Silent Information Regulator T1 Research Articles

SIRT1 maintains bone homeostasis by regulating osteoblast glycolysis through GOT1.

The silent information regulator T1 (SIRT1) is linked to longevity and is a crucial mediator of osteoblast function. We investigated the direct role of Sirt1 during bone modeling and remodeling stages in vivo using Tamoxifen-inducible osteoblast-specific Sirt1 conditional knockout (cKO) mice. cKO mice exhibited lower trabecular and cortical bone mass in the distal femur. These phenotypes were coupled with lower bone formation and bone resorption. Metabolomics analysis revealed that the metabolites involved in glycolysis were significantly decreased in cKO mice. Further analysis of the quantitative acetylome revealed 11 proteins with upregulated acetylation levels in both the femur and calvaria of cKO mice. Cross-analysis identified four proteins with the same upregulated lysine acetylation site in both the femur and calvaria of cKO mice. A combined analysis of the metabolome and acetylome, as well as immunoprecipitation, gene knockout, and site-mutation experiments, revealed that Sirt1 deletion inhibited glycolysis by directly binding to and increasing the acetylation level of Glutamine oxaloacetic transaminase 1 (GOT1). In conclusion, our study suggested that Sirt1 played a crucial role in regulating osteoblast metabolism to maintain bone homeostasis through its deacetylase activity on GOT1. These findings provided a novel insight into the potential targeting of osteoblast metabolism for the treatment of bone-related diseases.

Interference in the nutrient-sensing and inflammatory signaling pathways by renal autophagy activation in mice with late stage diabetic nephropathy.

Disturbance in metabolism and inflammation are the main causes of kidney injury in patients with late stage diabetic nephropathy (DN). Here, we explored whether autophagy was activated in mice with late stage DN and whether it was associated with disturbance in metabolism and inflammation. In total, mice were divided into the control group (db/m) and DN group (db/db). Mice were raised for 7months, and their biochemical indices were measured. Subsequently, their kidneys were collected to detect autophagy and the related nutrient-sensing and inflammatory signaling pathways in late stage DN. The expression levels of autophagy markers LC3-I and LC3-II were significantly increased in mice with late stage DN, whereas that of autophagy flux marker P62 was significantly decreased, indicating activation of autophagy. Concurrently, mechanistic target of rapamycin was highly expressed as a cellular nutrient-sensing and energy regulator in mice with late stage DN. Additionally, the expression levels of markers of nutrient-sensing signaling pathways adenosine monophosphate-activated protein kinase (AMPK) were increased markedly in mice with late stage DN. Additionally, the expression levels of the marker of nutrient-sensing signaling pathways silent information regulator T1 (SIRT1), the marker of inflammatory signaling pathways high mobility group box protein 1 (HMGB1), and interferon regulatory factor 3 (IRF3) were significantly increased in mice with late stage DN. The findings of our study indicate that autophagy activation in late stage DN may interfere with nutrient-sensing and inflammatory signaling pathways involving AMPK, SIRT1, HMGB1, and IRF3.

Thermo-TRP channels are involved in BAT thermoregulation in cold-acclimated Brandt's voles.

Transient receptor potential (TRP) channels, which can sense temperature, pressure and mechanical stimuli, were involved in many physiological and biochemical reactions. Whether thermosensitive TRP channels (Thermo-TRPs) are involved in thermoregulation in small mammals is still not clear. We measured the changes of thermo-TRPs at 4°C, 23°C and 30°C in Brandt's voles (Lasiopodomys brandtii) to test the hypothesis that Thermo-TRPs are involved in cold-induced thermogenesis of brown adipose tissue (BAT) in small mammals. Results showed that air temperatures had no effect on body mass and rectal temperature, but the food intake and basal metabolic rate (BMR) in the 4°C group were significantly higher than in the 30°C group. Compared with 30°C group, the protein contents of uncoupling protein 1(UCP1), TRP vanilloid 2 (TRPV2), TRP ankyrin 1 (TRPA1), TRP melastatin 2 (TRPM2), silent Information Regulator T1 (SIRT1), AMP-activated protein kinase (AMPK) and Calcium/calmodulin-dependent protein kinase II (CaMKII) in BAT increased significantly in 4°C group, but there was no significant difference in the protein content of Thermo-TRPs in the hypothalamus among groups. Further, the expression of PRDM16 (PR domain containing 16) in inguinal white adipose tissue (iWAT) at 4°C was significantly higher than that at 30°C, but no difference was observed in the expression of other browning-related genes or TRPV2. In conclusion, TRP channels may participate in BAT thermoregulation through the CaMKII, AMPK, SIRT1 and UCP1 pathway in cold-acclimated Brandt's voles.

Translocator Protein (TSPO) Alleviates Neuropathic Pain by Activating Spinal Autophagy and Nuclear SIRT1/PGC-1α Signaling in a Rat L5 SNL Model.

