Resveratrol Alleviates Osteoporosis by Promoting Osteogenic Differentiation of Bone Marrow Mesenchymal Stem Cells via SITR1/PI3K/AKT Pathway / El Resveratrol Alivia la Osteoporosis al Promover la Diferenciación Osteogénica de las Células Madre Mesenquimales de la Médula Ósea a Través de la Vía SITR1/PI3K/AKT

SUMMARY: Senile osteoporosis is mainly caused by reduced osteoblast differentiation and has become the leading cause of fractures in the elderly worldwide. Natural organics are emerging as a potential option for the prevention and treatment of osteoporosis. This study was designed to study the effect of resveratrol on osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) in osteoporosis mice. A mouse model of osteoporosis was established by subcutaneous injection of dexamethasone and treated with resveratrol administered by gavage. In vivo and in vitro, we used western blot to detect protein expression, and evaluated osteogenic differentiation of BMSCs by detecting the expression of osteogenic differentiation related proteins, calcium deposition, ALP activity and osteocalcin content. Resveratrol treatment significantly increased the body weight of mice, the level of serum Ca2+, 25(OH)D and osteocalcin, ration of bone weight, bone volume/total volume, trabecular thickness, trabecular number, trabecular spacing and cortical thickness in osteoporosis mice. In BMSCs of osteoporosis mice, resveratrol treatment significantly increased the expression of Runx2, osterix (OSX) and osteocalcin (OCN) protein, the level of calcium deposition, ALP activity and osteocalcin content. In addition, resveratrol treatment also significantly increased the expression of SIRT1, p-PI3K / PI3K and p-AKT / AKT in BMSCs of osteoporosis mice. In vitro, resveratrol increased the expression of SIRT1, p-PI3K / PI3K and p-AKT / AKT, Runx2, OSX and OCN protein, the level of calcium deposition, ALP activity and osteocalcin content in BMSCs in a concentration-dependent manner, while SIRT1 knockdown significantly reversed the effect of resveratrol. Resveratrol can attenuate osteoporosis by promoting osteogenic differentiation of bone marrow mesenchymal stem cells, and the mechanism may be related to the regulation of SIRT1/PI3K/AKT pathway.

La osteoporosis senil es causada principalmente por una diferenciación reducida de osteoblastos y se ha convertido en la principal causa de fracturas en las personas mayores en todo el mundo. Los productos orgánicos naturales están surgiendo como una opción potencial para la prevención y el tratamiento de la osteoporosis. Este estudio fue diseñado para estudiar el efecto del resveratrol en la diferenciación osteogénica de las células madre mesenquimales de la médula ósea (BMSC) en ratones con osteoporosis. Se estableció un modelo de osteoporosis en ratones mediante inyección subcutánea de dexametasona y se trató con resveratrol administrado por sonda. In vivo e in vitro, utilizamos Western blot para detectar la expresión de proteínas y evaluamos la diferenciación osteogénica de BMSC detectando la expresión de proteínas relacionadas con la diferenciación osteogénica, la deposición de calcio, la actividad de ALP y el contenido de osteocalcina. El tratamiento con resveratrol aumentó significativamente el peso corporal de los ratones, el nivel sérico de Ca2+, 25(OH)D y osteocalcina, la proporción de peso óseo, el volumen óseo/ volumen total, el espesor trabecular, el número trabecular, el espaciado trabecular y el espesor cortical en ratones con osteoporosis. En BMSC de ratones con osteoporosis, el tratamiento con resveratrol aumentó significativamente la expresión de las proteínas Runx2, osterix (OSX) y osteocalcina (OCN), el nivel de deposición de calcio, la actividad de ALP y el contenido de osteocalcina. Además, el tratamiento con resveratrol también aumentó significativamente la expresión de SIRT1, p-PI3K/PI3K y p-AKT/AKT en BMSC de ratones con osteoporosis. In vitro, el resveratrol aumentó la expresión de las proteínas SIRT1, p-PI3K/PI3K y p- AKT/AKT, Runx2, OSX y OCN, el nivel de deposición de calcio, la actividad de ALP y el contenido de osteocalcina en BMSC de manera dependiente de la concentración, mientras que La caída de SIRT1 revirtió significativamente el efecto del resveratrol. El resveratrol puede atenuar la osteoporosis al promover la diferenciación osteogénica de las células madre mesenquimales de la médula ósea, y el mecanismo puede estar relacionado con la regulación de la vía SIRT1/PI3K/AKT.

