRT1 knockouts, we have developed an inducible system that permits whole-body deletion of SIRT1 in adult mice. Mice treated with a moderate dose of resveratrol showed increased mitochondrial biogenesis and function, AMPK activation, and increased NAD(+) levels in skeletal muscle, whereas SIRT1 knockouts displayed none of these benefits. A mouse overexpressing SIRT1 mimicked these effects. A high dose of resveratrol activated AMPK in a SIRT1-independent manner, demonstrating that resveratrol dosage is a critical factor. Importantly, at both doses of resveratrol no improvements in mitochondrial function were observed in animals lacking SIRT1. Together these data indicate that SIRT1 plays an essential role in the ability of moderate doses of resveratrol to stimulate AMPK and improve mitochondrial function both in vitro and in vivo.
Key points
Resveratrol, an antioxidant found in red wine, has beneficial effects on cardiac and skeletal muscle function, similar to the effects of endurance exercise training.
Combining resveratrol supplementation with exercise training augments the beneficial effects of exercise alone.
We show that endurance capacity is enhanced in rats whose diet includes resveratrol during a 12 week endurance‐training programme.
Increased endurance was associated with increases in skeletal muscle force, cardiac function, and oxidative metabolism.
Our results establish that resveratrol is an effective ergogenic aid that enhances exercise performance over exercise alone.
Sir2 has been shown to be essential for transcriptional silencing and longevity provided by calorie restriction in Saccharomyces cerevisiae and Caenorhabditis elegans. In this study, we investigated the role for its mammalian homologue, SIRT1, in hematopoietic cells. SIRT1 inhibitor, nicotinamide (NA), promoted and its activator, resveratrol, inhibited the differentiation of murine bone marrow c-Kit(high)Sca-1(+)Lineage(-) (KSL) cells during the culture system ex vivo. To further clarify the roles of SIRT1 in hematopoietic cells, we isolated KSL cells from fetal liver of SIRT1 knockout (KO) mice and cultured them for 5days, because SIRT1 KO mice die shortly after the delivery. In agreement with the results from the experiments using NA and resveratrol, KSL cells isolated from SIRT1 KO mice more apparently differentiated and lost the KSL phenotype than those from wild-type (WT) mice. Furthermore, in each of colony assay, replating assay, or serial transplantation assay, SIRT1 KO KSL cells lost earlier the characteristics of stem cells than WT KSL cells. In addition, we found that SIRT1 maintains prematurity of hematopoietic cells through ROS elimination, FOXO activation, and p53 inhibition. These results suggest that SIRT1 suppresses differentiation of hematopoietic stem/progenitor cells and contributes to the maintenance of stem cell pool.
Malignant glioblastoma represents a challenge in the chemotherapy of brain tumors, because of its aggressive behavior characterized by chemoresistance, infiltrative diffusion, and high rate of recurrence and death. In this study, we used cultured human U87MG cells and primary human glioblastoma cultures to test the anticancer properties of resveratrol (RV), a phytoalexin abundantly present in a variety of dietary products. In U87MG cells, 100 μM RV elicited cell growth arrest by 48 h and bax-mediated cell toxicity by 96 h and greatly limited cell migration and invasion through matrigel. Both in U87MG cells and in primary glioblastoma cultures, the chronic administration of RV (100 μM for up to 96 h) decreased the expression of nestin (a brain (cancer) stem cells marker) but increased that of glial acidic fibrillary protein (a mature glial cell marker) and of βIII-tubulin (a neuronal differentiation marker). Chronic treatment with RV increased the proportion of cells positive for senescence-associated β-galactosidase activity. This is the first report showing the ability of RV to induce glial-like and neuronal-like differentiation in glioblastoma cells. The beneficial effects of chronic RV supplementation lasted up to 96 h after its withdrawal from the culture medium. The present findings support the introduction of pulsed administration of this food-derived molecule in the chemotherapy regimen of astrocytomas.
