Silent Information Regulator 1 mRNA Research Articles

Effect of moxibustion on the SIRT1/FOXO3a signaling pathway in ApoE-/- mice with atherosclerosis.

To observe the effects of moxibustion on blood lipid metabolism, pathological morphology of thoracic aorta, and the expression of silent information regulator 1 (SIRT1) and forkhead box transcription factor O3a (FOXO3a) in ApoE-/- atherosclerosis (AS) mice, so as to explore the potential mechanism of moxibustion in preventing and treating AS. Ten C57BL/6J mice were fed a normal diet as the control group, and 30 ApoE-/- mice were fed a high-fat diet to establish the AS model, which were randomly divided into the model group, simvastatin group, and moxibustion group, with 10 mice in each group. From the first day of modeling, mice in the moxibustion group received mild moxibustion treatment at "Shenque"(CV8), "Yinlingquan"(SP9), bilateral "Neiguan"(PC6) and "Xuehai"(SP10) for 30 min per time；the mice in the simvastatin group were given simvastatin orally (2.5 mg·kg-1·d-1), with both treatments given once daily, 5 times a week, with a total intervention period of 12 weeks. The body weight and general condition of the mice were observed and recorded during the intervention period. After the intervention, the contents of serum total cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C) were measured using an automated biochemistry analyzer. Hematoxylin eosin (HE) staining was used to observe the pathological morphology of the thoracic aorta. ELISA was used to measure the contents of serum oxidized low-density lipoprotein (ox-LDL) and superoxide dismutase (SOD) activity. Western blot and real-time fluorescent quantitative PCR analysis were used to detect the expression levels of SIRT1 and FOXO3a protein and mRNA in the thoracic aorta. Compared with the control group, body weight at the 8th and 12th week, serum TC, TG, LDL-C, and ox-LDL contents of the model group mice were significantly increased(P<0.05, P<0.01), while the HDL-C contents, SOD activity, and the expression levels of SIRT1 protein and mRNA in the thoracic aorta were significantly decreased(P<0.05, P<0.01). HE staining showed thickening of the aortic intima, endothelial cell degeneration, swelling, and shedding. Compared with the model group, body weight at the 8th and 12th week, serum TC, TG, LDL-C, and ox-LDL contents of mice in the simvastatin group and moxibustion group were significantly decreased(P<0.01), while the serum SOD activity, expression levels of SIRT1 protein and mRNA in the thoracic aorta were significantly increased(P<0.01). The HDL-C contents were significantly increased in the simvastatin group(P<0.05). The thoracic aortic structure was more intact in both groups, with a more regular lumen and orderly arrangement of the elastic membrane in the media, and a slight amount of endothelial cell degeneration and swelling in the intima. There was no significant difference in the evaluated indexes between the moxibustion group and the simvastatin group and the pathological changes in the thoracic aorta were similar between the two groups. Moxibustion can reduce the body weight of AS model mice, regulate lipid levels, repair vascular intima, and alleviate endothelial damage. Its mechanism of action may be related to the regulation of the SIRT1/FOXO3a signaling pathway to improve oxidative damage.

SIRT1 and miR-34a-5p Expression in PBMCs as Potential Biomarkers for Patients With Type 2 Diabetes With Cognitive Impairments.

Patients with type 2 diabetes mellitus (T2DM) are at significantly increased risk of Alzheimer disease (AD). However, no biomarkers are available for early identification of patients with T2DM with cognitive impairment (T2DM-CI). Mitochondrial dysfunction is linked to AD. Silent Information Regulator 1 (SIRT1), which is responsible for regulating mitochondrial biogenesis, and its related miRNAs were also altered in AD. This study aimed to determine whether mitochondrial function in peripheral blood mononuclear cells (PBMCs) of patients with T2DM-CI was altered and if these alterations could be used as biomarkers. A total of 374 subjects were enrolled, including AD, T2DM-CI, T2DM-nCI (T2DM without cognitive impairment), and healthy controls. The mitochondrial function was determined using a commercial assay kit. The mitochondrial DNA (mtDNA) content, the expression of SIRT1, and selected miRNAs in PBMCs were measured by quantitative polymerase chain reaction. The correlations and diagnostic accuracy were assessed using the Spearman correlation coefficient or receiver operating characteristics analysis, respectively. We found significant changes in mitochondrial function in PBMCs of patients with AD compared with controls (all P < .05), which were not found in T2DM-CI. However, mtDNA content and SIRT1 mRNA expression were lower in PBMCs of patients with T2DM-CI, while miR-34a-5p expression was higher than in patients with T2DM-nCI (all P < .05). A combination of SIRT1 and miR-34a-5p demonstrated excellent discrimination between T2DM-CI and T2DM-nCI (area under the curve = 0.793; sensitivity: 80.01%; specificity: 78.46%). Furthermore, correlation analysis revealed a link between miR-34a-5p expression and hyperglycemia in T2DM-CI. Our findings revealed that there was an alteration of mitochondria at the peripheral level in patients with T2DM-CI. SIRT1 combined with miR-34a-5p in PBMCs performed well in identifying patients with T2DM-CI and may be a promising biomarker.