PurposeRecent studies showed promotion of astrocyte autophagy in the spinal cord would provide analgesic effects. Silent information regulator T1 (SIRT1) and α subunit of peroxisome proliferator-activated receptor-γ coactivator-1 (PGC-1α) are two master regulators of endogenous antioxidant defense and mitochondrial biogenesis. They play vital roles in both autophagy and neuropathic pain (NP). Our previous study showed that TSPO agonist Ro5-4864 elicited potent analgesic effects against NP, but the mechanisms remain unclear. This study aims to investigate the effects of TSPO agonist Ro5-4864 on autophagy and nuclear SIRT1/PGC-1α signaling in spinal dorsal horn.MethodsA rat model of L5 spinal nerve ligation (SNL) was adopted. Rats were randomly assigned to the Sham group, the SNL group, the Ro (TSPO agonist Ro5-4864) group and the Ro+3-MA group. The behavior assessments were conducted at baseline, on Day 1, 3, 7 and 14 after SNL. The autophagy-related proteins (ATG7, Beclin1, LC3, and P62) in spinal dorsal horn were assayed and the nuclear SIRT1/PGC-1α and downstream factors were analyzed.ResultsRo5-4864 alleviated the mechanical allodynia induced by SNL (P < 0.01 vs the SNL group), which could be totally abrogated by autophagy inhibitor 3-MA (P < 0.01 vs the Ro group). SNL induced elevated ATG7 (P < 0.01), Beclin1 (P < 0.01) and LC3-II/LC3-I (P < 0.01) contents and P62 accumulation (P < 0.01) on Day 7 and Day 14, which suggested an autophagy flux impairment. Ro5-4864 augmented ATG7 (P < 0.01), Beclin1 (P < 0.01) and LC3-II/LC3-I (P < 0.05) with decreased P62 (P < 0.01), which indicated a more fluent autophagic process. These effects were also totally abrogated by 3-MA (P < 0.01). Furthermore, Ro5-4864 activated the spinal nuclear SIRT1/PGC-1α signaling pathway.ConclusionTSPO improved both autophagy impairment and mitochondrial biogenesis, which may provide a new strategy for the treatment of NP.

Fucoxanthin regulates Nrf2 signaling to decrease oxidative stress and improves renal fibrosis depending on Sirt1 in HG-induced GMCs and STZ-induced diabetic rats

Diabetic nephropathy (DN) is one of the major microvascular complications of diabetes. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a crucial cellular defense factor to cope with oxidative stress. Silent information regulator T1 (Sirt1) is a deacetylase with antioxidative stress activity. Fucoxanthin is a marine-derived carotenoid. This study was conducted to investigate whether fucoxanthin could alleviate oxidative stress by activating Sirt1/Nrf2 signaling to alleviate DN. In streptozotocin-induced diabetic rats, fucoxanthin treatment effectively improved renal function, alleviated glomerulosclerosis. Fucoxanthin reversed the decreased protein levels of Sirt1 and Nrf2 in the kidney of diabetic rats and glomerular mesangial cells cultured in high glucose. Conversely, EX527, a Sirt1 inhibitor, counteracted the effect of fucoxanthin on the expression of Nrf2. Furthermore, in vivo and vitro results showed that fucoxanthin treatment reversed the low expression and activity of superoxide dismutase and heme oxygenase 1, depending on Sirt1 activation. Our results suggest that fucoxanthin improves diabetic kidney function and renal fibrosis by activating Sirt1/Nrf2 signaling to reduce oxidative stress.

The hepatic AMPK-TET1-SIRT1 axis regulates glucose homeostasis.

Ten-eleven translocation methylcytosine dioxygenase 1 (TET1) is involved in multiple biological functions in cell development, differentiation, and transcriptional regulation. Tet1 deficient mice display the defects of murine glucose metabolism. However, the role of TET1 in metabolic homeostasis keeps unknown. Here, our finding demonstrates that hepatic TET1 physically interacts with silent information regulator T1 (SIRT1) via its C-terminal and activates its deacetylase activity, further regulating the acetylation-dependent cellular translocalization of transcriptional factors PGC-1α and FOXO1, resulting in the activation of hepatic gluconeogenic gene expression that includes PPARGC1A, G6PC, and SLC2A4. Importantly, the hepatic gluconeogenic gene activation program induced by fasting is inhibited in Tet1 heterozygous mice livers. The adenosine 5'-monophosphate-activated protein kinase (AMPK) activators metformin or AICAR-two compounds that mimic fasting-elevate hepatic gluconeogenic gene expression dependent on in turn activation of the AMPK-TET1-SIRT1 axis. Collectively, our study identifies TET1 as a SIRT1 coactivator and demonstrates that the AMPK-TET1-SIRT1 axis represents a potential mechanism or therapeutic target for glucose metabolism or metabolic diseases.

Coffeeberry Activates the CaMKII/CREB/BDNF Pathway, Normalizes Autophagy and Apoptosis Signaling in Nonalcoholic Fatty Liver Rodent Model.