Cancers are the largest cause of mortality and morbidity in industrialized countries. In the field of the medicinal chemistry of natural products, numerous studies have reported interesting properties of trans-resveratrol as a chemopreventing agent against cancers, inflammation, and viral infection. Tumor growth inhibition has been linked to the ability of resveratrol to arrest cell cycle progression and to trigger cell death. This review focuses on the pathways that mediate resveratrol-induced cell death. Resveratrol impacts on the mitochondrial functions (respiratory chain, oncoproteins, gene expression, etc), in which p53 protein can be involved and its acetylated or phosphorylated forms. This polyphenol also affects death receptor distribution in ceramide-enriched membrane platforms which serve to trap and cluster receptor molecules, and facilitates the formation of a death-inducing signaling complex in the cell. To induce apoptosis, resveratrol also activates the ceramide / sphingomyelin pathway, which promotes ceramide generation and the downstream activation of kinase cascades. Resveratrol can activate alternative pathways to cell death such as those leading to autophagy, senescence or mitotic catastrophe. Furthermore, numerous attempts have been made using resveratrol analogs to improve the molecule's ability to block cell proliferation and induce cell death. Moreover, structural modification of natural phenolics is expected to produce analogs that may be useful tools to study the structure-activity relationships. Lastly, in various cancer types, resveratrol behaves as a chemosensitizer that lowers the threshold of cell death induction by classical anticancer agents and counteracts tumor cell chemoresistance.
The bone protective effects of resveratrol have been demonstrated in several osteoporosis models while the underlying mechanism is largely unclear. In the present study, we evaluated the effects of resveratrol on differentiation and apoptosis of murine osteoclast progenitor RAW 264.7 cells. We found that resveratrol at non-toxic concentrations dose-dependently inhibited RANKL-induced osteoclast differentiation and induced apoptosis. Resveratrol has been shown to be an activator of Sirt1, a NAD(+) dependent protein deacetylase, and has been demonstrated to mimic estrogen. However, we found that although Sirt1 protein was abundantly expressed in RAW264.7 cells, the specific Sirt1 inhibitor EX-527 could not attenuate the inhibition of osteoclastogenesis mediated by resveratrol. Also, the effects of resveratrol could not be attenuated by ICI-182780, a high affinity estrogen receptor antagonist. The central role of reactive oxygen species (ROS) in RANKL-induced osteoclast differentiation has recently been clarified. We found that resveratrol suppressed RANKL-induced ROS generation in a concentration dependent manner. We postulate that the direct inhibitory effects of resveratrol on osteoclastogenesis are mediated via inhibition of ROS generation.
Resveratrol (trans-3,4',5-trihydroxystilbene) is a natural antioxidant with cardiovascular and cancer preventive properties that is currently at the stage of pre-clinical studies for human cancer prevention. Beside its known effects on protein coding genes, one possible mechanism for resveratrol protective activities is by modulating the levels of non-coding RNAs. Here, we analyzed the effects of resveratrol on microRNA populations in human SW480 colon cancer cells. We establish that resveratrol treatment decreases the levels of several oncogenic microRNAs targeting genes encoding Dicer1, a cytoplasmic RNase III producing mature microRNAs from their immediate precursors, tumor-suppressor factors such as PDCD4 or PTEN, as well as key effectors of the TGFβ signaling pathway, while increasing the levels of miR-663, a tumor-suppressor microRNA targeting TGFβ1 transcripts. We also show that, while upregulating several components of the TGFβ signaling pathway such as TGFβ receptors type I (TGFβR1) and type II (TGFβR2), resveratrol decreases the transcriptional activity of SMADs, the main effectors of the canonical TGFβ pathway. Our results establish that protective properties of resveratrol may arise at least in part from its capability to modify the composition of microRNA populations in cells, and suggest that the manipulation of the levels of key microRNAs, such as miR-663, may help to potentiate the anti-cancer and anti-metastatic effects of resveratrol.
Previously we found that a fruit-derived polyphenol fraction caused an inhibition of proliferation and an induction of differentiation markers in Caco-2 human colon cancer cells. In the present work, we sought to determine if individual polyphenols would exert similar actions. Proliferation was inhibited by several polyphenolic molecules including gallic acid, ellagic acid, quercetin and resveratrol. In Caco-2 cells, growth inhibition was accompanied by increased specific activities of two differentiation markers, alkaline phosphatase and dipeptidyl peptidase, but not of aminopeptidase. Increased enzyme activities were not seen in HT29 and SW1116 colon cancer cells. In Caco-2 cells there were additive effects of butyrate or valproate and polyphenolic molecules. Histone acetylation was not greatly affected by the polyphenols. Cycloheximide inhibited protein synthesis in the 3 cell types examined but paradoxically, in Caco-2 cells it caused increased specific activities of alkaline phosphatase and dipeptidyl peptidase. Several plant polyphenols can inhibit the growth of colon cancer cells but increased specific activity of some differentiation markers seen in Caco-2 cells did not appear to be a general phenomenon in colon cancer cells.