Exploring Effect and Mechanism of Astragaloside on Repair and Healing in Chronic Non-healing Wound Rat Model Based on SIRT1 and PI3K/Akt Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the promotional effect of astragaloside on the repair and healing of chronic non-healing wounds and its mechanism. MethodA total of 60 male SD rats were constructed with full-layer skin defect wounds on the back， and except for the control （Con） group， the rest were constructed with non-healing wounds， which were then randomly divided into the sham-operation （sham） group， the low-dose astragaloside group， the high-dose astragaloside group， the astragaloside + LY294002 ［phosphatidylinositol 3-kinase （PI3K） inhibitor］ group， and the astragaloside + EX527 ［silencing regulatory protein 1 （SIRT1） inhibitor］ group. The percentage of wound area in each group was observed on the 2nd， 4th， 6th， and 8th days after wound molding. Collagen type Ⅰ alpha 1 （COL1A1） and alpha smooth muscle actin （α-SMA） expressions in the wound tissue were detected by immunofluorescence. Hematoxylin and eosin （HE） staining was performed to determine the pathological structure of the wound. The mRNA expression of inflammatory factors in the wound was measured by real-time polymerase chain reaction （Real-time PCR）， and the expression of proteins related to the SIRT1/ nuclear factor （NF）-κB and PI3K/protein kinase B （Akt） signaling pathways in the wound was tested by Western blot. ResultCompared with the sham group， the percentage of postoperative wound area of rats in both low-dose and high-dose astragaloside groups gradually decreased with time， and the efficacy of the high-dose astragaloside group was better. Compared with the Con group， the fluorescence intensity of COL1A1 in wound tissue of the sham group decreased， while the expression of α-SMA increased. The epithelial tissue was severely damaged， with an increase in the thickness， and a large number of inflammatory cells were seen in the infiltration. The mRNA expression of tumor necrosis factor （TNF）-α， interleukin （IL）-1β， IL-6， and inducible nitric oxide synthase （iNOS） was elevated. The protein expression of NF-κB p65， p-PI3K/PI3K， and p-Akt/Akt was elevated， while SIRT1 expression was decreased （P<0.05）. Compared with the sham group， the fluorescence intensity of COL1A1 and α-SMA increased after astragaloside treatment. The number of epithelial cells increased， and the thickness decreased. The inflammatory cells decreased， and the amount of collagen increased. The mRNA expression of TNF-α， IL-1β， IL-6， and iNOS was decreased， and the protein expression of NF-κB p65， p-PI3K/PI3K， and p-Akt/Akt was decreased. SIRT1 was elevated， and the effect was better in the high-dose astragaloside group （P<0.05）. Compared with the high-dose astragaloside group， inhibition of the PI3K/Akt and SIRT1 pathways by LY294002 and EX527 prevented the therapeutic efficacy of astragaloside on chronic non-healing wounds. ConclusionThe topical application of astragaloside significantly promotes the healing of chronic non-healing wounds in rats， and the mechanism may be related to the activation of the PI3K/Akt pathway and the SIRT1/NF-κB pathway.