Nonalcoholic fatty liver disease (NAFLD) shows extensive liver cell destruction with lipid accumulation, which is frequently accompanied by metabolic comorbidities and increases mortality. This study aimed to investigate the effects of coffeeberry (CB) on regulating the redox status, the CaMKII/CREB/BDNF pathway, autophagy, and apoptosis signaling by a NAFLD rodent model senescence-accelerated mice prone 8 (SAMP8). Three-month-old male SAMP8 mice were divided into a control group and three CB groups (50, 100, and 200 mg/kg BW), and fed for 12 weeks. The results show that CB reduced hepatic malondialdehyde and carbonyl protein levels. CB significantly enhanced Ca2+/calmodulin-dependent protein kinase II (CaMKII) and brain-derived neurotrophic factor (BDNF) and reduced the phospho-cAMP response element-binding protein (p-CREB)/CREB ratio. In addition, CB increased the silent information regulator T1 level, promoted Beclin 1 and microtubule-associated protein light chain 3 II expressions, and reduced phosphorylated mammalian target of rapamycin and its downstream p-p70s6k levels. CB also inhibited the expressions of apoptosis-related factors poly (ADP-ribose) polymerase-1 and the apoptosis-inducing factor. In conclusion, CB might protect the liver by reducing oxidative stress, activating the CaMKII/CREB/BDNF pathway, and improving autophagic and apoptotic expressions in a dose-dependent manner.

Silent Information Regulator T1 in Aqueous Humor of Patients with Age-Related Macular Degeneration

Purpose: The purpose of this study is to compare the aqueous humor level of Silent Information Regulator T1 (SIRT1) between patients with Age-related Macular Degeneration (AMD) and cataract patients. Materials and Methods: Aqueous humor level of SIRT1 was measured by enzyme-linked immunosorbent assay in 13 patients with wet-type AMD (n=13, AMD group) and 13 patients with cataracts (cataract group). In addition, the thickness of each retinal layer was determined by optical coherence tomography. Results: The aqueous humor level of SIRT1 was significantly lower in the AMD group than in the cataract group (p=0.007). In the AMD group, the SIRT1 level was positively correlated with the thickness of the retinal ganglion cell layer (r=0.31) and the inner nuclear layer (r=0.76). Conclusion: The aqueous level of SIRT1 decreased as the ganglion cell layer and inner nuclear layer became thinner, suggesting that reduction of SIRT1 activity might be involved in the pathogenesis of this disease.

Evaluation of silent information regulator T (SIRT) 1 and Forkhead Box O (FOXO) transcription factor 1 and 3a genes in glaucoma.

Analysis of the reactive oxygen species (ROS)-detoxifying biomarkers may elucidate the mitochondrial dysfunction in glaucoma pathogenesis. Therefore, we purposed to investigate the effects of ROS-detoxifying molecules including Silent Information Regulator T1 (SIRT1) and Forkhead Box O 1 (FOXO1) and 3a (FOXO3a) transcription factors in patients with glaucoma. Our analyses included 20 eyes from patients with primary open-angle glaucoma (POAG) and 20 eyes from patients with pseudoexfoliation glaucoma (PXG) who were scheduled for trabeculectomy. After extraction of total RNA from trabecular meshwork tissue, we compared the levels of SIRT1, FOXO1and FOXO3a genes in the oxidative pathway with the level of glyceraldehyde-3 phosphate dehydrogenase (GAPDH), the reference gene, using real-time polymerase chain reaction. Relative gene expression was calculated using the threshold cycle (2-ΔΔCT) method. We observed similarly reduced expression levels of SIRT1, FOXO1, and FOXO3a genes versus GAPDH among patient groups (p = 0.40; p = 0.56; p = 0.35, respectively). This is the first study to identify the role of SIRT1 and FOXOs in human TM with glaucoma. Relative expression levels of SIRT1, FOXO1, and FOXO3a genes versus a control gene (GAPDH) were decreased in POAG and PXG groups. Our results show that SIRT1and FOXOs (1-3a) deserve special attention in the pathogenesis of glaucoma.

MicroRNA-34a Inhibition Alleviates Lung Injury in Cecal Ligation and Puncture Induced Septic Mice.