Moxibustion at "Zusanli"(ST36) mitigates senescence in subacute aging rats via regulating SIRT1/p53 signaling pathway

To observe the effect of moxibustion at "Zusanli"(ST36) on the silent information regulator 1 (SIRT1) /p53 signaling pathway in subacute aging model rats, so as to reveal its mechanisms in delaying aortic aging. Male SD rats were divided into blank group, model group, prevention group and treatment group, with 20 rats in each group. Subacute aging model was established by intraperitoneal injection of D-galactose(500 mg·kg-1·d-1). In the morning, rats in the prevention group received moxibustion at ST36 with 3 moxa cones after modeling operation, once every day for 42 d. From the day after the 42-day modeling, rats in the treatment group received the same moxibustion treatment as the prevent group for 28 d. Rats in the blank and model group were fixed in the similar way as the other two groups, for 5 min. Contents of serum SIRT1, p53, endothelial nitric oxide synthase(eNOS) and vascular endothelial growth factor(VEGF) were detected by ELISA. Histopathological changes of aortic tissue were observed after HE staining. Expressions of SIRT1 and p53 mRNAs and proteins in aortic tissue were detected by qPCR and Western blot. Compared with the blank group, the model group showed aging symptoms, the prevention group was similar to the blank group, and the treatment group was slightly better than the model group. Compared with the blank group, content of serum p53, expressions of p53 mRNA and protein in aortic tissues were significantly increased (P<0.05, P<0.01), while contents of serum SIRT1, VEGF, eNOS, and expressions of SIRT1 mRNA and protein in aortic tissues were significantly decreased (P<0.05, P<0.01) in the model group. Compared with the model group, content of serum p53, and expression of p53 mRNA and protein in aortic tissues were significantly decreased (P<0.05, P<0.01) in the prevention and treatment groups, while the contents of serum SIRT1, VEGF, eNOS, and the expressions of SIRT1 mRNA and protein in aortic tissues were significantly increased (P<0.05, P<0.01). Compared with the treatment group, rats in the prevention group displayed significant improvement of the above indexes (P<0.05). Compared with the blank group, the endothelial cells were disordered, the vessel wall was significantly thickened, and the senescent cells were increased in the model group; the blood vessel walls were thinner to varying degrees, and the senescent cells were reduced and unevenly distributed in the prevention and treatment groups. The histopathological lesion was improved more obviously in the prevention group than the treatment group. Moxibustion at ST36 can alleviate vascular endothelial injury and oxidative stress in subacute aging rats, which may be related to its effect in regulating the SIRT1/p53 signaling pathway.

Role and mechanism of SIRT1 in regulating Nrf2/HO-1 signaling pathway in septic liver injury

To investigate the role and mechanism of silent information regulator 1 (SIRT1) in regulating nuclear factor E2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) signaling pathway in oxidative stress and inflammatory response to sepsis-induced liver injury. A total of 24 male Sprague-Dawley (SD) rats were randomly divided into sham operation (Sham) group, cecal ligation and puncture (CLP) group, SIRT1 agonist SRT1720 pretreatment (CLP+SRT1720) group and SIRT1 inhibitor EX527 pretreatment (CLP+EX527) group, with 6 rats in each group. Two hours before operation, SRT1720 (10 mg/kg) or EX527 (10 mg/kg) were intraperitoneally injected into the CLP+SRT1720 group and CLP+EX527 group, respectively. Blood was collected from the abdominal aorta at 24 hours after modeling and the rats were sacrificed for liver tissue. The serum levels of interleukins (IL-6, IL-1β) and tumor necrosis factor-α (TNF-α) were detected by enzyme-linked immunosorbent assay (ELISA). The serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were detected by microplate method. Hematoxylin-eosin (HE) staining was used to observe the pathological injury of rats in each group. The levels of malondialdehyde (MDA), 8-hydroxydeoxyguanosine (8-OHdG), glutathione (GSH) and superoxide dismutase (SOD) in liver tissue were detected by corresponding kits. The mRNA and protein expressions of SIRT1, Nrf2 and HO-1 in liver tissues were detected by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blotting. Compared with the Sham group, the serum levels of IL-6, IL-1β, TNF-α, ALT and AST in the CLP group were significantly increased; histopathological results showed that liver cords were disordered, hepatocytes were swollen and necrotic, and a large number of inflammatory cells infiltrated; the contents of MDA and 8-OHdG in liver tissue increased, while the contents of GSH and SOD decreased; and the mRNA and protein expressions of SIRT1, Nrf2 and HO-1 in liver tissues were significantly decreased. These results suggest that sepsis rats have liver dysfunction, and the levels of SIRT1, Nrf2, HO-1 and antioxidant protein in liver tissues were decreased, while the levels of oxidative stress and inflammation were increased. Compared with the CLP group, the levels of inflammatory factors and oxidative stress were significantly decreased in the CLP+SRT1720 group, the mRNA and protein expressions of SIRT1, Nrf2 and HO-1 were significantly increased [IL-6 (ng/L): 34.59±4.21 vs. 61.84±3.78, IL-1β (ng/L): 41.37±2.70 vs. 72.06±3.14, TNF-α (ng/L): 76.43±5.23 vs. 130.85±5.30, ALT (U/L): 30.71±3.63 vs. 64.23±4.59, AST (U/L): 94.57±6.08 vs. 145.15±6.86, MDA (μmol/g): 6.11±0.28 vs. 9.23±0.29, 8-OHdG (ng/L): 117.43±10.38 vs. 242.37±11.71, GSH (μmol/g): 11.93±0.88 vs. 7.66±0.47, SOD (kU/g): 121.58±5.05 vs. 83.57±4.84, SIRT1 mRNA (2-ΔΔCt): 1.20±0.13 vs. 0.46±0.02, Nrf2 mRNA (2-ΔΔCt): 1.21±0.12 vs. 0.58±0.03, HO-1 mRNA (2-ΔΔCt): 1.71±0.06 vs. 0.48±0.07, SIRT1 protein (SIRT1/β-actin): 0.89±0.04 vs. 0.58±0.03, Nrf2 protein (Nrf2/β-actin): 0.87±0.08 vs. 0.51±0.09, HO-1 protein (HO-1/β-actin): 0.93±0.14 vs. 0.54±0.12, all P < 0.05], these results indicated that SIRT1 agonist SRT1720 pretreatment could improve liver injury in sepsis rats. However, pretreatment with SIRT1 inhibitor EX527 showed the opposite effect [IL-6 (ng/L): 81.05±6.47 vs. 61.84±3.78, IL-1β (ng/L): 93.89±5.83 vs. 72.06±3.14, TNF-α (ng/L): 177.67±5.12 vs. 130.85±5.30, ALT (U/L): 89.33±9.52 vs. 64.23±4.59, AST (U/L): 179.59±6.44 vs. 145.15±6.86, MDA (μmol/g): 11.39±0.51 vs. 9.23±0.29, 8-OHdG (ng/L): 328.83±11.26 vs. 242.37±11.71, GSH (μmol/g): 5.07±0.34 vs. 7.66±0.47, SOD (kU/g): 59.37±4.28 vs. 83.57±4.84, SIRT1 mRNA (2-ΔΔCt): 0.34±0.03 vs. 0.46±0.02, Nrf2 mRNA (2-ΔΔCt): 0.46±0.04 vs. 0.58±0.03, HO-1 mRNA (2-ΔΔCt): 0.21±0.03 vs. 0.48±0.07, SIRT1 protein (SIRT1/β-actin): 0.47±0.04 vs. 0.58±0.03, Nrf2 protein (Nrf2/β-actin): 0.32±0.07 vs. 0.51±0.09, HO-1 protein (HO-1/β-actin): 0.19±0.09 vs. 0.54±0.12, all P < 0.05]. SIRT1 can inhibit the release of proinflammatory factors and alleviate the oxidative damage of hepatocytes by activating Nrf2/HO-1 signaling pathway, thus playing a protective role against CLP-induced liver injury.

