Modulation of Sirtuins to address aging related disorders through the use of selected phytochemicals.

NAMPT regulates senescence, proliferation, and migration of endothelial progenitor cells through the SIRT1 AS lncRNA/miR-22/SIRT1 pathway

Corrigendum to “NAMPT regulates senescence, proliferation, and migration of endothelial progenitor cells through the SIRT1 AS lncRNA/miR-22/SIRT1 pathway”

Melatonin has potent antioxidant, analgesic, and antinociceptive properties. However, the effects of melatonin against oxidative stress-induced cytotoxicity and inflammatory mediators in human chondrocytes remain poorly understood. This study examined the effects and underlying mechanism of melatonin in hydrogen peroxide (H(2) O(2) )-stimulated human chondrocytes and rabbit osteoarthritis (OA) model. Melatonin markedly inhibited hydrogen peroxide (H(2) O(2) )-stimulated cytotoxicity, iNOS, and COX-2 protein and mRNA expression, as well as the downstream products, NO and PGE(2) . Incubation of cells with melatonin decreased H(2) O(2) -induced Sirtuin 1 (SIRT1) mRNA and protein expression. SIRT1 inhibition by sirtinol or Sirt1 siRNA reversed the effects of melatonin on H(2) O(2) -mediated induction of pro-inflammatory cytokines (NO, PGE(2) , TNF-α, IL-1β, and IL-8) and the expression of iNOS, COX-2, and cartilage destruction molecules. Melatonin blocked H(2) O(2) -induced phosphorylation of PI3K/Akt, p38, ERK, JNK, and MAPK, as well as activation of NF-κB, which was reversed by sirtinol and SIRT1 siRNA. In rabbit with OA, intra-articular injection of melatonin significantly reduced cartilage degradation, which was reversed by sirtinol. Taken together, this study shows that melatonin exerts cytoprotective and anti-inflammatory effects in an oxidative stress-stimulated chondrocyte model and rabbit OA model, and that the SIRT1 pathway is strongly involved in this effect.

Carbon monoxide protects against hepatic ischemia/reperfusion injury by modulating the miR-34a/SIRT1 pathway

BackgroundInteractions between environmental factors, such as diet and lifestyle, and metabolic pathways are pivotal in understanding aging mechanisms. hNAA40, Nicotinamide phosphoribosyltransferase (NAMPT), and NAD-dependent protein deacetylase sirtuin-1 (SIRT-1) have been shown to exert important biological processes, including stress response and aging.MethodshNAA40, NAMPT, and SIRT-1 mRNA expression in peripheral blood mononuclear cells (PBMC) were quantitated in 30 lean adult volunteers of normal weight, 30 obese, 20 drug-naïve obese Type 2 diabetes mellitus (T2DM), and 30 obese T2DM on Metformin. Similarly, hNAA40, NAMPT, and SIRT-1 expression in PBMC were quantitated in 36 normal healthy adults randomly assigned to three different groups (Glucose or Whey proteins or lipids; 300 kcal). Blood samples were obtained at 1, 2, and 3 hrs after the macronutrient intake.ResultsThere was an increase in hNAA40 and a decrease in NAMPT and SIRT-1 expression in PBMC from T2DM. Metformin treatment reverted hNAA40, NAMPT, and SIRT-1 expression levels to normal levels. Glucose intake resulted in a significant increase in expression of hNAA40 at 1 hr and decreased significantly at 3 hrs post intake. Lipid intake resulted in an increase in expression of hNAA40 at 2 hr post intake and returned to normal levels at 3 hrs. Neither glucose nor lipid intake resulted in a significant change in NAMPT or SIRT-1 expression. Whey proteins resulted in significantly lower expression of NAMPT at 3 hrs and did not alter the expression levels of SIRT-1 significantly.ConclusionhNAA40, NAMPT, and SIRT-1 pathway could play a role in the determination of the healthy life-span. Metformin modulates this pathway.

