Icariin ameliorate Alzheimer's disease by influencing SIRT1 and inhibiting Aß cascade pathogenesis.

Of all types of dementia, Alzheimer's disease is the type that has the highest proportion of cases and is the cause of substantial medical and economic burden. The mechanism of Alzheimer's disease is closely associated with the aggregation of amyloid-ß protein and causes neurotoxicity and extracellular accumulation in the brain and to intracellular neurofibrillary tangles caused by tau protein hyperphosphorylation in the brain tissue. Previous studies have demonstrated that sirtuin1 downregulation is involved in the pathological mechanism of Alzheimer's disease. The decrease of sirtuin1 level would cause Alzheimer's disease by means of promoting the amyloidogenic pathway to generate amyloid-ß species and thereby triggering amyloid-ß cascade reaction, such as tau protein hyperphosphorylation, neuron autophagy, neuroinflammation, oxidative stress, and neuron apoptosis. Currently, there is no effective treatment for Alzheimer's disease, it is necessary to develop new treatment strategies. According to the theory of traditional Chinese medicine and based on the mechanism of the disease, tonifying the kidneys is one of the principles for the treatment of Alzheimer's disease and Epimedium is a well-known Chinese medicine for tonifying kidney. Therefore, investigating the influence of the components of Epimedium on the pathological characteristics of Alzheimer's disease may provide a reference for the treatment of Alzheimer's disease in the future. In this article, we summarise the effects and mechanism of icariin, the main ingredient extracted from Epimedium, in ameliorating Alzheimer's disease by regulating sirtuin1 to inhibit amyloid-ß protein and improve other amyloid-ß cascade pathogenesis.

Hyperoside Attenuate Inflammation in HT22 Cells via Upregulating SIRT1 to Activities Wnt/ß-Catenin and Sonic Hedgehog Pathways.

Neuroinflammation plays important roles in the pathogenesis and progression of altered neurodevelopment, sensorineural hearing loss, and certain neurodegenerative diseases. Hyperoside (quercetin-3-O-ß-D-galactoside) is an active compound isolated from Hypericum plants. In this study, we investigate the protective effect of hyperoside on neuroinflammation and its possible molecular mechanism. Lipopolysaccharide (LPS) and hyperoside were used to treat HT22 cells. The cell viability was measured by MTT assay. The cell apoptosis rate was measured by flow cytometry assay. The mRNA expression levels of interleukin-1ß (IL-1ß), interleukin-6 (IL-6), interleukin-8 (IL-8), and tumor necrosis factor-α (TNF-α) were determined by quantitative reverse transcription polymerase chain reaction. The levels of oxidative stress indices superoxide dismutase (SOD), reactive oxygen species (ROS), catalase (CAT), glutathione (GSH), and malondialdehyde (MDA) were measured by the kits. The expression of neurotrophic factor and the relationship among hyperoside, silent mating type information regulation 2 homolog-1 (SIRT1) and Wnt/ß-catenin, and sonic hedgehog was examined by western blotting. In the LPS-induced HT22 cells, hyperoside promotes cell survival; alleviates the level of IL-1ß, IL-6, IL-8, TNF-α, ROS, MDA, Bax, and caspase-3; and increases the expression of CAT, SOD, GSH, Bcl-2, BDNF, TrkB, and NGF. In addition, hyperoside upregulated the expression of SIRT1. Further mechanistic investigation showed that hyperoside alleviated LPS-induced inflammation, oxidative stress, and apoptosis by upregulating SIRT1 to activate Wnt/ß-catenin and sonic hedgehog pathways. Taken together, our data suggested that hyperoside acts as a protector in neuroinflammation.

Modulation of Sirt1 and FoxO1 on Hypothalamic Leptin-Mediated Sympathetic Activation and Inflammation in Diet-Induced Obese Rats.