Sirt1 Regulates Corneal Epithelial Migration by Deacetylating Cortactin.

Silent information regulator 1 (SIRT1) is a nicotinamide adenine dinucleotide (NAD+) dependent deacetylase, which plays an essential role in cellular metabolism, autophagy, and chromatin accessibility. Our study aimed to determine its role in controlling corneal epithelial wound healing (CEWH). Corneal epithelial (CE)-specific Sirt1 deletion mice were created using the Cre-lox system. CE debridement was used to create a CEWH model. Corneal epithelial cells (CECs) were collected with an Algerbrush. Western blot analysis and RT-qPCR were performed to determine protein and mRNA expression levels. SiRNA transfection technology knocked down SIRT1 and cortactin expression levels in human corneal epithelial cells. Scratch wound assay, MTS assay, and TUNEL assay determined cell migratory, proliferative, and apoptotic behavior, respectively. Co-immunoprecipitation probed for SIRT1 and cortactin interaction. Immunofluorescence staining evaluated the location and expression levels of SIRT1, cortactin, acetylated-cortactin, and F-actin. During CEWH, increases in SIRT1 mRNA and protein expression levels accompanied the downregulation of acetylated lysine in non-histone proteins. The loss of SIRT1 function reduced cell migration and, in turn, delayed CEWH. SIRT1 bound to and deacetylated cortactin in vitro and in vivo. Loss of either SIRT1 or cortactin suppressed wound edge lamellipodia formation, which is consistent with migration retardation. During CEWH, SIRT1 upregulation and its modification of cortactin boost CEC migration by increasing the development of lamellipodia at the wound edge. Therefore SIRT1 may serve as a potential target to enhance CEWH.

Effect of mild moxibustion on SIRT1/NF-κB signaling in atherosclerotic rabbits

To observe the effect of mild moxibustion on blood lipid, histopathological structure of the aortic arch, thoracic aortic silent information regulator 1 (SIRT1)/nuclear factor κB (NF-κB) signaling pathway in atherosclerosis (AS) rabbits, so as to explore its underlying mechanisms in improving AS. Sixty male rabbits were randomly divided into control group (n=12), model group(n=11), mild moxibustion group (n=11), mild moxibustion + blocker (blocker) group (n=12). The AS model was established by feeding the rabbits with high-fat forage for 8 weeks, followed by immune response damage. Mild moxibustion was applied to "Danzhong"(CV17), "Shenque"(CV8) and "Neiguan" (PC6, bilateral) and "Xuehai" (SP10, bilateral) for 30 min, once daily, 3 times a week for 4 weeks. Rabbits of the blocker group received intraperitoneal injection of EX527 (a selective inhibitor of SIRT1, 5 mg·kg-1·d-1) 30 min before moxibustion. Rabbits of the control and model groups were only grabbed and fixed without intervention. After the intervention, the contents of serum triglyceride (TG) and total cholesterol (TC) were determined by enzymatic method, and those of serum low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) were determined by colorimetric method. The Sudan Ⅳ staining was employed to observe the histopathological structure of the aortic arch, and Western blot and fluorescence quantitative real time-PCR were used to detect the expressions of SIRT1 and NF-κB proteins and mRNAs in the thoracic aorta, respectively. Compared with the control group, the contents of serum TG, TC and LDL-C and the expression levels of NF-κB protein and mRNA were significantly increased (P<0.01, P<0.05), whereas the content of HDL-C and the expression of SIRT1 mRNA markedly decreased in the model group (P<0.01). After mild moxibustion, the contents of serum TG, TC, and LDL-C and the expression of NF-κB protein and mRNA were significantly down-regulated (P<0.01, P<0.05), while the content of HDL-C and the expression levels of SIRT1 protein and mRNA significantly up-regulated in the mild moxibustion group (P<0.05, P<0.01). There were no significant differences between the blocker and model groups in all the indexes (P>0.05). Compared with the mild moxibustion group, the serum TG, TC, and LDL-C contents and NF-κB protein expression were significantly increased (P<0.01, P<0.05), and HDL-C content and the expression of SIRT1 protein and mRNA significantly decreased (P<0.05, P<0.01) in the blocker group. Sudan Ⅳ staining showed vague structure of the aortic arch with obvious lipid infiltration in the model group, which was relatively milder in the mild moxibustion. Mild-moxibustion can reduce blood lipid levels and endothelial damage in atherosclerotic rabbits, which may be related to its function in regulating SIRT1/NF-κB signaling pathway.