Inhibition of Oxidative Stress of Sciatic Nerve in Diabetic Rats by Tangbikang Granules Regulating AMPK/PGC-1α/SIRT3 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effect of Tangbikang granules on oxidative stress of sciatic nerve in diabetic rats by regulating adenylate activated protein kinase/peroxisome proliferator-activated receptor γ coactivator-1α/mitochondrial Sirtuins 3 （AMPK/PGC-1α/SIRT3） signaling pathway. MethodThe spontaneous obesity type 2 diabetes model was established using ZDF rats. After modeling， they were randomly divided into high， medium， and low dose Tangbikang granule groups （2.5， 1.25， 0.625 g·kg-1·d-1） and lipoic acid group （0.026 8 g·kg-1·d-1）， and the normal group was set up. The rats were administered continuously for 12 weeks after modeling. The blood glucose of rats was detected before intervention and at 4， 8， 12 weeks after intervention. At the 12th week， motor nerve conduction velocity （MNCV）， sensory nerve conduction velocity （SNCV）， nerve blood flow velocity， mechanical pain threshold， and thermal pain threshold were detected. The sciatic nerve was taken for hematoxylin-eosin （HE） staining to observe the tissue morphology. The ultrastructure of the sciatic nerve was observed by transmission electron microscope. The expression levels of superoxide dismutase （SOD）， malondialdehyde （MDA）， interleukin-1β （IL-1β）， and tumor necrosis factor-α （TNF-α） in sciatic nerve were determined by enzyme-related immunosorbent assay （ELISA）. The mRNA expressions of AMPKα， AMPKβ， PGC-1α， and SIRT3 in sciatic nerve were determined by real-time polymerase chain reaction （Real-time PCR）. ResultCompared with the normal group， fasting blood glucose in the model group was increased at each time point （P<0.01）. The mechanical pain threshold was decreased （P<0.05）， and the incubation time of the hot plate was extended （P<0.01）. MNCV， SNCV， and nerve blood flow velocity decreased （P<0.05）. The expression level of SOD was decreased （P<0.01）. The expression levels of MDA， IL-1β， and TNF-α were increased （P<0.01）. The mRNA expression levels of AMPKα， AMPKβ， PGC-1α， and SIRT3 were decreased （P<0.01）. The structure of sciatic nerve fibers in the model group was loose， and the arrangement was disordered. The demyelination change was obvious. Compared with the model group， the fasting blood glucose of rats in the high dose Tangbikang granule group was decreased after the intervention of eight weeks and 12 weeks （P<0.01）. The mechanical pain threshold increased （P<0.05）. The incubation time of the hot plate was shortened （P<0.01）. MNCV， SNCV， and Flux increased （P<0.05）. The expression level of SOD was increased （P<0.01）. The expression levels of MDA， IL-1β， and TNF-α were decreased （P<0.01）. The mRNA expression levels of AMPKα， AMPKβ， PGC-1α， and SIRT3 were increased （P<0.01）. The sciatic nerve fibers in the high-dose Tangbikang granule group were tighter and more neatly arranged， with only a few demyelinating changes. The high， medium， and low dose Tangbikang granule groups showed a significant dose-effect trend. ConclusionTangbikang granules may improve sciatic nerve function in diabetic rats by regulating AMPK/PGC-1α/SIRT3 signaling pathway partly to inhibit oxidative stress.

Effect of Linggui Zhugantang on Mitochondrial Fusion-fission and Sirt3/ AMPK Signaling Pathway in Chronic Heart Failure Rats After Myocardial Infarction / 中国实验方剂学杂志