Background Hypothalamic leptin-mediated signaling contributes to the exaggerated sympatho-excitation and increased blood pressure in obesity-associated hypertension. The aim of the study was to investigate the roles of energy-sensing enzyme sirtuin1 (Sirt1) and forkhead box protein O1 (FoxO1) on the hypothalamic leptin-mediated high sympathetic nerve activity and inflammation in obesity. Methods and Results Sprague Dawley rats were fed with high-fat diet (HFD) for 12 weeks. In vivo, the potential of Srit1 and FoxO1 in the sympathetic effects of leptin was investigated via siRNA injection to knockdown Sirt1 or FoxO1 gene in the arcuate nucleus (ARCN) of hypothalamus in rats. In vitro, the effects of Sirt1 or FoxO1 on leptin-mediated inflammation were observed in proopiomelanocortin (POMC) and microglial cells. Knockdown Sirt1 by siRNA significantly reduced the renal sympathetic nerve activity (RSNA) and blood pressure responses to leptin injection in the ARCN in the HFD rats. Conversely, knockdown FoxO1 significantly enhanced the RSNA and blood pressure responses to leptin injection in the HFD rats. Knockdown Sirt1 reduced the levels of pro-inflammatory cytokines interleukin 6 (IL-6), tumor necrosis factor α (TNF-α), interleukin 1ß (IL-1ß), C1q/TNF-related protein-1 (CTRP1), and immune cell infiltration in the ARCN in the HFD rats. Knockdown FoxO1 significantly increased the level of IL-6 in the ARCN of HFD rats. In cultured hypothalamic POMC and microglial cells, knockdown Sirt1 significantly reduced leptin-induced IL-6 expression, affected the levels of AMP-activated protein kinase (AMPK) and serine/threonine-specific protein kinase (Akt). Knockdown FoxO1 significantly increased leptin-induced IL-6 in both POMC cells and microglial cells. Conclusions These data suggest that both Sirt1 and FoxO1 are the key modulators of leptin signaling in the hypothalamus contributed to the over sympathetic activation and inflammation in obesity.

Juzentaihoto Suppresses Muscle Atrophy and Decreased Motor Function in SAMP8 Mice.

Sarcopenia is a disease whose symptoms include decreased muscle mass and weakened muscle strength with age. In sarcopenia, decreased production of insulin-like growth factor-1 (IGF-1) increases ubiquitin ligases, such as Atrogin1 and Muscle RING-Finger Protein-1 (MuRF1), by activating forkhead box O (FOXO), and inflammatory cytokines and oxidative stress increase the expression of ubiquitin ligases by activating the transcription factor nuclear factor-kappa B (NF-κB). In addition, increased levels of ubiquitin ligases cause skeletal muscle atrophy. Conversely, sirtuin 1 (Sirt1) is known to regulate the expression of ubiquitin ligases by suppressing the activities of NF-κB and FOXO. In this study, we evaluated the effect that juzentaihoto hot water extract (JTT) has on skeletal muscle atrophy and motor function by administering it to senescence-accelerated mouse prone-8 (SAMP8). The group treated with JTT displayed larger gastrocnemius muscle (GA) and extensor digitorum longus (EDL) weights, larger GA muscle fiber cross-sectional areas, and motor function decline during rota-rod tests. JTT also increased IGF-1 serum levels, as well as mRNA Sirt1 levels in GA. Serum levels of tumor necrosis factor-α, interleukin-6, and mRNA levels of Atrogin1 and MuRF1 in GA were reduced by JTT. The muscle fiber cross-sectional area of GA was correlated with the mRNA levels of Sirt1 in GA. The results of this study suggested that JTT administration suppresses skeletal muscle atrophy and motor function decline in SAMP8 mice. This effect may be associated with the increased expression levels of Sirt1 and IGF-1 by JTT.

A Synthetic Snake-Venom-Based Tripeptide Protects PC12 Cells from the Neurotoxicity of Acrolein by Improving Axonal Plasticity and Bioenergetics.

The synthetic peptide p-BTX-I is based on the native peptide (formed by glutamic acid, valine and tryptophan) isolated from Bothrops atrox venom. We have previously demonstrated its neuroprotective and neurotrophic properties in PC12 cells treated with the dopaminergic neurotoxin 1-methyl-4-phenylpyridinium (MPP+). Now, we have investigated the neuroprotective effects and mechanisms of p-BTX-I against the toxicity of acrolein in PC12 cells. Studies have demonstrated that acrolein might play an important role in the etiology of Alzheimer's disease (AD), which is characterized by neuronal and synaptic loss. Our results showed that not only acrolein reduced cell differentiation and cell viability, but also altered the expression of markers of synaptic communication (synapsin I), energy metabolism (AMPK-α, Sirt I and glucose uptake), and cytoskeleton (ß-III-tubulin). Treatment with p-BTX-I increased the percentage of differentiation in cells treated with acrolein and significantly attenuated cell viability loss, besides counteracting the negative effects of acrolein on synapsin I, AMPK-α, Sirt I, glucose uptake, and ß-III-tubulin. Additionally, p-BTX-I alone increased the expression of apolipoprotein E (apoE) gene, associated with the proteolytic degradation of ß-amyloid peptide aggregates, a hallmark of AD. Taken together, these findings demonstrate that p-BTX-I protects against acrolein-induced neurotoxicity and might be a tool for the development of novel drugs for the treatment of neurodegenerative diseases.