Zearalenone promotes follicle development through activating the SIRT1/PGC-1α signaling pathway in the ovaries of weaned gilts.

This study aimed to investigate the effect of zearalenone (ZEA) exposure on follicular development in weaned gilts, and its mechanism based on the silent information regulator 1 (SIRT1)/peroxisome proliferator-activated receptor-γ co-activator 1α (PGC-1α) signaling pathway. A total of 32 healthy female weaned piglets (Landrace × Yorkshire × Duroc) with an average body weight of 12.39±0.24kg were randomly allotted to a basal diet supplemented with 0, 0.15, 1.5, or 3.0mg/kg ZEA for a 32-d feeding test. Blood and ovarian samples were obtained at the end of the experiment to determine serum toxin concentrations, ovarian histology, and the expressions of proliferating cell nuclear antigen (PCNA) and SIRT1/PGC-1α signaling pathway-related genes. Results showed that the vulva area, serum concentrations of ZEA, α-zearalenol and β-zearalenol, the thickness of the growing follicular layer, and the diameter of the largest growing follicles, as well as the expressions of SIRT1, PGC-1α, estrogen-related receptor α (ERRα), ATP synthase subunit beta (ATP5B), and PCNA, increased linearly (P < 0.05) with increasing dietary ZEA, whereas the thickness of the primordial follicle layer decreased linearly (P < 0.05). Immunohistochemical analysis showed that the immunoreactive substances of SIRT1 and PGC-1α in the ovaries enhanced with the increasing dietary ZEA (P < 0.05). In addition, the thickness of the growing follicular layer and the diameter of the largest growing follicle were positively correlated with relative mRNA and protein expressions of SIRT1, PGC-1α, ERRα, ATP5B, and PCNA (P < 0.05). However, the thickness of the primordial follicle layer was negatively correlated with the mRNA and protein expression of SIRT1, PGC-1α, ERRα, ATP5B, and PCNA (P < 0.05). Interestingly, the 1.5mg/kg ZEA treatment had highly hyperplastic follicles, whereas 3.0mg/kg ZEA resulted in a large number of follicular atresia, which indicated that low-dose ZEA exposure accelerated follicular proliferation, while high-dose ZEA promoted follicular atresia, although the critical value interval needs further confirmation. Results provide a theoretical basis for finding the therapeutic target of ZEA-induced reproductive disorders in weaned gilts.

Peripheral Blood Levels of miR-448 and SIRT1 in Patients with Deep Venous Thrombosis and Their Relationship.

The goal of this study was to investigate the changes in peripheral blood levels of miR-448 and silent information regulator 1 (SIRT1) in patients with deep venous thrombosis (DVT) and to analyze their relationship. A total of 112 patients treated from January 2019 to June 2020 were divided into DVT group (n = 40) and non-DVT group (n = 72). Fasting venous blood was extracted to separate serum and peripheral blood mononuclear cells (PBMCs). Enzyme-linked immunosorbent assay (ELISA) was employed to measure serum SIRT1 protein, and qPCR was utilized to detect miR-448 expression in PBMCs. The clinical data, serum indicators, and expressions of SIRT1 and miR-448 were compared, and the correlations of miR-448 and SIRT1 with DVT were analyzed using a multivariate Cox regression model. TargetScan Release 7.1 was used to predict the possibility of binding sites between miR-448 and SIRT1 mRNA 3'-untranslated region (3'-UTR), and dual-luciferase reporter assay was used to determine the targeting of miR-448 and SIRT1. HeLa cells were divided into overexpression, inhibition, and blank control groups. The cells were harvested 24 hours after transfection, followed by detection of SIRT1 mRNA expression by qPCR and measurement of supernatant SIRT1 protein expression by ELISA. Serum SIRT1 protein level was lower and miR-448 expression in PBMCs was higher in DVT group than those in non-DVT group (p < 0.05). DVT group had a larger number of patients with vascular diseases and history of venous thrombosis than that of non-DVT group (p < 0.05). miR-448 was an independent risk factor for postoperative DVT, and SIRT1 was a protective factor (p < 0.01). There were potential complementary base binding sites between miR-448 and SIRT1 mRNA 3'-UTR. Dual-luciferase reporter assay verified the targeted regulation between miR-448 and SIRT1. HeLa cell SIRT1 mRNA expression and supernatant SIRT1 protein expression were lower in overexpression group while higher in inhibition group than those in blank control group (p < 0.05). Serum SIRT1 protein level decreases while miR-448 expression in PBMCs increases in patients with DVT, and miR-448 inhibits SIRT1 expression through binding to SIRT1 mRNA 3'-UTR with complementary bases, thus inducing inflammatory response to participate in the formation of DVT. Targeting miR-448 to regulate cytokine expression may become an effective target and approach for the treatment of DVT. miR-488 combined with SIRT1 has a high predictive value for the occurrence of DVT.