β-Nicotinamide mononucleotide activates NAD+/SIRT1 pathway and attenuates inflammatory and oxidative responses in the hippocampus regions of septic mice

Sirtuin 1 (SIRT1) NAD+-dependent deacetylase regulates energy metabolism by modulating expression of genes involved in gluconeogenesis and other liver fasting responses. While many effects of SIRT1 on gene expression are mediated by deacetylation and activation of peroxisome proliferator activated receptor coactivator α (PGC-1α), SIRT1 also binds directly to DNA bound transcription factors, including nuclear receptors (NRs), to modulate their activity. Since thyroid hormone receptor β1 (TRβ1) regulates several SIRT1 target genes in liver and interacts with PGC-1α, we hypothesized that SIRT1 may influence TRβ1. Here, we confirm that SIRT1 cooperates with PGC-1α to enhance response to triiodothyronine, T3. We also find, however, that SIRT1 stimulates TRβ1 activity in a manner that is independent of PGC-1α but requires SIRT1 deacetylase activity. SIRT1 interacts with TRβ1 in vitro, promotes TRβ1 deacetylation in the presence of T3 and enhances ubiquitin-dependent TRβ1 turnover; a common response of NRs to activating ligands. More surprisingly, SIRT1 knockdown only strongly inhibits T3 response of a subset of TRβ1 target genes, including glucose 6 phosphatase (G-6-Pc), and this is associated with blockade of TRβ1 binding to the G-6-Pc promoter. Drugs that target the SIRT1 pathway, resveratrol and nicotinamide, modulate T3 response at dual TRβ1/SIRT1 target genes. We propose that SIRT1 is a gene-specific TRβ1 co-regulator and TRβ1/SIRT1 interactions could play important roles in regulation of liver metabolic response. Our results open possibilities for modulation of subsets of TR target genes with drugs that influence the SIRT1 pathway.

Objective(s):Trans-chalcone as the parent member of the chalcone series reduces circulating levels of insulin and glucose. However, the cellular mechanism of these effects is poorly understood. Sirtuin 1 (SIRT1) as a direct target of miR-34a controls homeostasis of glucose, and also improves insulin sensitivity. Therefore, the present study for the first time investigated the influence of trans-chalcone on the miR-34a/SIRT1 pathway as a possible mechanism for its hypoglycemic and hypoinsulinemic effects.Materials and Methods:In this study, thirty male rats were randomly divided into three groups (n=10): solvent control (NS), oral administration of trans-chalcone for 2 (N2T) and 6 weeks (N6T) groups. Then, hepatic levels of miR-34a and SIRT1 were measured through the qRT-PCR method.Results:Trans-chalcone reduced food intake, body weight gain, and serum glucose as well as insulin levels. Also, this chalcone inhibited hepatic miR-34a expression and significantly elevated SIRT1 mRNA level.Conclusion:Trans-chalcone as an insulin-sensitizing chalcone partly acts through the miR-34a/SIRT1 pathway.

Atherosclerosis is one of the most important causes of cardiovascular disease and studies have showed that adventitial fibroblasts, which are considered to be the most common cell type of the vascular adventitia, are involved in the development of early atherosclerotic plaques. Resveratrol is a plant polyphenolic compound confirmed to have anti-atherosclerotic and cardioprotective effects. The aim of the present study was to investigate the effects of resveratrol on adventitial fibroblasts in vitro and to clarify the underlying mechanism. Adventitial fibroblasts were isolated from the thoracic aorta of 8-week-old SPF Sprague-Dawley rats. Following pre-treatment with different concentrations of resveratrol, cell viability, DNA synthesis ability, cell apoptosis and cell migration ability were assessed in vitro. Through transfection with small interfering (si)RNA targeting sirtuin 1 (SIRT1), the role of the SIRT1 pathway in these processes was evaluated. Western blot analysis was used to assess the protein expression of SIRT1. It was demonstrated that resveratrol inhibited the cell viability, DNA synthesis and migratory ability of the adventitial fibroblasts, and induced cell apoptosis in a concentration-dependent manner in vitro. These effects were partly through the SIRT1 pathways. siRNA targeting SIRT1 successfully reversed the antiproliferative, antimigratory and pro-apoptotic effects of resveratrol on adventitial fibroblasts. In conclusion, the data showed that resveratrol inhibited cell viability, DNA synthesis and cell migration, and induced cell apoptosis in the rat adventitial fibroblasts in vitro through the SIRT1 signaling pathway. As the activation and migration of adventitial fibroblasts contributes to the early development of atherosclerosis, this may be a mechanism underlying the anti-atherosclerotic effect of resveratrol.