Chronic whole-body heat treatment relieves atherosclerotic lesions, cardiovascular and metabolic abnormalities, and enhances survival time restoring the anti-inflammatory and anti-senescent heat shock response in mice.

Unhealthy lifestyle persistently feeds forward inflammation in metabolic organs thus imposing senescence-associated secretory phenotype (SASP), as observed in obesity and type 2 diabetes. However, SASP blocks physiological resolution of inflammation by suppressing the anti-inflammatory and anti-senescent heat shock (HS) response, i.e., the gene program centered in heat shock factor-1 (HSF1)-dependent expression heat shock proteins (HSPs). As SASP-inducing factors are not removed, leading to the perpetuation of inflammation, we argued that SIRT1-HSF1-HSP axis might also be suppressed in atherosclerosis, which could be reversible by heat treatment (HT), the most powerful HS response trigger. LDLr-/- adult mice were fed on high-fat/high-cholesterol diet from the age of 90 days until the end of study (age of 270 days). After 120 days under atherosclerotic diet, the animals were submitted to either whole-body HT (n = 42; 40 °C) or sham (n = 59; 37 °C) treatment (15 min/session), under anesthesia, once a week, for 8 weeks, being echographically and metabolically monitored. Aortic expressions of SIRT1, HSF1, HSP27, HSP72 and HSP73 were progressively depressed in atherosclerotic animals, as compared to normal (LDLr+/+; n = 25) healthy counterparts, which was paralleled by increased expression of NF-κB-dependent VCAM1 adhesion molecule. Conversely, HT completely reversed suppression of the above HS response proteins, while markedly inhibiting both VCAM1 expression and NF-κB DNA-binding activity. Also, HT dramatically reduced plasma levels of TG, total cholesterol, LDL-cholesterol, oxidative stress, fasting glucose and insulin resistance while rising HDL-cholesterol levels. HT also decreased body weight gain, visceral fat, cellular infiltration and aortic fatty streaks, and heart ventricular congestive hypertrophy, thereby improving aortic blood flow and myocardial performance (Tei) indices. Remarkably, heat-treated mice stopped dying after the third HT session (= 8 human years), suggesting a curative effect. Therefore, evolution of atherosclerosis is associated with suppression of the anti-inflammatory and anti-senescent SIRT1-HSF1-HSP molecular axis, which is refreshed by chronic heat treatment.

The protective effects of compatibility of Aconiti Lateralis Radix Praeparata and Zingiberis Rhizoma on rats with heart failure by enhancing mitochondrial biogenesis via Sirt1/PGC-1α pathway.

Aconiti Lateralis Radix Praeparata ("Fuzi" in Chinese) in combination with Zingiberis Rhizoma ("Ganjiang" in Chinese) is commonly applied for the treatment of heart failure for thousands of years in China. However, its therapeutic mechanism is still poorly defined. This study aimed to investigate whether the compatibility of Fuzi and Ganjiang can protect rats with acute heart failure by enhancing mitochondrial biogenesis via Sirt1/PGC-1α signaling pathway. Hemodynamic parameters, including heart rate and left ventricular maximal rate of pressure rise and decline, were recorded in rats with acute heart failure induced by Propafenone hydrochloride. The serum levels of cardiac enzymes, including creatine kinase, lactate dehydrogenase, brain natriuretic peptide and cardiac troponin T, were also determined. The gene and protein levels of Sirtuin 1 (Sirt1), peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) and their downstream transcription factors were measured as well. The results indicated that Fuzi-Ganjiang herbal couple provided more significant benefits by restoring the left ventricular function and cardiac enzyme activities in comparison with their single use. Moreover, this herbal couple possessed a significant cardio-protection by increasing both gene and protein levels of Sirt1 and PGC-1α. In conclusion, the compatibility of Fuzi and Ganjiang had better therapeutic effect than their single use against failing heart, and the underlying mechanisms were partially through increasing mitochondrial biogenesis via Sirt1/PGC-1α pathway.