Effect of electroacupuncture combined with Zhuang-medicine-thread moxibustion on silent information regulator-1/nuclear factor κB signaling pathway in gastric antrum of diabetic gastroparesis rats

To observe the effect of electroacupuncture (EA) combined with Zhuang-medicine-thread moxibustion on silent information regulator-1 (SIRT1)/nuclear factor kappa B (NF-κB) signal pathway and inflammatory factor expression in gastric antrum tissue of diabetic gastroparesis (DGP) rats, so as to explore its mechanism underlying improvement of DGP. Male SD rats were randomly divided into normal, model, medication, EA, Zhuang-medicine-thread moxibustion (moxibustion) and EA+moxibustion groups (n=12 per group). The DGP model was established by intraperitoneal injection of streptozotocin (STZ). Rats of the medication group were treated by gavage of 0.15 mg/mL mosapride citrate suspension. EA (10 Hz /50 Hz, 2 mA) or moxibustion (3 cones) or EA+moxibustion was applied to "Zhongwan"(CV12), bilateral "Neiguan"(PC6) and bilateral "Sanyinjiao"(SP6) of the related group for 20 min, once a day for 3 weeks. Blood glucose, gastric emptying rate and intestinal propulsion rate were measured. The levels of serum interleukin (IL)-6, IL-8, IL-10 and tumor necrosis factor-α(TNF-α) were detected by ELISA; the phosphorylation level of the phosphorylated inhibitor of nuclear factor κBα inhibitor (pIκ-Bα), the protein and mRNA expression of NF-κB p65 and SIRT1 in the gastric antrum tissue were detected by Western blot and real-time quantifitative PCR, respectively. (1) Compared with the normal group, the levels of blood glucose, serum IL-6, IL-8, TNF-α, and gastric pIκ-Bα and NF-κB p65 protein and mRNA expressions were significantly increased (P<0.01), and the gastric emptying rate, intestinal propulsion rate, serum IL-10 level, and SIRT1 protein and mRNA expressions were considerably decreased in the model group (P<0.01). (2) In contrast to the model group, the blood glucose in the EA, moxibustion and EA+moxibustion groups, serum IL-6, IL-8 and TNF-α levels in the 4 treatment groups, as well as NF-κB p65 protein expression in the medication and EA+moxibustion groups, and NF-κB p65 mRNA expression and pIκ-Bα protein and mRNA expression in the 4 treatment groups were significantly decreased (P<0.01, P<0.05); while the gastric emptying rate and intestinal propulsive rate and IL-10 content in the 4 treatment groups, and SIRT1 protein and mRNA expression in the medication and EA+moxibustion groups were obviously increased (P<0.05, P<0.01). (3) The effects of EA+moxibustion were significantly superior to those of simple EA and moxibustion in increasing gastric emptying rate, IL-10, SIRT1 protein expression (P<0.05, P<0.01), and in lowering IL-8 and TNF-α contents, pIκ-Bα protein and mRNA expression and NF-κB p65 mRNA expression (P<0.05, P<0.01). No significant differences were found among the 4 intervention groups in promoting the intestinal propulsive rate and among the EA, moxibustion and EA+moxibustion groups in lowering blood glucose (P>0.05). EA combined with Zhuang-medicine-thread moxibustion can effectively reduce the level of serum inflammatory factors and regulate SIRT1/NF-κB signal pathway in DGP rats, which may contribute to its function in improving gastrointestinal movement.

Berberine mitigates nonalcoholic hepatic steatosis by downregulating SIRT1-FoxO1-SREBP2 pathway for cholesterol synthesis

ObjectiveTo investigate effects of berberine (BBR) on cholesterol synthesis in HepG2 cells with free fatty acid (FFA)-induced steatosis and to explore the underlying mechanisms. MethodsA steatosis cell model was induced in HepG2 cell line fed with FFA (0.5 mmol/L, oleic acid:palmitic acid = 2:1), and then treated with three concentrations of BBR; cell viability was assessed with cell counting kit-8 assays. Lipid accumulation in cells was observed through oil red O staining and total cholesterol (TC) content was detected by TC assay. The effects of BBR on cholesterol synthesis mediators were assessed by Western blotting and quantitative polymerase chain reaction. In addition, both silent information regulator 1 (SIRT1) and forkhead box transcription factor O1 (FoxO1) inhibitors were employed for validation. ResultsFFA-induced steatosis was successfully established in HepG2 cells. Lipid accumulation and TC content in BBR groups were significantly lower (P < 0.05, P < 0.01), associated with significantly higher mRNA and protein levels of SIRT1(P < 0.05, P < 0.01), significantly lower sterol regulatory element-binding protein 2 (SREBP2) and 3-hydroxy 3-methylglutaryl-CoA reductase levels (P < 0.05, P < 0.01), as well as higher Acetyl-FoxO1 protein level (P < 0.05, P < 0.01) compared to the FFA only group. Both SIRT1 inhibitor SIRT1-IN-1 and FoxO1 inhibitor AS1842856 blocked the BBR-mediated therapeutic effects. Immunofluorescence showed that the increased SIRT1 expression increased FoxO1 deacetylation, and promoted its nuclear translocation. ConclusionBBR can mitigate FFA-induced steatosis in HepG2 cells by activating SIRT1-FoxO1-SREBP2 signal pathway. BBR may emerge as a potential drug candidate for treating nonalcoholic hepatic steatosis.

The effects of compound centella formula on OxInflammation and silent information regulator 1 in a high-fat diet/streptozotocin-induced diabetic kidney disease rat model.

The Chinese decoction compound centella formula (CCF) is clinically effective against diabetic kidney disease (DKD), but the exact mechanism remains unclear. The present study aimed to investigate the effects of CCF on OxInflammation and silent information regulator 1 (SIRT1) levels in rats with streptozotocin (STZ)-induced diabetes. Sprague-Dawley rats were divided into CCF, losartan, diabetic control (DC) and normal control (NC) groups (n=7). Except for the NC, all subgroups of rats were fed a high-fat diet for 112 days and received a single intraperitoneal injection of 35 mg/kg STZ on day 29. All rats were sacrificed on day 112. High-performance liquid chromatography was performed to analyse asiaticoside, astragaloside and triptolide levels in CCF (0.3400, 0.0640 and 0.0001 mg/ml, respectively). Fasting blood glucose, urine protein-to-creatinine ratio, serum creatinine and blood urea nitrogen were quantified. Periodic acid Schiff staining, H&E staining and transmission electron microscopy were used to examine kidney pathological changes. The mRNA and protein expression levels of SIRT1 in renal tissues were analysed by reverse transcription-quantitative PCR, western blotting and immunohistochemistry. Oxidative stress was evaluated by measuring the levels of superoxide dismutase (SOD), malondialdehyde (MDA) and nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4) in renal tissues. TNF-α and NF-κB p65 subunit in renal tissues were assessed for inflammation. Compared with the rats in the NC group, the rats in the DC group exhibited renal injury with proteinuria, decreased expression levels of SIRT1 and SOD (P<0.01) and increased levels of MDA, NOX4, TNF-α and NF-κB p65 (P<0.01). CCF treatment reduced proteinuria (P<0.01), alleviated renal damage, decreased MDA, NOX4, TNF-α and NF-κB p65 levels (P<0.01), increased SOD levels (P<0.05) and increased SIRT1 mRNA and protein expression levels (P<0.01). The present study indicates that CCF effectively protects the kidney from diabetes by inhibiting OxInflammation and upregulating SIRT1.