Currently, a lack of consensus exists on how best to perform and interpret quantitative real-time PCR (qPCR) experiments. The problem is exacerbated by a lack of sufficient experimental detail in many publications, which impedes a reader's ability to evaluate critically the quality of the results presented or to repeat the experiments.
The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines target the reliability of results to help ensure the integrity of the scientific literature, promote consistency between laboratories, and increase experimental transparency. MIQE is a set of guidelines that describe the minimum information necessary for evaluating qPCR experiments. Included is a checklist to accompany the initial submission of a manuscript to the publisher. By providing all relevant experimental conditions and assay characteristics, reviewers can assess the validity of the protocols used. Full disclosure of all reagents, sequences, and analysis methods is necessary to enable other investigators to reproduce results. MIQE details should be published either in abbreviated form or as an online supplement.
Following these guidelines will encourage better experimental practice, allowing more reliable and unequivocal interpretation of qPCR results.
1. Electrophysiological and immunohistochemical properties during the early stages of muscle differentiation were studied in two myoblastic cell lines, mouse C2C12 and rat L6, and compared to those in myogenic clonal cells derived from the mouse mesodermal stem cell line C3H10T1/2, studied in the preceding paper. 2. Mouse C2C12 cells were induced to differentiate to muscle by changing from 10% fetal calf serum to 2% horse serum in the medium. Most of the C2C12 cells before serum reduction showed ATP-induced slow K+ current. Twelve per cent showed inward rectifier K+ current. They expressed fibronectin and Neural Cell Adhesion Molecule (NCAM). Small spindle-shaped cells at an early stage of muscle differentiation began to appear 24 h after serum reduction. In contrast to cells before serum reduction, only 13% of these spindle-shaped cells showed an ATP response. Most showed tetrodotoxin (TTX)-resistant Na+ current and outward K+ current. Thirty-eight per cent had inward rectifier K+ current. They expressed NCAM but not fibronectin. The T-type Ca2+ current was not observed up to the latest stage of differentiation investigated. 3. Rat L6 cells in maintaining culture medium showed only infrequent ATP responses, but already showed TTX-resistant Na+ current. No clear T-type Ca2+, inward rectifier K+ or outward K+ currents were observed. About one-third of the cells did not express fibronectin. From these results, L6 cells appear to be at a stage near to but slightly earlier than that of C2C12 cells after serum reduction. 4. The properties of the early stages of muscle differentiation in C3H10T1/2 cells, such as the disappearance of ATP-induced K+ current and fibronectin, and the appearance of NCAM, were also seen in C2C12 and L6. However, T-type Ca2+ and inward K+ currents, which were found in the initial stages of C3H10T1/2 muscle differentiation, were not clearly observed in C2C12 and L6. Instead, C2C12 and L6 showed a TTX-resistant Na+ current which was never observed in C3H10T1/2 cells. 5. The properties of the TTX-resistant Na+ current were investigated. In L6 cells, it was reduced to 60% by 1 microM-TTX. It could be evoked by depolarizations to a level above -50 mV with a maximum amplitude at around -15 mV. Steady-state inactivation was detectable with pre-pulses to -100 mV for 100 ms and reached half at pre-pulses of -78 mV. These parameters of inactivation are clearly different from those of the TTX-sensitive Na+ current observed in C3H10T1/2-derived mature muscle cells in the preceding paper.
lin-4 is essential for the normal temporal control of diverse postembryonic developmental events in C. elegans. lin-4 acts by negatively regulating the level of LIN-14 protein, creating a temporal decrease in LIN-14 protein starting in the first larval stage (L1). We have cloned the C. elegans lin-4 locus by chromosomal walking and transformation rescue. We used the C. elegans clone to isolate the gene from three other Caenorhabditis species; all four Caenorhabditis clones functionally rescue the lin-4 null allele of C. elegans. Comparison of the lin-4 genomic sequence from these four species and site-directed mutagenesis of potential open reading frames indicated that lin-4 does not encode a protein. Two small lin-4 transcripts of approximately 22 and 61 nt were identified in C. elegans and found to contain sequences complementary to a repeated sequence element in the 3' untranslated region (UTR) of lin-14 mRNA, suggesting that lin-4 regulates lin-14 translation via an antisense RNA-RNA interaction.