Carnosic acid (CA), found in rosemary, has been reported to have antioxidant and antiadipogenic properties. Here, we investigate the molecular mechanism by which CA inhibits hydrogen peroxide (H2O2)-induced injury in HepG2 cells. Cells were pretreated with 2.5-10 μmol/L CA for 2 h and then exposed to 3 mmol/L H2O2 for an additional 4 h. CA dose-dependently increased cell viability and decreased lactate dehydrogenase activities. Pretreatment with CA completely attenuated the inhibited expression of manganese superoxide dismutase (MnSOD) and the B-cell lymphoma-extra large (Bcl-xL), and reduced glutathione activity caused by H2O2, whereas it reversed reactive oxygen species accumulation and the increase in cleaved caspase-3. Importantly, sirtuin 1 (SIRT1), a NAD(+)-dependent deacetylase, was significantly increased by CA. Considering the above results, we hypothesized that SIRT1 may play important roles in the protective effects of CA in injury induced by H2O2. As expected, SIRT1 suppression by Ex527 (6-chloro-2,3,4,9-tetrahydro-1H-carbazole-1-carboxamide) and siRNA-mediated SIRT1 silencing (si-SIRT1) significantly aggravated the H2O2-induced increased level of cleaved caspase-3 but greatly reduced the decreased expression of MnSOD and Bcl-xL. Furthermore, the positive regulatory effect of CA was inhibited by si-SIRT1. Collectively, the present study indicated that CA can alleviate H2O2-induced hepatocyte damage through the SIRT1 pathway.

This study aimed to determine the effects of lidocaine on cell proliferation and cisplatin resistance in A431 human cutaneous squamous cell carcinoma (cSCC) cells and elucidate the underlying mechanism. Cell proliferation, colony numbers, and cisplatin resistance were determined in A431 or cisplatin-resistant A431 (A431-R) cells that were first transfected with miR-30c-inhibitor or miR-30c-mimic, respectively, and then treated with different concentrations of lidocaine, cisplatin, or both. The expression levels of miR-30c and Sirtuin 1 (SIRT1) in A431 and A431-R cells were determined by quantitative real-time polymerase chain reaction and Western blotting. Lidocaine suppressed A431 cell proliferation and cisplatin resistance in a dose- and time-dependent manner via the miR-30c/SIRT1 pathway. MiR-30c overexpression also suppressed cell proliferation and cisplatin resistance in A431 cells by directly targeting and downregulating SIRT1, thus enhancing the protective effects of lidocaine. Conversely, SIRT1 upregulation or miR-30c inhibition antagonized the inhibitory effects of lidocaine. Our results suggest that lidocaine may suppress the progression of cSCC by activating the miR-30c/SIRT1 pathway, indicating its promising potential as a treatment strategy for cSCC.

Nicotinamide adenine dinucleotide (NAD+) is indispensable for the anti-aging activity of the sirtuin (SIRT) family enzymes. AMP-activated protein kinase (AMPK) upregulates NAD+ synthesis and SIRT activity in a nicotinamide phosphoribosyltransferase (NAMPT)-dependent manner. However, the molecular mechanisms that affect AMPK-driven NAMPT expression and NAD+/SIRT activation remain unclear. In this study, we tried to identify senescence-associated microRNAs (miRNAs) that negatively regulate the cascade linking AMPK and NAMPT expression. miRNA-screening experiments showed that the expression of miR-146a increased in senescent cells but decreased following AMPK activation. Additionally, miR-146a overexpression weakened the metformin-mediated upregulation of NAMPT expression, NAD+ synthesis, SIRT activity, and senescence protection, whereas treatment with the miR-146a inhibitor reversed this effect. Importantly, these findings were observed both in vitro and in vivo. Mechanistically, miR-146a directly targeted the 3′-UTR of Nampt mRNA to reduce the expression of NAMPT. AMPK activators metformin and 5-aminoimidazole-4-carboxamide (AICAR) hindered miR-146a expression at the transcriptional level by promoting IκB kinase (IKK) phosphorylation to attenuate nuclear factor-kappaB (NF-κB) activity. These findings identified a novel cascade that negatively regulates the NAD+/SIRT pathway by suppressing miR-146a-mediated NAMPT downregulation. Furthermore, our results showed that miR-146a impedes the anti-aging effect of AMPK. This mutual inhibitory relationship between miR-146a and AMPK enriches our understanding of the molecular connections between AMPK and SIRT and provides new insight into miRNA-mediated NAD+/SIRT regulation and an intervention point for the prevention of aging and age-related diseases.