Sepsis is the leading cause of death in intensive care units. MicroRNA-34a (miR-34a) is involved in sepsis progression, while its underlying mechanisms on sepsis-induced lung injury remain obscure. Oxidative stress, pyroptosis, and inhibition of autophagy can result in organ injury. MiR-34a has been reported to regulate oxidative stress and autophagy via inhibiting silent information regulator T1 (SIRT1) and autophagy gene 4B (ATG4B) signaling. This study aimed at identifying the function of miR-34a in oxidative stress, inflammation, pyroptosis, and autophagy in sepsis-induced lung injury. Male 8-week-old C57BL/6 mice were subjected to cecal ligation and puncture and treated with miR-34a antagomir/agomir. Survival (n = 10), histopathological changes (n = 6), and lung wet-to-dry ratio (n = 6) were recorded and assayed. Other detection (n = 6) was performed to investigate the level of oxidative stress, inflammation, pyroptosis, and autophagy in lung tissues. Results showed that miR-34a down-regulation ameliorated lung injury in septic mice as reflected by decreased lung injury scores (decrease from 3.00 ± 0.32 to 2.00 ± 0.32) and wet-to-dry ratio (0.36-fold decrease). MiR-34a down-regulation also decreased reactive oxygen species accumulation (0.36-fold decrease), and promoted superoxide dismutase activity and the expression of SIRT1 (1.24-fold increase), heme oxygenase-1 and nuclear factor erythroid 2 like 2 to inhibit oxidative stress in septic mice. Moreover, miR-34a down-regulation suppressed inflammatory response and pyroptosis in septic mice, as evidenced by decreased level of pro-inflammatory factors including tumor necrosis factor α, interleukin-6 (IL-6), IL-1β, and IL-18, activity of caspase-1 (0.51-fold decrease) and expression of nucleotide-binding domain and leucine-rich repeat protein-3 (0.48-fold decrease), apoptosis-associated speck-like protein containing a CARD, cleaved-caspase-1, and cleaved-gasdermin D (0.36-fold decrease), and increased level of anti-inflammatory factors IL-10. MiR-34a down-regulation also enhanced autophagy in septic mice as evidenced by more autolysosomes and elevated expressions of ATG4B (0.90-fold increase), beclin1, ATG9, and LC3 II/I. Among these experiments, miR-34a up-regulation showed opposite effects on oxidative stress, inflammatory response, pyroptosis, and autophagy in septic mice. Additionally, miR-34a could bind to the 3′-untranslated region of SIRT1 and ATG4B. In conclusion, our findings demonstrated that miR-34a was implicated in oxidative stress, inflammation, pyroptosis, and autophagy in the development of sepsis. MiR-34a inhibition had a potential to alleviate sepsis-induced lung injury.

Pathogenesis of Photoaging in Human Dermal Fibroblasts

Pathogenesis of Photoaging in Human Dermal Fibroblasts

Cordyceps cicadae Mycelia Ameliorate Cisplatin-Induced Acute Kidney Injury by Suppressing the TLR4/NF-κB/MAPK and Activating the HO-1/Nrf2 and Sirt-1/AMPK Pathways in Mice.

Acute kidney injury (AKI) is a common clinical problem, characterized by a sudden loss of renal function, a high risk of death, and the eventual development of renal fibrosis and renal failure. Cordyceps cicadae is a traditional Chinese medicine with the potential function of kidney protection. We analyze two sputum extracts, a water extract (WCC), and an ethanol extract (ECC), to assess the potential of treating AKI in an animal model of kidney injury induced by cisplatin. A nephrotoxic mouse model was first established by intraperitoneal injection of cisplatin. Subsequently, WCC and ECC were orally administered in these mice. The results show that WCC and ECC significantly alleviated cisplatin-induced AKI renal histological changes, serum creatinine (CRE) and blood urea nitrogen (BUN) production, and the levels of NO, TNF-α, IL-1β, and IL-6. The levels of malondialdehyde (MDA) and glutathione (GSH) were suppressed by administration of WCC and ECC. However, WCC treatment prevented these changes significantly better than ECC treatment. In addition, Western blot data showed that WCC attenuated the cisplatin-induced protein expression of cyclooxygenase-2 (COX-2) and inducible NO synthase (iNOS), as well as inhibiting nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) activation in the kidney tissues. Furthermore, WCC greatly inhibited the expression of Toll-like receptor 4 (TLR4) and cisplatin-induced NF-κB activation, as well as dramatically increasing the production of antioxidative enzymes (i.e., superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase, nuclear factor erythroid 2-related factor 2 (Nrf2), and heme oxygenase 1 (HO-1)), silent information regulator T1 (Sirt1), and p-AMP-activated protein kinase (AMPK) in the kidney tissues. In addition, we found that WCC increased the expression levels of the autophagy-related proteins LC3B and Beclin-1; proapoptotic proteins, including cleaved caspase-3 and cleaved poly (ADP-ribose) polymerase (PARP) 1; and organic anion transporters 1 (OAT1) and 3 (OAT3) in the kidney tissues. Finally, WCC, ECC, and two bioactive compounds—adenosine and N6-(2-hydroxyethyl) adenosine (HEA)—inhibited the production of nitrite oxide (NO) and intracellular reactive oxygen species (ROS) triggered by lipopolysaccharide- (LPS-) stimulated RAW264.7 macrophages in vitro. Collectively, WCC could provide a potential therapeutic candidate for the prevention of cisplatin-induced kidney injury through the inhibition of oxidative stress and inflammation.