Hydrogen Rich Water Attenuates Renal Injury and Fibrosis by Regulation Transforming Growth Factor-ß Induced Sirt1.

The current research was designed to study the role of hydrogen in renal fibrosis and the renal epithelial to mesenchymal transition (EMT) induced by transforming growth factor-ß1 (TGF-ß1). Hydrogen rich water (HW) was used to treat animal and cell models. Unilateral ureteral obstruction (UUO) was performed on Balb/c mice to create a model of renal fibrosis. Human kidney proximal tubular epithelial cells (HK-2 cells) were treated with TGF-ß1 for 36 h to induce EMT. Serum creatinine (Scr) and blood urea nitrogen (BUN) were measured to test renal function, in addition, kidney histology and immunohistochemical staining of alpha-smooth muscle actin (α-SMA) positive cells was performed to examine the morphological changes. The treatment with UUO induced a robust fibrosis of renal interstitium, shrink of glomerulus and partial fracture of basement membrane. Renal function was also impaired in the experimental group with UUO, with an increase of Scr and BUN in serum. After that, Western-blot was performed to examine the expression of α-SMA, fibronectin, E-cadherin, Smad2 and Sirtuin-1 (Sirt1). The treatment with HW attenuated the development of fibrosis and deterioration of renal function in UUO model. In HK-2 cells, the pretreatment of HW abolished EMT induced by TGF-ß1. The down-regulation the expression of Sirt1 induced by TGF-ß1 which was dampened by the treatment with HW. Sirtinol, a Sirt1 inhibitor, reversed the effect of HW on EMT induced by TGF-ß1. HW can inhibit the development of fibrosis in kidney and prevents HK-2 cells from undergoing EMT which is mediated through Sirt1, a downstream molecule of TGF-ß1.

Omega-3 polyunsaturated fatty acids suppress the inflammatory responses of lipopolysaccharide-stimulated mouse microglia by activating SIRT1 pathways.

Obesity and diabetes are known risk factors for dementia, and it is speculated that chronic neuroinflammation contributes to this increased risk. Microglia are brain-resident immune cells modulating the neuroinflammatory state. Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), the major ω-3 polyunsaturated fatty acids (PUFAs) of fish oil, exhibit various effects, which include shifting microglia to the anti-inflammatory phenotype. To identify the molecular mechanisms involved, we examined the impact of EPA, DHA, and EPA+DHA on the lipopolysaccharide (LPS)-induced cytokine profiles and the associated signaling pathways in the mouse microglial line MG6. Both EPA and DHA suppressed the production of the pro-inflammatory cytokines TNF-α and IL-6 by LPS-stimulated MG6 cells, and this was also observed in LPS-stimulated BV-2 cells, the other microglial line. Moreover, the EPA+DHA mixture activated SIRT1 signaling by enhancing mRNA level of nicotinamide phosphoribosyltransferase (NAMPT), cellular NAD+ level, SIRT1 protein deacetylase activity, and SIRT1 mRNA levels in LPS-stimulated MG6. EPA+DHA also inhibited phosphorylation of the stress-associated transcription factor NF-κB subunit p65 at Ser536, which is known to enhance NF-κB nuclear translocation and transcriptional activity, including cytokine gene activation. Further, EPA+DHA increased the LC3-II/LC3-I ratio, an indicator of autophagy. Suppression of TNF-α and IL-6 production, inhibition of p65 phosphorylation, and autophagy induction were abrogated by a SIRT1 inhibitor. On the other hand, NAMPT inhibition reversed TNF-α suppression but not IL-6 suppression. Accordingly, these ω-3 PUFAs may suppress neuroinflammation through SIRT1-mediated inhibition of the microglial NF-κB stress response and ensue pro-inflammatory cytokine release, which is implicated in NAMPT-related and -unrelated pathways.