Activation of NOD-like receptor protein 3 inflammasome mediates inflammatory response and apoptosis in septic intestinal injury model

To investigate the expression of NOD-like receptor protein 3 (NLRP3) inflammasome in intestinal injury models with different severity of sepsis and the inflammatory response and apoptosis mediated by NLRP3 inflammasome. Human colorectal adenocarcinoma cells (Caco-2) were cultured in vitro. The logarithmic growth phase cells were divided into blank control group (normal culture in complete medium) and lipopolysaccharide (LPS) 1, 2 and 4 mg/L groups (complete medium containing 1, 2 and 4 mg/L LPS, respectively). The supernatant were collected at 6, 12 and 24 hours, and the levels of tumor necrosis factor-α (TNF-α), interleukins (IL-6, IL-1β, IL-18) were detected by enzyme linked immunosorbent assay (ELISA). The apoptotic level of cells was detected by flow cytometry. The cells were harvested, and the real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to detect the mRNA expressions of NLRP3 and silent information regulator 1 (SIRT1). Western blotting was used to detect the protein expressions of NLRP3, SIRT1, caspase-1 and apoptosis-associated speck-like protein (ASC). ELISA results showed that the levels of IL-6, TNF-α, IL-1β, and IL-18 in cell supernatant of LPS groups increased in a dose-dependent and time-dependent manner as compared with the blank control group during the same intervention period. The increase was most significant in LPS 4 mg/L group at 24 hours [IL-6 (ng/L): 3.55±0.06 vs. 0.67±0.09, TNF-α (ng/L): 15.37±0.19 vs. 5.04±0.14, IL-1β (ng/L): 2.26±0.10 vs. 0.56±0.09, IL-18 (ng/L): 433.92±22.55 vs. 93.55±21.13, all P < 0.05]. The results of the apoptotic test showed that, compared with the blank control group, the apoptotic rate of LPS groups increased in a dose-dependent and time-dependent manner, and the apoptotic rate of LPS 4 mg/L group increased most significantly at 24 hours [(14.83±3.73)% vs. (5.87±1.17)%, P < 0.05]. RT-qPCR results showed that the expression level of NLRP3 mRNA was increased, while the expression level of SIRT1 mRNA was decreased with the increase of LPS intervention dose and the prolonging of intervention time. At 24 hours, there were significant differences between LPS 4 mg/L group and blank control group [NLRP3 mRNA (2-ΔΔCt): 8.20±2.82 vs. 1.00±0.36, SIRT1 mRNA (2-ΔΔCt): 0.58±0.01 vs. 1.03±0.06, both P < 0.05]. Western blotting showed that compared with the blank control group, the protein expression levels of NLRP3, caspase-1 and ASC in LPS groups were significantly increased, while the protein expression levels of SIRT1 were significantly decreased. During each intervention period, with the increase of LPS dose, the expressions of NLRP3, caspase-1 and ASC protein increased gradually, while the expression of SIRT1 protein decreased gradually. At 24 hours, the difference between the LPS 4 mg/L group and the blank control group was significant [NLRP3 protein (NLRP3/β-actin): 1.48±0.03 vs. 0.90±0.12, caspase-1 protein (caspase-1/β-actin): 1.18±0.11 vs. 0.72±0.09, ASC protein (ASC/β-actin) : 1.09±0.01 vs. 0.82±0.03, SIRT1 protein (SIRT1/β-actin): 0.48±0.03 vs. 0.76±0.05, all P < 0.05]. In vitro, in the sepsis induced intestinal inflammation model, with the increase of LPS intervention dose and the prolongation of intervention time, intestinal inflammatory response and cell apoptosis showed an increasing trend, which may be related to the up-regulation of NLRP3 inflammasome and its downstream products ASC and caspase-1, and to the down-regulation of SIRT1 expression.

Protective effects of ginsenoside Rc against acute cold exposure-induced myocardial injury in rats.

Ginsenoside Rc is one of the cardinal bioactive components of Panax ginseng. The present study aimed to investigate whether ginsenoside Rc exerted protective effects against acute cold exposure-induced myocardial injury in rats. Forty rats were randomly assigned into four groups: Control, model, ginsenoside Rc 10 mg/kg, and 20 mg/kg groups. Rats were intragastrically administrated with ginsenoside Rc (10, 20 mg/kg) or vehicle daily for 7 days. On the seventh day, all rats except the control group were exposed to low temperature. Cardiac function, myocardial enzyme activities, hemorheology, and inflammatory response were detected. Histopathological examination and apoptosis of cardiac tissues were performed. The expressions of silent information regulator 1 (SIRT1), B-cell lymphoma (Bcl-2), Bcl-2-associated X (Bax), procaspase-3, and the mRNA (messenger RNA) level of SIRT1 were measured by western blot and real-time quantitative polymerase chain reaction (PCR) analysis. Ginsenoside Rc significantly improved cardiac function, diminished the activities of lactate dehydrogenase (LDH), aspartate aminotransferase, and creatine kinase isoenzyme (CK-MB), and regulated abnormal hemorheology in acute cold-exposed rats (p < 0.05 or p < 0.01). Furthermore, ginsenoside Rc could attenuate myocardial histological changes and structural abnormalities, decrease apoptotic cells and reduce the mRNA levels and activity of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and IL-6 (p < 0.01). In addition, ginsenoside Rc upregulated the expressions of SIRT1, Bcl-2, and procaspase-3 and downregulated that of Bax (p < 0.01). The changes in both the mRNA and protein expression levels of SIRT1 were similar. The results of the current study suggested that ginsenoside Rc could alleviate acute cold exposure-induced myocardial injury in rats by inhibiting cardiomyocyte apoptosis via regulating SIRT1 expression and attenuating the inflammatory responses. PRACTICAL APPLICATION: The current study indicated that ginsenoside Rc could alleviate acute cold exposure-induced myocardial injury in rats. Ginsenoside Rc could be potentially used as a bioactive ingredient in processed functional food products or food supplements to prevent from acute cold exposure-induced myocardial injury.