ObjectiveTo investigate the effects of Linggui Zhugantang on mitochondrial fission and fusion and silencing information regulator 3（Sirt3）/adenosine monophosphate dependent protein kinase （AMPK） signaling pathway in chronic heart failure （CHF） rats after myocardial infarction （MI）. MethodSD rats randomly divide into sham operation group （normal saline ，thread only without ligature）， model group （normal saline， ligation of the left anterior descending coronary artery proximal to the heart）， Linggui Zhugantang group （4.8 g·kg-1） and Captopril group （0.002 57 g·kg-1）， with 10 rats in each group. Administere drug continuously for 28 days. Echocardiography detected cardiac function parameters. Hematoxylin eosin （HE） staining observed the pathological changes of the heart. Immunofluorescence detected the levels of reactive oxygen species （ROS）. JC-1 detect mitochondrial membrane potential. Colorimetry measure adenosine triphosphate （ATP）， superoxide dismutase （SOD）， malondialdehyde （MDA）， mitochondrial respiratory chain complex activity （Ⅰ-Ⅳ）. TdT-mediated dUTP nick end labeling （TUNEL） staining detected the apoptosis rate of myocardial tissue. Western blot detected protein expression levels of Sirt3， phosphorylated AMPK （p-AMPK）， phosphorylated dynamic-related protein 1（p-Drp1）， mitochondrial fission protein 1（Fis1）， mitochondrial fission factor （MFF）， optic atrophy protein 1（OPA1）. ResultCompared to the sham group， the left ventricular end diastolic diameter （LVIDd） and left ventricular end systolic diameter （LVIDs） were significantly increased in model group （P<0.01）， while the left ventricular short axis shortening rate （LVFS） and left ventricular ejection fraction （LVEF） were significantly decreased （P<0.01）. There were inflammatory cell infiltration and obvious pathological injury in myocardial tissue. ROS， MDA levels and myocardial cell apoptosis rate were significantly increased （P<0.01）， SOD level， ATP content， and membrane potential were significantly decreased （P<0.01）. The activity of mitochondrial respiratory chain complexes （Ⅰ-Ⅳ） was significantly decreased （P<0.01）. Levels of p-Drp1， Fis1， MFF proteins were significantly up-regulated （P<0.01）， while Sirt3， p-AMPK， OPA1 proteins level were significantly down-regulated （P<0.01）. Compared with model group， LVIDd and LVIDs were significantly decreased （P<0.01）， LVEF and LVFS were significantly increased （P<0.01）. Inflammatory cell infiltration and pathological damage of myocardial tissue were significantly relieved. ROS， MDA levels and myocardial cell apoptosis rate were significantly decreased in Linggui Zhugantang group and Captopril group （P<0.01）， SOD level， ATP content， and membrane potential significantly increased （P<0.01）. The activity of mitochondrial respiratory chain complexes （Ⅰ-Ⅳ） increased significantly （P<0.01），and p-Drp1， Fis1， MFF protein levels were significantly down-regulated （P<0.01）， Sirt3， p-AMPK， OPA1 protein were significantly up-regulated （P<0.01）. ConclusionLinggui Zhugantang can alleviate oxidative stress and apoptosis damage of myocardial cells， maintain mitochondrial function stability， and its effect may be related to mitochondrial mitosis fusion and Sirt3/AMPK signaling pathway.

Gegen Qinliantang Ameliorates Glucose and Lipid Metabolism Disorders in Rat Model of Catch-up Growth by Regulating SIRT1/PGC1α/Nrf1 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effect of Gegen Qinliantang on glucose and lipid metabolism in the rat model of catch-up growth （CUG） induced by a high-fat diet and the underlying mechanism. MethodA total of 60 SD rats were randomized into a normal control group （n=18） and a modeling group （n=42）. The rat model of CUG was established with a restricted diet followed by a high-fat diet， and the changes of general status and body weight were observed. The levels of fasting blood glucose （FBG）， fasting insulin （FINS）， triglyceride （TG）， and total cholesterol （TC） were measured in 6 rats in each group at the end of the 4th and 8th week， respectively. The homeostasis model assessment of insulin resistance index （HOMA-IR） was calculated， and the insulin sensitivity and body composition changes of CUG rats were evaluated. The successfully modeled rats were assigned into 6 groups： normal control， model， high-， medium-， and low-dose Gegen Qinliantang （2.5， 5， 10 g·kg-1）， and pioglitazone （3.125 mg·kg-1）. The rats were administrated with corresponding drugs by gavage for 6 weeks， and the normal control group and model group were administrated with the same amount of normal saline. During the experiment period， the changes of body weight were recorded， and the FBG， FINS， HOMA-IR， TG， and TC were determined at the end of the experiment. Hematoxylin-eosin （HE） staining was employed to observe the pathological changes of skeletal muscle in rats. The levels of reactive oxygen species （ROS） and malondialdehyde （MDA） in the skeletal muscle were measured strictly according to the manuals of the reagent kits. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was performed to measure the mRNA levels of silencing information regulator 1 （SIRT1）， peroxisome proliferator-activated receptor-gamma coactivator1α （PGC1α）， and nuclear respiratory factor 1 （Nrf1） in the skeletal muscle. Western blot and immunohistochemistry were employed to assess the expression of SIRT1， PGC1α， and Nrf1 in the skeletal muscle. ResultCompared with the normal control group， the model group presented elevated levels of FBG， FINS， TG， and TC （P<0.05， P<0.01）， increased HOMA-IR （P<0.01）， increased diameter of muscle fibers and adipocytes between muscle cells in the skeletal muscle， rising levels of ROS and MDA in the skeletal muscle （P<0.01）， and down-regulated mRNA and protein levels of SIRT1， PGC1α， and Nrf1 （P<0.05， P<0.01）. Compared with the model group， Gegen Qinliantang （especially the medium and high doses） and pioglitazone decreased the body weight， FINS， HOMA-IR， and TG （P<0.05， P<0.01） and reduced interstitial components such as intermuscular fat in the skeletal muscles and the diameter of muscle fibers. Furthermore， the drugs lowerd the levels of ROS and MDA （P<0.05， P<0.01） and up-regulated the mRNA and protein levels of SIRT1， PGC1α， and Nrf1 （P<0.05， P<0.01） in the skeletal muscle. ConclusionGegen Qinliantang can ameliorate the glucose and lipid metabolism disorders and insulin resistance in CUG rats by regulating the SIRT1/PGC1α/Nrf1 signaling pathway.