Ibrutinib in Combination with Venetoclax Decreases Mitochondrial Bioenergetics through the Impaired BTK, AKT and AMPK/SIRT/PGC-1α Signaling Pathway in CLL

Objective: Chronic lymphocytic leukemia (CLL) is one of the most common types of leukemia in adults. Altered mitochondrial metabolism has been shown to be involved in the pathogenesis of CLL. Ibrutinib, an inhibitor of Bruton's tyrosine kinase (BTK), and venetoclax, an inhibitor of B-cell lymphoma 2 (Bcl-2) protein are approved therapies for CLL. The adenosine monophosphate-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) signaling axis senses the metabolic demands of cells and regulates mitochondrial function. Silent information regulator T1 (SIRT1) is a cytoplasmic enzyme that mediates NAD+-dependent deacetylation of target substrates. The importance of BTK, AKT, and AMPK/SIRT/PGC-1α signaling pathway have to be explored yet in CLL. Ibrutinib and venetoclax may trigger a switching off of AMPK and /or SIRT1 signaling leading to impaired PGC-1α expression/activity and diminished mitochondrial activity. The effects of these drugs on mitochondrial bioenergetics profile, cell viability, mitochondrial membrane potential (MMP), reactive oxygen species (ROS) and protein levels of p-BTK, p-AKT, Bcl-2, p-AMPK, SIRT1, and PGC-1α were analyzed. We hypothesize that ibrutinib, along with venetoclax, will alter CLL mitochondrial bioenergetics profiles and function, cell viability, and ROS levels through the involvement of BTK, AKT and AMPK/SIRT/PGC-1α signaling pathway and enable lower doses and profound effects for combination therapies rather than single agent therapy. Methods: The high resolution Oroboros Oxygraph 2K (Oroboros Instruments, Austria), a Clarke-type oxygen electrode, is used to measure mitochondrial respiration rates of CLL cells at 370C. Freshly isolated CLL cells (10 mil. cells/ml) were added to the chamber in 2 ml of RPMI. After the measurement of basal respiration rates, the following chemicals were added: oligomycin (2uM), FCCP: carbonylcyanide p-trifluoromethoxyphenylhydrazone (2-12 uM), and antimycin A (2 uM). Oxygen consumption rate is expressed in pmol oxygen/s /mil. cells. Protein levels in CLL cell lysates were determined by Western blotting. Cell viability, MMP, and ROS were assessed by the flow cytometer (NovoCyte). Results: CLL cells obtained ex vivo from ibrutinib treated patients had decreased mitochondrial bioenergetics compared to pre-treatment samples. Primary CLL cells treated in vitro with sub-lethal doses of ibrutinib for 24 or 48h showed significantly decreased basal respiration rates, maximal respiration rates, and spare respiratory capacity compared to DMSO vehicle control. These parameters were also significantly affected by low doses (0.5 - 2 nM) of venetoclax. The combination treatment of ibrutinib with venetoclax for 24 or 48h significantly decreased these bioenergetics parameters compared to each individual drug alone except for the spare respiratory capacity vs venetoclax alone at 24 h. MMP was significantly decreased by these drug treatments and its combinations compared to DMSO. Cell viability was used to confirm a sub-lethal phenotype. ROS levels were significantly increased by venetoclax compared to DMSO, and in combination with ibrutinib vs ibrutinib alone. However, ROS levels by ibrutinib were not significantly changed compared to DMSO or venetoclax. Protein levels of p-BTK, p-AKT, p-AMPK, SIRT1, and PGC-1α were decreased by ibrutinib or venetoclax alone compared to DMSO and further decreased by the combination of these two drugs compared to single agent. Conclusion: CLL cells from ibrutinib treated patients, a clinically approved BTK inhibitor, demonstrated decreased bioenergetics similar to normal B-lymphocytes suggesting ibrutinib treatment normalizes the mitochondrial bioenergetics in CLL. Ibrutinib and venetoclax affected bioenergetics profiles in CLL cells at low doses. The combined effect of these drugs on the mitochondrial bioenergetics profiles and cell viability is more profound than each BTK or Bcl-2 inhibitor agent alone. Protein levels of p-BTK, p-AKT, p-AMPK, SIRT1, and PGC-1α in ibrutinib and venetoclax treated samples were reduced compared to DMSO and each single agent inhibitor. These novel data suggest the involvement of BTK, AKT and AMPK/SIRT/PGC-1α signaling pathway to target mitochondrial metabolism and provide rational therapeutic combinations that may lead to reduced toxicity and increased drug efficacy in CLL. Disclosures Johnston: Janssen: Research Funding. Banerji:Janssen: Consultancy, Honoraria, Research Funding; Roche: Honoraria, Licensing fee, Research Funding; Abbvie: Consultancy, Honoraria; CIHR: Research Funding; LLSC: Research Funding; Research Manitoba: Research Funding; Astra-Zeneca: Consultancy, Honoraria; Gilead: Consultancy, Honoraria, Research Funding; Dana-Farber Cancer Institute: Other: Licencing fee; CCMF: Research Funding; CancerCare Manitoba/University of Manitoba: Employment; CAPhO: Honoraria; BIOGEN: Other: Licensing fee.