Metabolism of 20(S)-Ginsenoside Rg2 by Rat Liver Microsomes: Bioactivation to SIRT1-Activating Metabolites.

20(S)-Ginsenoside Rg2 (1) has recently become a hot research topic due to its potent bioactivities and abundance in natural sources such as the roots, rhizomes and stems-leaves of Panax ginseng. However, due to the lack of studies on systematic metabolic profiles, the prospects for new drug development of 1 are still difficult to predict, which has become a huge obstacle for its safe clinical use. To solve this problem, investigation of the metabolic profiles of 1 in rat liver microsomes was first carried out. To identify metabolites, a strategy of combined analyses based on prepared metabolites by column chromatography and ultra-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (UPLC-Q-TOF/MS) was performed. As a result, four metabolites M1-M4, including a rare new compound named ginsenotransmetin A (M1), were isolated and the structures were confirmed by spectroscopic analyses. A series of metabolites of 1, MA-MG, were also tentatively identified by UPLC-Q-TOF/MS in rat liver microsomal incubate of 1. Partial metabolic pathways were proposed. Among them, 1 and its metabolites M1, M3 and M4 were discovered for the first time to be activators of SIRT1. The SIRT1 activating effects of the metabolite M1 was comparable to those of 1, while the most interesting SIRT1 activatory effects of M3 and M4 were higher than that of 1 and comparable with that of resveratrol, a positive SIRT1 activator. These results indicate that microsome-dependent metabolism may represent a bioactivation pathway for 1. This study is the first to report the metabolic profiles of 1 in vitro, and the results provide an experimental foundation to better understand the in vivo metabolic fate of 1.

High dose of epigallocatechin-3-gallate inhibits proliferation and induces apoptosis of H9C2 cardiomyocytes through down-regulation of SIRT1.

BACKGROUND: Previous studies have suggested that high doses of (-)-epigallocatechin-3-gallate (EGCG) can induce toxicity in the liver, kidneys, and intestine. However, there have been no reports of myocardiotoxicity following treatment with EGCG. In this study, we investiged the proliferation and apoptosis of H9C2 cardiomyocytes treated with high dose of EGCG. METHODS: Cell proliferation was measured by CCK8 assay, cell apoptosis rate was evaluated by TUNEL assay, and the expression alterations of Sirtuin 1 (SIRT1) protein was detected by Western blotting. RESULTS: EGCG inhibits proliferation and induces apoptosis in time- and dose-dependent manner in H9C2 cardiomyocytes. SIRT1 participates in the inhibitory effect of EGCG on cell proliferation and apoptosis induction in H9C2 cardiomyocytes. CONCLUSION: This study demonstrates that high doses of EGCG inhibit proliferation and induce apoptosis in H9C2 cardiomyocytes. Down-regulation of SIRT 1 protein expression may be involved.

Novel role of resveratrol: suppression of high-mobility group protein box 1 nucleocytoplasmic translocation by the upregulation of sirtuin 1 in sepsis-induced liver injury.

BACKGROUND: High-mobility group protein box 1 (HMGB1) is essential in the response to injury during sepsis. We hypothesized that resveratrol (RESV) administration would inhibit nuclear-cytoplasmic HMGB1 translocation in hepatocytes, which is associated with sirtuin 1 (SIRT1) upregulation. We investigated the regulatory role of SIRT1 in HMGB1 nucleocytoplasmic translocation and its effect on sepsis-induced liver injury. METHODS: Rats were randomly assigned to pretreatment with RESV (60 mg/kg per day), nicotinamide (60 mg/kg per day), or vehicle (olive oil), which was administered by gavage for 3 days directly before cecal ligation and puncture was performed to induce sepsis. Parallel control groups were established. Rats were killed 24 h after surgery, and cytokine production, histology, apoptosis, SIRT1, serum HMGB1, nuclear and cytoplasmic HMGB1/ac-HMGB1, and the interaction between SIRT1 and HMGB1 were evaluated. In vitro evaluations were performed in human liver L02 cells subjected to lipopolysaccharide-induced injury, and siRNA-mediated SIRT1 knockdown experiments were performed. RESULTS: Sepsis-induced serum aminotransferase activities and proinflammatory chemokine levels were reduced by RESV pretreatment, which also improved liver histological parameters in association with SIRT1 upregulation. Resveratrol inhibited HMGB1 cytoplasmic translocation. Nicotinamide, an SIRT1 inhibitor, reduced the SIRT1-mediated suppression of HMGB1 translocation and aggravated cecal ligation and puncture-induced liver damage. Sirtuin 1 knockdown in vitro confirmed that RESV increased the SIRT1-mediated repression of HMGB1 translocation. In vivo, SIRT1 and HMGB1 physically interacted in the nucleus, and SIRT1 regulated HMGB1 acetylation in response to septic liver injury. CONCLUSIONS: Resveratrol protects against sepsis-induced liver injury through the SIRT1-mediated HMGB1 nucleocytoplasmic translocation pathway, a new potential therapeutic target in sepsis-induced liver injury.