Sirtuin 1 (SIRT1) plays a critical role in the maintenance of metabolic homeostasis and promotes fat mobilization in white adipose tissue. However, regulation of SIRT1 during adipogenesis, particularly through microRNAs, remains unclear. We observed that miR-146b expression was markedly increased during adipogenesis in 3T3-L1 cells. Differentiation of 3T3-L1 was induced by overexpression of miR-146b. Conversely, inhibition of miR-146b decreased adipocyte differentiation. Bioinformatics-based studies suggested that SIRT1 is a target of miR-146b. Further analysis confirmed that SIRT1 was negatively regulated by miR-146b. We also observed that miR-146b bound directly to the 3′-untranslated region of SIRT1 and inhibited adipogenesis through SIRT1 downregulation. The miR-146b/SIRT1 axis mediates adipogenesis through increased acetylation of forkhead box O1 (FOXO1). Expression of miR-146b was increased and SIRT1 mRNA subsequently decreased in the adipose tissues of diet-induced and genetically obese mice. Furthermore, in vivo knockdown of miR-146b by a locked nucleic acid miR-146b antagomir significantly reduced body weight and fat volume in accordance with upregulation of SIRT1 and subsequent acetylation of FOXO1. Therefore, the miR-146b/SIRT1 pathway could be a potential target for obesity prevention and treatment.

Disc degeneration is the main cause of discogenic low back pain, disc herniation, degenerative stenosis of spinal canal, lumbar spondylolisthesis and other diseases. In the process of intervertebral disc degeneration, water and extracellular matrix of nucleus pulposus tissues are lost, so the normal tension in the intervertebral disc cannot be maintained, which worsens the living environment of nucleus pulposus cells. Low back pain (LBP), with a high incidence rate of disability, has become an increasing health concern and a social and economic problem. The present study aimed to analyze the action mechanisms of nicotinamide phosphoribosyl transferase (Nampt) and sirtuin 1 (SIRT1) in intervertebral disc degeneration (IVDD). In total 26 patients with lumbar disc herniation who had surgical resection at The Third Affiliated Hospital of Jinzhou Medical University were recruited as the experimental group and their degenerative nucleus pulposus (DNP) tissues of intervertebral disc were collected. In addition, nucleus pulposus tissues of intervertebral disc were collected from 20 patients with burst fracture of lumbar spine at the same hospital (control). Nucleus pulposus cells from primary culturing were separated for subsequent experimentation. LC3 II/I, beclin-1, SIRT1 and NAMPT mRNA and protein expression levels were determined using reverse transcription-quantitative PCR and western blotting, respectively. Nicotinamide adenine dinucleotide (NAD) contents in nucleus pulposus cells was determined by NAD assay kit. The mRNA and protein expression levels of SIRT1 in DNP tissues were reduced compared with the control tissues and decreased with increasing disease severity. The expression of autophagy-associated LC3 II/I and beclin-1 in DNP tissues was reduced compared with control tissues. SIRT1 regulated the LC3 II/I and beclin-1 expression levels in nucleus pulposus cells. Treatment with resveratrol and inhibitor of SIRT1 showed that Nampt/NAD+/SIRT1 pathway participated in the process of IVDD by regulating autophagy of nucleus pulposus cells. SIRT1 serves a role in the process of IVDD through Nampt/NAD+/SIRT1 pathway that regulates autophagy of nucleus pulposus cells. SIRT1 may become a biological target for the treatment of IVDD.

ObjectiveHepatic insulin resistance is a major initiating factor for type 2 diabetes mellitus. In previous study, Gegen Qinlian Decoction containing berberine could enhance hepatic insulin sensitivity by SIRT1-dependent deacetylation of FOXO1. However, it is not clear whether berberine also can improve hepatic insulin sensitivity by SIRT1/FOXO1 pathway. This study aimed to evaluate the efficacy of berberine for improving hepatic insulin resistance and the possible molecular mechanisms involved.MethodsIn vitro, HepG2 cells were induced with palmitic acid, and glycogen synthesis was examined. In vivo, a high-fat diet (HFD)-fed mouse model was established, and metabolic parameters were assessed. The expressions of miR-146b and sirtuin 1 (SIRT1) in liver were also examined. The relationship between miR-146b and SIRT1 was examined by the dual-luciferase reporter gene assay.ResultsSerum biochemical parameters, such as glucose and HOMA-IR index, were increased in HFD mice; miR-146b and SIRT1 were abnormally expressed in HFD mice and palmitic acid-treated HepG2 cells. Interestingly, berberine reduced body weight and caused a significant improvement in glucose tolerance and HOMA-IR index without altering food intake in mice. Overexpression of miR-146b abolished the protective effect of berberine on palmitic acid-induced impaired glycogen synthesis in HepG2 cells. Luciferase assay showed that miR-146b directly targeted SIRT1.ConclusionThe present findings suggest that berberine could attenuate hepatic insulin resistance through the miR-146b/SIRT1 pathway, which may represent a potential therapeutic target for the prevention and treatment of metabolic diseases, particularly diabetes.