SIRT1 Modulators in Experimentally Induced Liver Injury

This article is directed at highlighting the involvement of the endogenous stress sensor SIRT1 (silent information regulator T1) as a possible factor involved in hepatoprotection. The selective SIRT1 modulators whether activators (STACs) or inhibitors are being tried experimentally and clinically. We discuss the modulation of SIRT1 on cytoprotection or even cytotoxicity in the liver chemically injured by hepatotoxic agents in rats, to shed light on the crosstalk between SIRT1 and its modulators. A combination of D-galactosamine and lipopolysaccharide (D-GalN/LPS) downregulated SIRT1 expression, while SIRT1 activators, SRT1720, resveratrol, and quercetin, upregulated SIRT1 and alleviated D-GalN/LPS-induced acute hepatotoxicity. Liver injury markers exhibited an inverse relationship with SIRT1 expression. However, under subchronic hepatotoxicity, quercetin decreased the significant increase in SIRT1 expression to lower levels which are still higher than normal ones and mitigated the liver-damaging effects of carbon tetrachloride. Each of these STACs was hepatoprotective and returned the conventional antioxidant enzymes to the baseline. Polyphenols tend to fine-tune SIRT1 expression towards normal in the liver of intoxicated rats in both acute and subchronic studies. Together, all these events give an impression that the cytoprotective effects of SIRT1 are exhibited within a definite range of expression. The catalytic activity of SIRT1 is important in the hepatoprotective effects of polyphenols where SIRT1 inhibitors block and the allosteric SIRT1 activators mimic the hepatoprotective effects of polyphenols. Our findings indicate that the pharmacologic modulation of SIRT1 could represent both an important move in alleviating hepatic insults and a future major step in the treatment of xenobiotic-induced hepatotoxicity.

Long noncoding RNA SNHG7 inhibits high glucose-induced human retinal endothelial cells angiogenesis by regulating miR-543/SIRT1 axis

Diabetic retinopathy (DR) is the serious complication of type 2 diabetes mellitus, which could lead to visual impairment. Growing evidence have revealed the involvement of long non-coding RNAs (lncRNAs) in the pathogenesis of DR. Thus, this study was performed to investigate the role of lncRNA SNHG7 (small nucleolar RNA host gene 7) in high glucose (HG)-induced proliferation, migration, and angiogenesis of human retinal endothelial cells (hRECs). We discovered that SNHG7 was decreased in hRECs under HG stimuli. Although SNHG7 had no influence on cell viability, migration and angiogenesis under condition, overexpression of SNHG7 inhibited the HG-induced cell proliferation, migration and angiogenesis, as well as vascular endothelial growth factor (VEGF) expression in HG condition. In terms of mechanism, we found that SNHG7 directly inhibited miR-543, which targeted the 3′-UTR of Silent information regulator T1 (SIRT1) mRNA and subsequently downregulated the VEGF expression in hRECs. Ultimately, upregulation of miR-543 or inhibition of SIRT1 both abrogated the effect of SNHG7 on HG-induced angiogenesis. Collectively, our results suggested that SNHG7 is a potential molecular target for attenuating HG-induced angiogenesis in the DR through regulation of the miR-543-mediated SIRT1/VEGF pathway.

Effects of Preventive Treadmill Exercise on the Recovery of Metabolic and Mitochondrial Factors in the 6-Hydroxydopamine Rat Model of Parkinson's Disease.

Metabolic and mitochondrial dysfunction has been implicated in Parkinson's disease, while exercise can induce essential pathways of mitochondrial biogenesis. Here, we tested whether long-term preventive treadmill training (16weeks, 21m/min, and 0° inclinations for 50min/d, 5 d/week) effects the mitochondrial and neurodegeneration markers, in the striatum of rats in the 6-hydroxydopamine (6-OHDA) model of Parkinson's disease. Following 16weeks of exercise or no exercise period (n = 16 rats per group), the animals were divided into four experimental groups (n = 8 per group): (1) no exercise and saline (SED), (2) exercise and saline (EX), (3) no exercise and 6-OHDA (SED + 6-OHDA), and (4) exercise and 6-OHDA (EX + 6-OHDA). For the model, 8μg of 6-OHDA (2μg/μL prepared in a solution of 0.2% ascorbic acid and 0.9% saline) was injected into the right medial forebrain bundle. Exposure to 6-OHDA resulted in a significant reduction (P < 0.05) of mitochondrial factors AMP-activated protein kinase, peroxisome proliferator-activated receptor gamma coactivator-1 alpha, and tyrosine hydroxylase, and increased expression of silent information regulator T1, mitochondrial transcription factor A, and p53 in the SED + 6-OHDA group relative to SED group. By contrast, gene and protein expressions upon exercise were higher and p53 protein level was lower in the EX + 6-OHDA group compared with SED + 6-OHDA. Further, exercise reduced the extent of weight loss associated with the 6-OHDA injection. In conclusion, exercise might be used to reduce mitochondrial disorders in Parkinson's disease.