Liquid fructose downregulates Sirt1 expression and activity and impairs the oxidation of fatty acids in rat and human liver cells.

Fructose ingestion is associated with the production of hepatic steatosis and hypertriglyceridemia. For fructose to attain these effects in rats, simultaneous induction of fatty acid synthesis and inhibition of fatty acid oxidation is required. We aimed to determine the mechanism involved in the inhibition of fatty acid oxidation by fructose and whether this effect occurs also in human liver cells. Female rats were supplemented or not with liquid fructose (10% w/v) for 7 or 14 days; rat (FaO) and human (HepG2) hepatoma cells, and human hepatocytes were incubated with fructose 25mM for 24h. The expression and activity of the enzymes and transcription factors relating to fatty acid ß-oxidation were evaluated. Fructose inhibited the activity of fatty acid ß-oxidation only in livers of 14-day fructose-supplemented rats, as well as the expression and activity of peroxisome proliferator activated receptor α (PPARα). Similar results were observed in FaO and HepG2 cells and human hepatocytes. PPARα downregulation was not due to an osmotic effect or to an increase in protein-phosphatase 2A activity caused by fructose. Rather, it was related to increased content in liver of inactive and acetylated peroxisome proliferator activated receptor gamma coactivator 1α, due to a reduction in sirtuin 1 expression and activity. In conclusion, fructose inhibits liver fatty acid oxidation by reducing PPARα expression and activity, both in rat and human liver cells, by a mechanism involving sirtuin 1 down-regulation.

Extract of Bryophyllum laetivirens reverses etoposide resistance in human lung A549 cancer cells by downregulation of NF-κB.

Since multidrug resistance (MDR) is one of the main reasons for failure in cancer treatment, its suppression may increase the efficacy of cancer therapy. In the present study we attempted to identify a new and effective anticancer drug against MDR cancer cells. We first found that lung cancer A549 cells resistant to etoposide (A549RT-eto) exhibit upregulation of NF-κB and SIRT1 in comparison to A549 parental cells. During a search for anticancer drug candidates from medicinal plant sources, we found that an extract fraction (F14) of Bryophyllum laetivirens leaves downregulated expression of NF-κB and SIRT1, sensitizing the levels of A549RT-eto cells to apoptosis through downregulation of P-glycoprotein (P-gp), which is encoded by the MDR1 gene. To address whether NF-κB is involved in resistance to etoposide through P-gp, we treated A549RT-eto cells with Bay11-7802, an inhibitor of NF-κB. We then observed that Bay11-7802 treatment reduced P-gp expression levels, and furthermore combined treatment with the F14 extract and Bay11-7802 accelerated apoptosis through a decrease in P-gp levels, suggesting that NF-κB is involved in MDR. To address whether upregulation of SIRT1 is involved in resistance to etoposide through P-gp, we treated A549RT-eto cells with SIRT1 siRNA or nicotinamide (NAM), an inhibitor of SIRT1. we found that suppression of SIRT1 did not reduce P-gp levels. furthermore, the combined treatment with the F14 extract, and SIRT1 siRNA or NAM did not accelerate apoptosis, indicating that SIRT1 is not involved in the regulation of P-gp levels in A549RT-eto cells. Taken together, we suggest that upregulation of NF-κB determines etoposide resistance through P-gp expression in human A549 lung cancer cells. We herein demonstrated that B. laetivirens extract reverses etoposide resistance in human A549 lung cancer cells through downregulation of NF-κB.

SirT1 mediates hyperbaric oxygen preconditioning-induced ischemic tolerance in rat brain.