In this study a comparison was made between results obtained from resveratrol dosages which have been shown to be pharmacologically active, in vitro and in vivo, and the results of plasma and tissue concentrations obtained after a single administration or after prolonged administration of red wine with a known resveratrol content. The dosages used by different investigators in the tests are very different and, in general, rather high in relation to the concentrations which are found in wine or grapes. The results of our tests on platelet aggregation confirm that even with modest dosages of resveratrol, a pharmacological effect can be observed, and that these dosages can be compatible with the resveratrol concentrations obtained after oral administration. The data obtained from these tests on animals can lead to the conclusion that even an average drinker of wine can, particularly in the long term, absorb a sufficient quantity of resveratrol to explain the beneficial effect of red wine on health, which has been observed in epidemiological studies carried out in populations whose daily diet includes the drinking of wine.
Real-time reverse transcription followed by polymerase chain reaction (RT-PCR) is the most suitable method for the detection and quantification of mRNA. It offers high sensitivity, good reproducibility and a wide quantification range. Today, relative expression is increasingly used, where the expression of a target gene is standardised by a non-regulated reference gene. Several mathematical algorithms have been developed to compute an expression ratio, based on real-time PCR efficiency and the crossing point deviation of an unknown sample versus a control. But all published equations and available models for the calculation of relative expression ratio allow only for the determination of a single transcription difference between one control and one sample. Therefore a new software tool was established, named REST (relative expression software tool), which compares two groups, with up to 16 data points in a sample and 16 in a control group, for reference and up to four target genes. The mathematical model used is based on the PCR efficiencies and the mean crossing point deviation between the sample and control group. Subsequently, the expression ratio results of the four investigated transcripts are tested for significance by a randomisation test. Herein, development and application of REST is explained and the usefulness of relative expression in real-time PCR using REST is discussed. The latest software version of REST and examples for the correct use can be downloaded at http://www.wzw.tum.de/gene-quantification/.
This work reports significant advances on the transport in hepatic cells of resveratrol, a natural polyphenol with potent protective properties. First, we describe a new simple technique to qualitatively follow resveratrol cell uptake and intracellular distribution, based on resveratrol fluorescent properties. Second, the time-course study and the quantification of (3)H-labelled resveratrol uptake have been performed using human hepatic derived cells (HepG2 tumor cells) and hepatocytes. The temperature-dependence of the kinetics of uptake as well as the cis-inhibition experiments agree with the involvement of a carrier-mediated transport in addition to passive diffusion. The decrease of passive uptake resulting from resveratrol binding to serum proteins brings to light a mediated mechanism in physiological situation.
5'-AMP-activated protein kinase (AMPK) is important for metabolic sensing. We used AMPKgamma3 mutant-overexpressing Tg-Prkag3(225Q) and AMPKgamma3-knockout Prkag3-/- mice to determine the role of the AMPKgamma3 isoform in exercise-induced metabolic and gene regulatory responses in skeletal muscle. Mice were studied after 2 h swimming or 2.5 h recovery. Exercise increased basal and insulin-stimulated glucose transport, with similar responses among genotypes. In Tg-Prkag3(225Q) mice, acetyl-CoA carboxylase (ACC) phosphorylation was increased and triglyceride content was reduced after exercise, suggesting that this mutation promotes greater reliance on lipid oxidation. In contrast, ACC phosphorylation and triglyceride content was similar between wild-type and Prkag3-/- mice. Expression of genes involved in lipid and glucose metabolism was altered by genetic modification of AMPKgamma3. Expression of lipoprotein lipase 1, carnitine palmitoyl transferase 1b, and 3-hydroxyacyl-CoA dehydrogenase was increased in Tg-Prkag3(225Q) mice, with opposing effects in Prkag3-/- mice after exercise. GLUT4, hexokinase II (HKII), and glycogen synthase mRNA expression was increased in Tg-Prkag3(225Q) mice after exercise. GLUT4 and HKII mRNA expression was increased in wild-type mice and blunted in Prkag3-/- mice after recovery. In conclusion, the Prkag3(225Q) mutation, rather than presence of a functional AMPKgamma3 isoform, directly promotes metabolic and gene regulatory responses along lipid oxidative pathways in skeletal muscle after endurance exercise.