Venetoclax in Combination with NAMPT Inhibitors Decreases Mitochondrial Bioenergetics through the Impaired AMPK/SIRT/PGC1α Signaling Pathway in CLL

Objective: Chronic lymphocytic leukemia (CLL) is one of the most common types of leukemia in adults. Altered mitochondrial metabolism has been shown to be involved in the pathogenesis of CLL. Nicotinamidephospho-ribosyltransferase (NAMPT) is a key enzyme in the nicotinamide adenine dinucleotide (NAD) salvage pathway. FK866 and GMX1778, chemical inhibitors of NAMPT, deplete cellular NAD and ATP levels and trigger apoptosis in CLL cells, suggesting NAMPT contributes to the prolonged survival of these cells. Venetoclax is an approved therapy for CLL and blocks the anti-apoptotic B-cell lymphoma-2 (Bcl-2) protein, leading to programmed cell death of CLL cells. The adenosine monophosphate-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) signaling axis senses the metabolic demands of cells and regulates mitochondrial function. Silent information regulator T1 (SIRT1) is a cytoplasmic enzyme that mediates NAD+-dependent deacetylation of target substrates. The importance of the AMPK/SIRT/PGC1α signaling pathway has to be explored yet in CLL. Venetoclax may trigger a switching off of AMPK and /or SIRT1 signaling leading to impaired PGC-1α expression/activity and diminished mitochondrial activity. The effects of these drugs on mitochondrial bioenergetics profile, cell viability, mitochondrial membrane potential (MMP), reactive oxygen species (ROS) and protein levels of Bcl-2, AMPK, SIRT1, and PGC1α were analyzed. We hypothesize that venetoclax, along with FK866 and GMX1778, will alter CLL mitochondrial bioenergetics profiles and function, cell viability, and ROS levels through the involvement of the AMPK/SIRT/PGC1α signaling pathway and enable lower doses and profound effects for combination therapies rather than single agent therapy.Methods: A high resolution Oroboros Oxygraph 2K (Oroboros Instruments, Innsbruck, Austria), a Clarke-type oxygen electrode is used to measure mitochondrial respiration rates of CLL cells at 370C. Freshly isolated CLL cells (10 mil./ml) were added to the chamber in 2 ml of RPMI. After the measurement of basal respiration rates, the following chemicals were added: oligomycin (2uM), FCCP: carbonyl cyanide p-trifluoromethoxy phenyhydrazone (2.5-12.5 uM), and antimycin A (2 uM). Oxygen consumption rate is expressed in pmol oxygen/s/mil. cells. Protein levels in CLL cell lysates were determined by Western blotting. Cell viability, MMP, and ROS were assessed by Novocyte flow cytometer.Results: Primary CLL cells treated with 1.25 nM of venetoclax for 24 or 48h showed significantly decreased basal respiration rates, maximal respiration rates, and spare respiratory capacity compared to DMSO vehicle control. These parameters were also significantly affected by 1nM of FK866 or 1 nM of GMX1778 except basal respiration rates. The combination treatment of venetoclax with FK866 or GMX1778 for 24 or 48h significantly decreased all these bioenergetics parameters compared to each individual drug. MMP was not affected by these drug treatments and cell viability corresponded to the effect of the mitochondrial bioenergetics profile. ROS levels after 24 hours in venetoclax treated CLL cells were significantly increased compared to DMSO, any single agent or in combination. However, after 48 hours of treatment, increased ROS levels were not statistically significant compared to DMSO. Protein levels of Bcl-2, P-AMPK, AMPK, SIRT1, and PGC1α were decreased by venetoclax alone or in combination with FK866 or GMX1778.Conclusion: Venetoclax, FK866 and GMX1778 affected bioenergetics profiles in CLL cells at doses as low as 1.25 nM, 1nM, and 1 nM, respectively. The combined effect of these drugs on the mitochondrial bioenergetics profiles and cell viability is more profound than each NAMPT inhibitor agent alone. Protein levels of Bcl-2, AMPK, SIRT1, and PGC1α in venetoclax treated samples were reduced compared to DMSO and each single agent NAMPT inhibitor. These novel data suggest the involvement of the AMPK/SIRT/PGC1α signaling pathway to target mitochondrial metabolism and provide rational therapeutic combinations that may lead to reduced toxicity and increased drug efficacy in CLL. DisclosuresBanerji:Roche: Other: Unrestricted grant received in the past; Janssen: Other: Unrestricted grant received in the past; Gilead: Other: Unrestricted grant received in the past; Abbvie: Other: Unrestricted grant received in the past; Teva: Other: Unrestricted grant received in the past.