Our previous studies have shown that hyperbaric oxygen preconditioning (HBO-PC) induces tolerance to cerebral ischemia/reperfusion (I/R). This study aimed to investigate whether SirT1, a class III histone deacetylase, is involved in neuroprotection elicited by HBO-PC in animal and cell culture models of ischemia. Rats were subjected to middle cerebral artery occlusion for 120 minutes after HBO-PC (once a day for 5 days). Primary cultured cortical neurons were exposed to 2 hours of HBO-PC after 2 hours of oxygen-glucose deprivation (OGD). We showed that HBO-PC increased SirT1 protein and mRNA expression, promoted neurobehavioral score, reduced infarct volume, and improved morphology at 24 hours and 7 days after cerebral I/R. Neuroprotection of HBO-PC was attenuated by SirT1 inhibitor EX527 and SirT1 knockdown by short interfering RNA (siRNA), whereas it was mimicked by SirT1 activator resveratrol. Furthermore, HBO-PC enhanced SirT1 expression and cell viability and reduced lactate dehydrogenase release 24 hours after OGD/re-oxygenation. The neuroprotective effect of HBO-PC was emulated through upregulating SirT1 and, reversely, attenuated through downregulating SirT1. The modulation of SirT1 was made by adenovirus infection carrying SirT1 or SirT1 siRNA. Besides, SirT1 increased B-cell lymphoma 2 (Bcl-2) expression and decrease cleaved caspase 3. These results indicate that SirT1 mediates HBO-PC-induced tolerance to cerebral I/R through inhibition of apoptosis.

Relevance of vascular peroxisome proliferator-activated receptor γ coactivator-1α to molecular alterations in atherosclerosis.

Peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) is emerging as a novel factor that plays a critical role in integrating signalling pathways in the control of cellular and systemic metabolism. We investigated the role of vascular expression of PGC-1α and related factors, such as sirtuin 1 (SIRT1), peroxisome proliferator-activated receptor γ (PPARγ) and adiponectin, during the atherosclerotic process. Endothelial function, vascular superoxide anion production and inflammatory mediators were also evaluated. This study was carried out in male New Zealand rabbits fed a diet containing 0.5% cholesterol and 14% coconut oil for 8 weeks. Animals developed mixed dyslipidaemia and atherosclerotic lesions, which were associated with endothelial dysfunction, aortic overproduction of superoxide anions and inflammation. Expression of PGC-1α, SIRT1, PPARγ and adiponectin was reduced (P<0.05) in aorta from atherosclerotic rabbits. Levels of PGC-1α were correlated negatively (P<0.05) with total cholesterol levels, aortic superoxide anion production and tumour necrosis factor-α expression, and positively (P<0.05) with maximal relaxation in response to acetylcholine. The observed results suggest that PGC-1α could be considered to be a link between the main atherosclerotic processes (endothelial dysfunction, oxidation and inflammation) and alterations of other factors involved in vascular wall integrity, such as SIRT1, PPARγ and adiponectin.

Evidence for possible period 2 gene mediation of the effects of alcohol exposure during the postnatal period on genes associated with maintaining metabolic signaling in the mouse hypothalamus.

BACKGROUND: Animals exposed to alcohol during the developmental period develop circadian disturbances and metabolic problems that often persist during their adult period. In order to study whether alcohol and the circadian clock interact to alter metabolic signaling in the hypothalamus, we determined whether postnatal alcohol feeding in mice permanently alters metabolic sensing in the hypothalamus. Furthermore, we evaluated whether the effect of circadian disruption via Period 2 (Per2) gene mutation prevents alcohol's effects on metabolic signaling in the hypothalamus. METHODS: Per2 mutant and wild-type male and female mice of the same genetic background were given a milk formula containing ethanol (EtOH; 11.34% vol/vol) from postnatal day (PD) 2 to 7 and used for gene expression and peptide level determinations in the hypothalamus at PD7 and PD90. RESULTS: We report here that postnatal alcohol feeding reduces the expression of proopiomelanocortin (Pomc) gene and production of ß-endorphin and α-melanocyte stimulating hormone (α-MSH) in the hypothalamus that persists into adulthood. In addition, expressions of metabolic sensing genes in the hypothalamus were also reduced as a consequence of postnatal alcohol exposure. These effects were not sex-specific and were observed in both males and females. Mice carrying a mutation of the Per2 gene did not show any reductions in hypothalamic levels of Pomc and metabolic genes and ß-endorphin and α-MSH peptides following alcohol exposure. CONCLUSIONS: These data suggest that early-life exposure to alcohol alters metabolic sensing to the hypothalamus possibly via regulating Per2 gene and/or the cellular circadian clock mechanism.