While fruits and vegetables are recommended for prevention of cancer and other diseases, their active ingredients (at the molecular level) and their mechanisms of action less well understood. Extensive research during the last half century has identified various molecular targets that can potentially be used not only for the prevention of cancer but also for treatment. However, lack of success with targeted monotherapy resulting from bypass mechanisms has forced researchers to employ either combination therapy or agents that interfere with multiple cell-signaling pathways. In this review, we present evidence that numerous agents identified from fruits and vegetables can interfere with several cell-signaling pathways. The agents include curcumin (turmeric), resveratrol (red grapes, peanuts and berries), genistein (soybean), diallyl sulfide (allium), S-allyl cysteine (allium), allicin (garlic), lycopene (tomato), capsaicin (red chilli), diosgenin (fenugreek), 6-gingerol (ginger), ellagic acid (pomegranate), ursolic acid (apple, pears, prunes), silymarin (milk thistle), anethol (anise, camphor, and fennel), catechins (green tea), eugenol (cloves), indole-3-carbinol (cruciferous vegetables), limonene (citrus fruits), beta carotene (carrots), and dietary fiber. For instance, the cell-signaling pathways inhibited by curcumin alone include NF-kappaB, AP-1, STAT3, Akt, Bcl-2, Bcl-X(L), caspases, PARP, IKK, EGFR, HER2, JNK, MAPK, COX2, and 5-LOX. The active principle identified in fruit and vegetables and the molecular targets modulated may be the basis for how these dietary agents not only prevent but also treat cancer and other diseases. This work reaffirms what Hippocrates said 25 centuries ago, let food be thy medicine and medicine be thy food.
Diminished mitochondrial oxidative phosphorylation and aerobic capacity are associated with reduced longevity. We tested whether resveratrol (RSV), which is known to extend lifespan, impacts mitochondrial function and metabolic homeostasis. Treatment of mice with RSV significantly increased their aerobic capacity, as evidenced by their increased running time and consumption of oxygen in muscle fibers. RSV's effects were associated with an induction of genes for oxidative phosphorylation and mitochondrial biogenesis and were largely explained by an RSV-mediated decrease in PGC-1alpha acetylation and an increase in PGC-1alpha activity. This mechanism is consistent with RSV being a known activator of the protein deacetylase, SIRT1, and by the lack of effect of RSV in SIRT1(-/-) MEFs. Importantly, RSV treatment protected mice against diet-induced-obesity and insulin resistance. These pharmacological effects of RSV combined with the association of three Sirt1 SNPs and energy homeostasis in Finnish subjects implicates SIRT1 as a key regulator of energy and metabolic homeostasis.
Resveratrol (trans-3,5,4'-trihydroxystilbene), a polyphenolic compound found largely in the skins of red grapes, has been used as a nutritional supplement or an investigational new drug for prevention of cardiovascular diseases. Previous reports showed that resveratrol had a protective effect against oxidative agent-induced cell injury. Our studies indicate that resveratrol plays a role in the differentiation of cardiomyoblasts. The cardiomyoblast cell line, H9c2, was exposed to 30-120 microM resveratrol for up to 5 days. Resveratrol inhibits cardiomyoblast proliferation without causing cells injury. Moreover, resveratrol treatment modulated the differentiation of morphological characteristics including elongation and cell fusion in cardiomyoblasts. Proliferation and differentiation of H9c2 cells were further revealed by measurement of the mRNA expression of a cell cycle marker (CDK2), a differentiation marker (myogenin), and a contractile apparatus protein (MLC-2). Gene expression analysis revealed that resveratrol promoted entry into cell cycle arrest but extended the myogenic differentiation progress. These results have implications for the role of resveratrol in modulating cell cycle control and differentiation in cardiomyoblasts.
these changes may potentially have a substantial impact in the differentiation/proliferation process. Indeed, miR-20b, downregulated by resveratrol putatively targets transcripts encoding Myo1d and Myf5 myogenic regulatory factors
- Interestingly
Interestingly, these changes may potentially have a substantial impact in the differentiation/proliferation process. Indeed, miR-20b, downregulated by resveratrol putatively targets transcripts encoding Myo1d and Myf5 myogenic regulatory factors [25].