The protective effect of resveratrol on vascular aging by modulation of the renin–angiotensin system

Background and aimsThis study evaluated the effects of resveratrol on arterial aging and the renin–angiotensin system (RAS) in mice and vascular smooth muscle cells (VSMCs). MethodsAging mice were divided into control and resveratrol groups. Histological changes, inflammation, oxidative stress, RAS components, and the expression of AMP-activated protein kinase (AMPK), silent information regulator T1 (SIRT1), peroxisome proliferator-activated receptor-γ co-activator 1α (PGC-1α), and anti-oxidative enzymes was measured in thoracic aortas of 24-month-old mice. The effect of resveratrol on fibrosis, cell senescence, and RAS components was also investigated in VSMCs stimulated by angiotensin (Ang) II. ResultsAorta media thickness, inflammation, fibrosis, and oxidative stress were significantly lower in the resveratrol group than in the control group. Resveratrol treatment decreased serum Ang II level and the aortic expression of prorenin receptor (PRR) and angiotensin converting enzyme (ACE), and increased serum Ang-(1–7) level and the expression of ACE2, Ang II type 2 receptor (AT2R), and Mas receptor (MasR). Resveratrol increased the expression of phosphorylated AMPK, SIRT1, PGC-1α, phosphorylated endothelial nitric oxide synthase and superoxide dismutase 1 and 2, and decreased that of NADPH oxidase 2 and 4. In Ang II-stimulated VSMCs, resveratrol treatment markedly decreased the number of senescence associated β-galactosidase stained cells and pro-fibrotic protein expression and increased the expression of AT2R and MasR. ConclusionsResveratrol protects against arterial aging and this effect is associated with reduced activity of the PRR–ACE–Ang II axis and stimulation of the ACE2–Ang-(1–7)–ATR2–MasR axis.

Aquatide Activation of SIRT1 Reduces Cellular Senescence through a SIRT1-FOXO1-Autophagy Axis

Ultraviolet (UV) irradiation is a relevant environment factor to induce cellular senescence and photoaging. Both autophagy- and silent information regulator T1 (SIRT1)-dependent pathways are critical cellular processes of not only maintaining normal cellular functions, but also protecting cellular senescence in skin exposed to UV irradiation. In the present studies, we investigated whether modulation of autophagy induction using a novel synthetic SIRT1 activator, heptasodium hexacarboxymethyl dipeptide-12 (named as Aquatide), suppresses the UVB irradiation-induced skin aging. Treatment with Aquatide directly activates SIRT1 and stimulates autophagy induction in cultured human dermal fibroblasts. Next, we found that Aquatide-mediated activation of SIRT1 increases autophagy induction via deacetylation of forkhead box class O (FOXO) 1. Finally, UVB irradiation-induced cellular senescence measured by SA-β-gal staining was significantly decreased in cells treated with Aquatide in parallel to occurring SIRT1 activation-dependent autophagy. Together, Aquatide modulates autophagy through SIRT1 activation, contributing to suppression of skin aging caused by UV irradiation.

SIRT1 activation inhibits hyperglycemia-induced apoptosis by reducing oxidative stress and mitochondrial dysfunction in human endothelial cells.

Sustained hyperglycemic stimulation of vascular cells is involved in the pathogenesis of diabetes mellitus‑induced cardiovascular complications. Silent information regulator T1 (SIRT1), a mammalian sirtuin, has been previously recognized to protect endothelial cells against hyperglycemia‑induced oxidative stress. In the present study, human umbilical vein endothelial cells (HUV‑EC‑C) were treated with D‑glucose, and the levels of oxidative stress, mitochondrial dysfunction, the rate of apoptosis and SIRT1 activity were measured. The effect of manipulated SIRT1 activity on hyperglycemia‑induced oxidative stress, mitochondrial dysfunction and apoptosis was then assessed using the SIRT1 activator, resveratrol (RSV), and the SIRT1 inhibitor, sirtinol. The present study confirmed that hyperglycemia promotes oxidative stress and mitochondrial dysfunction in HUV‑EC‑C cells. The accumulation of reactive oxygen species, the swelling of mitochondria, the ratio of adenosine 5'‑diphosphate to adenosine 5'‑triphosphate and localized mitochondrial superoxide levels were all increased following D‑glucose treatment, whereas the mitochondrial membrane potential was significantly reduced by >50mg/ml D‑glucose treatment. In addition, hyperglycemia was confirmed to induce apoptosis in HUV‑EC‑C cells. Furthermore, the results confirmed the prevention and aggravation of hyperglycemia‑induced apoptosis by RSV treatment and sirtinol treatment, via the amelioration and enhancement of oxidative stress and mitochondrial dysfunction in HUV‑EC‑C cells, respectively. In conclusion, the present study revealed that hyperglycemia promotes oxidative stress, mitochondrial dysfunction and apoptosis in HUV‑EC‑C cells, and manipulation of SIRT1 activity regulated hyperglycemia‑induced mitochondrial dysfunction and apoptosis in HUV‑EC‑C cells. The data revealed the protective effect of SIRT1 against hyperglycemia‑induced apoptosis via the alleviation of mitochondrial dysfunction and oxidative stress.