SIRT1 suppresses burn injury-induced inflammatory response through activating autophagy in RAW264.7 macrophages

The present study sought to investigate the association between silent information regulator 1 (SIRT1) and autophagy during systemic inflammatory response syndrome following burn injury. The experimental burn model in mice...

Regulation of the SIRT1 signaling pathway in NMDA-induced Excitotoxicity

Silent Information Regulator 1 (SIRT1), an NAD+-dependent deacetylase, contributes to the neuroprotective effect. However, intracellular signaling pathways that affect SIRT1 function remain unknown. It is well known that N-methyl-D-aspartate (NMDA) receptor activation induces calcium influx which then activates PKC, and SIRT1 is a mRNA target for HuR protein. We hypothesize that Ca2+-PKC-HuR-SIRT1 pathway modulates SIRT1 function. The present study is to investigate the potential pathway of SIRT1 in the SH-SY5Y cell line as an in vitro model of NMDA-induced neurotoxicity. The results showed that: (1) SIRT1 levels were downregulated in NMDA model; (2) NMDA induced an increase in serine phosphorylation of HuR, while inhibition of serine phosphorylation of HuR increased SIRT1 levels, promoting cell survival; (3) PKC inhibitor (Gö 6976) reversed NMDA insults and also suppressed serine phosphorylation of HuR; (4) 1,2-bis(2-aminophenoxy)ethane-N,N,N’,N’-tetraacetic acid (BAPTA-AM), an intracellular calcium chelator, fully reversed NMDA insults and also inhibited PKC activity evoked by NMDA. These results indicate that intracellular elevated Ca2+ activates PKC, which phosphorylates HuR and then promotes SIRT1 mRNA decay and subsequent neuronal death in NMDA model. Therefore, the study suggests that inhibition of Ca2+-PKC-HuR-SIRT1 pathway could be an effective strategy for preventing certain neurological diseases related to NMDA excitotoxicity.

Xuefu Zhuyu Oral Liquid () Prevents Apoptosis of Ischemic Myocardium Cells in Rats by Regulating SIRT1 and Its Pathway-Related Genes.

To observe the changes of ischemic myocardial cells apoptosis in rats following intervention with Xuefu Zhuyu Oral Liquid (, XFZY), as well as changes of protein expression of silent information regulator 1 (SIRT1) and SIRT1 pathway-related genes. H9c2 rat myocardial cells were divided into 6 groups: control group, oxygen glucose deprivation (OGD) group, SIRT1 siRNA group, OGD+SIRT1 siRNA group, OGD+XFZY group, and OGD+SIRT1 siRNA+XFZY group. Quantitative fluorescent polymerase chain reaction (PCR) and Western blot were used to detect the concentration variations of SIRT1 and its pathway-related genes and corresponding protein expression after XFZY intervention and SIRT1 transfection. Compared with the control group, the mRNA and protein expressions of SIRT1 were decreased obviously, while the mRNA and protein levels of P53, FoxO1, FoxO3, FoxO4 and nuclear factor kappa B (NF-ΚB) were increased in the OGD group, SIRT1 siRNA group, and OGD+SIRT1 siRNA group (P<0.01). Compared with the OGD group and OGD+SIRT1 siRNA group, the treatment of XFZY inhibited the decline in SIRT1 mRNA and protein expressions (P<0.01), and down-regulated the mRNA and protein levels of P53, FoxO1, FoxO3, FoxO4 and NF-ΚB, respectively (P<0.05 or P<0.01). XFZY could prevent myocardial cells apoptosis probably by increasing the mRNA and protein expressions of SIRT1 and inhibiting the mRNA and protein expressions of P53, NF- K B, FoxO1, FoxO3 and FoxO4.

The resistant effect of SIRT1 in oxidative stress-induced senescence of rat nucleus pulposus cell is regulated by Akt-FoxO1 pathway.

Objective: The senescence of nucleus pulposus (NP) cells induced by oxidative stress is one of the important causes of intervertebral disc degeneration (IDD). Herein, we investigated the role and action mechanism of silent information regulator 1 (SIRT1) in oxidative stress-induced senescence of rat NP cell.Methods: Premature senescence of rat NP cells was induced by sublethal concentration of hydrogen peroxide (H2O2) (100 μM). SIRT1 was activated with SRT1720 (5 μM) to explore its effect on NP cells senescence. FoxO1 and Akt were inhibited by AS1842856 (0.2 μM) and MK-2206 (5 μM), respectively, to explore the role of Akt-FoxO1-SIRT1 axis in rat NP cells. Pretreatment with the resveratrol (20 μM), a common antioxidant and indirect activator of SIRT1, was done to investigate its role in senescent rat NP cells.Results: The mRNA and protein levels of SIRT1 were decreased in H2O2-induced senescent rat NP cells, and that specific activation of SIRT1 suppresses senescence. And the Akt-FoxO1 pathway, as the upstream of SIRT1, might be involved in the regulation of H2O2-induced senescence of rat NP cells by affecting the expression of SIRT1. In addition, the resveratrol played an anti-senescence role in rat NP cells, which might affect the Akt-FoxO1-SIRT1 axis.Conclusion: SIRT1 ameliorated oxidative stress-induced senescence of rat NP cell which was regulated by Akt-FoxO1 pathway, and resveratrol exerted anti-senescence effects by affecting this signaling axis.