Resveratrol decreases inflammation and increases utrophin gene expression in the mdx mouse model of Duchenne muscular dystrophy.

BACKGROUND & AIMS: Duchenne muscular dystrophy (DMD) is a lethal genetic disease with no cure. Reducing inflammation or increasing utrophin expression can alleviate DMD pathology. Resveratrol can reduce inflammation and activate the utrophin promoter. The aims of this study were to identify an active dose of resveratrol in mdx mice and examine if this dose decreased inflammation and increased utrophin expression. METHODS: 5-week old mdx mice were given 0, 10, 100, or 500 mg/kg of resveratrol everyday for 10 days. Sirt1 was measured by qRT-PCR and used to determine the most active dose. Muscle inflammation was measured by H&E staining, CD45 and F4/80 immunohistochemistry. IL-6, TNFα, PGC-1α, and utrophin gene expression were measured by qRT-PCR. Utrophin, Sirt1, and PGC-1α protein were quantified by western blot. RESULTS: The 100 mg/kg dose of resveratrol, the most active dose, increased Sirt1 mRNA 60 ± 10% (p < 0.01), reduced immune cell infiltration 21 ± 6% (H&E) and 42 ± 8% (CD45 immunohistochemistry (p < 0.05)), reduced macrophage infiltration 48 ± 10% (F4/80 immunohistochemistry (p < 0.05)), and increased IL-6, PGC-1α, and utrophin mRNA 247 ± 77%, 27 ± 17%, and 43 ± 23% respectively (p ≤ 0.05). Utrophin, Sirt1, and PGC-1α protein expression did not change. CONCLUSIONS: Resveratrol may be a therapy for DMD by reducing inflammation.

Leucine supplementation increases SIRT1 expression and prevents mitochondrial dysfunction and metabolic disorders in high-fat diet-induced obese mice.

Leucine supplementation has been shown to prevent high-fat diet (HFD)-induced obesity, hyperglycemia, and dyslipidemia in animal models, but the underlying mechanisms are not fully understood. Recent studies suggest that activation of Sirtuin 1 (SIRT1) is an important mechanism to maintain energy and metabolic homeostasis. We therefore examined the involvement of SIRT1 in leucine supplementation-prevented obesity and insulin resistance. To accomplish this goal, male C57BL/6J mice were fed normal diet or HFD, supplemented with or without leucine. After 2 mo of treatment, alterations in SIRT1 expression, insulin signaling, and energy metabolism were analyzed. Eight weeks of HFD induced obesity, fatty liver, mitochondrial dysfunction, hyperglycemia, and insulin resistance in mice. Addition of leucine to HFD correlated with increased expression of SIRT1 and NAMPT (nicotinamide phosphoribosyltransferase) as well as higher intracellular NAD(+) levels, which decreased acetylation of peroxisome proliferator-activated receptor-γ coactivator 1α (PGC1α) and forkhead box O1 (FoxO1). The deacetylation of PGC1α may contribute to upregulation of genes controlling mitochondrial biogenesis and fatty acid oxidation, thereby improving mitochondrial function and preventing HFD-induced obesity in mice. Moreover, decreased acetylation of FoxO1 was accompanied by decreased expression of pseudokinase tribble 3 (TRB3) and reduced the association between TRB3 and Akt, which enhanced insulin sensitivity and improved glucose metabolism. Finally, transfection of dominant negative AMPK prevented activation of SIRT1 signaling in HFD-Leu mice. These data suggest that increased expression of SIRT1 after leucine supplementation may lead to reduced acetylation of PGC1α and FoxO1, which is associated with attenuation of HFD-induced mitochondrial dysfunction, insulin resistance, and obesity.

Calorie restriction-like effects of 30 days of resveratrol supplementation on energy metabolism and metabolic profile in obese humans.

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.