Buyang Huanwu Decoction Targets SIRT1/VEGF Pathway to Promote Angiogenesis After Cerebral Ischemia/Reperfusion Injury

Background: Ischemia stroke is known as the major cause of morbidity and mortality. Buyang Huanwu Decoction (BHD), a classical traditional Chinese medicine (TCM) formula, has been used to prevent and treat stoke for hundreds of years. The purpose of present study is to investigate the effects of BHD on angiogenesis in rats after cerebral ischemia/reperfusion (I/R) injury targeting Silent information regulator 1 (SIRT1) / Vascular endothelial growth factor (VEGF) pathway.Methods: The cerebral I/R injury model was induced by middle cerebral artery occlusion (MCAO). Adult Sprag-Dawley (SD) rats were randomly divided into five groups: sham group, normal saline (NS) group, BHD group, BHD+EX527 (SIRT1 specific inhibitor) group, and NS+EX527 group. Each group was divided into the subgroups according to 1, 3, 7, or 14 days time-point after cerebral ischemia/reperfusion, respectively. Neurological function score (NFS) was evaluated by the Rogers scale; microvascular density (MVD) in brain tissue around infarction area was observed by immunofluorescence; and the expression of SIRT1 and VEGF was assessed by Western Blot and Quantitative Real-time-PCR.Results: BHD can significantly improve NFS (P < 0.05), increase the MVD in the boundary ischemic area (P < 0.01) and elevate the expression of protein and mRNA of SIRT1 and VEGF following I/R injury (P < 0.01). In contrast, treatment with EX527 reversed the BHD-induced improvements in NFS (P < 0.01) and decreased the MVD (P < 0.01) and the expression of SIRT1 and VEGF (P < 0.05).Conclusion: BHD exerts neuroprotection targeting angiogenesis through the up-regulation of SIRT1/VEGF pathway against cerebral ischemic injury in rats.

RNA-binding protein RPS3 contributes to hepatocarcinogenesis by post-transcriptionally up-regulating SIRT1.

Although several studies indicate that RNA-binding proteins (RBPs) contribute to key steps in a variety of physiological processes and cancer, the detailed biological functions and mechanisms remain to be determined. By performing bioinformatics analysis using well-established hepatocellular carcinoma (HCC) datasets, we identified a set of HCC progression-associated RBPs (HPARBPs) and found that the global expression of HPARBPs was significantly correlated with patient prognosis. Among the 42 HPARBPs, human ribosomal protein S3 (RPS3) was one of the most abundant genes whose role remains uncharacterized in HCC. Gain- and loss-of-function analyses demonstrated that RPS3 promoted HCC tumorigenesis both in vitro and in vivo. Mechanistically, we revealed that silent information regulator 1 (SIRT1) was a critical target of RPS3 and was essential for sustaining the RPS3-induced malignant phenotypes of HCC cells. RPS3 stabilized SIRT1 mRNA by binding to AUUUA motifs in the 3448–3530 region of the 3′ untranslated region (UTR) of SIRT1 mRNA. In addition, we found that (5-formylfuran-2-yl) methyl 4-hydroxy-2-methylenebutanoate (FMHM) inhibited HCC progression by repressing the RPS3/SIRT1 pathway. Our study unveils a novel extra-ribosomal role of RPS3 in facilitating hepatocarcinogenesis via the post-transcriptional regulation of SIRT1 expression and proposes that the RPS3/SIRT1 pathway serves as a potential therapeutic target in HCC.

Circular RNA hsa_circ_0076248 promotes oncogenesis of glioma by sponging miR-181a to modulate SIRT1 expression.

Glioma is one of the most common primary malignancies of the central nervous system, which has aggressive clinical behavior and a poorer prognosis. MicroRNAs (miRs) are a class of small noncoding RNAs that function as mediators of gene expression, which can be sponged by circRNA provided with a closed circular structure. Dysregulations of circular RNAs (circRNAs) and miRs have been implicated in the development and progression of glioma. In the current study, we investigated the role of circular RNA hsa_circ_0076248 in mediating the oncogenesis of glioma by sponging miR‐181a to modulate silent information regulator 1 (SIRT1) expression in vitro and in vivo. The quantitative real‐time polymerase chain reaction results showed that the expression of miR‐181a was significantly decreased in glioma tissues and cell lines compared with normal brain tissues and normal gliocyte, respectively, and the expression of hsa_circ_0076248 and SIRT1 demonstrated the opposite. Bioinformatics analysis identified hsa_circ_0076248 could sponge miR‐181a, and miR‐181a could target the mRNA of SIRT1. Our results verified that downregulating hsa_circ_0076248 or upregulating miR‐181a could depress the proliferation and invasion of glioma in vitro and in vivo. The experiment also showed that downregulating hsa_circ_0076248 or upregulating miR‐181a could remarkably promote the temozolomide chemotherapy sensitivity. Furthermore, Western blot analysis testified that downregulating hsa_circ_0076248 or upregulating miR‐181a could promote the expression of p53 and SIRT1. In summary, our study sheds light on the regulatory mechanism of hsa_circ_0076248 in glioma growth and invasion via sponging miR‐181a, which downregulates the SIRT1 expression and also suggests that hsa_circ_0076248, miR‐181a, and SIRT1 may serve as potential therapeutic targets for glioma.