Role of sirtuin 5 in cardiovascular diseases / 中国药理学通报

Cardiovascular diseases ( CVDs ) are the leading cause of death worldwide and pose a serious threat to human health. Silent information regulator 5 ( SIRT5 ) , which is widely distributed in cardiac myocytes, vascular smooth muscle cells and endothelial cells,as a novel deacylation-modifying enzyme,plays an important role in CVDs through deacetylation, desuccinylation and demalonylation. This review summarizes the pathophysiolog-ical mechanism of SIRT5 from the aspects of energy metabolism, regulation of inflammatory response and oxidative stress, apart from the role of SIRT5 in CVDs such as myocardial infarction, myocardial hypertrophy, arrhythmia, atherosclerosis and heart failure. This review also figures out the current research progress of SIRT5 -related inhibitors and agonists, so as to provide strategies for targeting SIRT5 to prevent and treat CVDs.

Role of SIRTl/Nrf2/HO-l pathway in attenuation of learning and memory impairment by sevoflurane postcondition in a mouse model of hemorrhagic shock and resuscitation / 中国药理学通报

Aim To explore the role of SIRT1/Nrf2 / HO-1 in alleviating the cognitive function impairment by sevoflurane treatment in a mouse model of postoperative cerebral reperfusion. Methods C57BL/6J mice were randomly divided into five groups: sham operation group, hemorrhagic shock reperfusion group, sevoflurane postconditioning group, sevoflurane postcondition-ing + SIRT1 inhibitor group and sevoflurane postconditioning + Nrf2 inhibitor group. Mice were subjected to Morris water maze test after cerebral ischemia reperfusion. The ATP, superoxide dismutase (SOD), ROS and MDA contents in tissue of mice were detected. SIRT1, Nrf2 and HO-1 proteins in tissue were detected by Western blot. Results After hemorrhagic shock, the learning and memory ability of mice was reduced.ATP and SOD concentration in hippocampus was reduced , MDA and ROS concentration increased, and the SIRT, Nrf2 and HO-1 concentration was reduced. Sevoflurane improved the cognitive dysfunction and oxi-dative damage in postoperative mice, and the neuro-protective effect of sevoflurane on hemorrhagic shock and resuscitation mice was weakened followed with SIRT1 and Nrf2 inhibitors. Conclusion Sevoflurane probably alleviates the oxidative reaction damage and cognitive impairment caused by cerebral reperfusion in mice through SIRT1/Nrf2/H0-1 pathway.

Improvement mechanism of proanthocyanidins on gentamicin-induced acute kidney injury of rats through SIRT1/AMPK signaling pathway / 中国药房

OBJECTIVE To explore the improvement mechanism of proanthocyanidins on acute kidney injury （AKI） induced by gentamicin in rats. METHODS Gentamicin sulfate was injected intraperitoneally to construct the AKI rat model； the model rats were randomly divided into model control group， benazepril hydrochloride 5 mg/kg group （positive control）， proanthocyanidins 50 mg/kg group， proanthocyanidins 100 mg/kg group， and proanthocyanidins 200 mg/kg group， with 10 rats in each group； in addition， 10 normal rats were selected to be treated as the normal control group. The rats in each administration group were given corresponding liquid intragastrically， and the normal control group and model control group were given equal volumes of normal saline intragastrically， once a day， for 28 consecutive days. After the last administration， the levels of serum creatinine （SCr）， blood urea nitrogen （BUN）， malondialdehyde （MDA）， superoxide dismutase （SOD）， glutathione peroxidase （GSH-Px） and 24 h urinary protein （UP） were detected； the renal index was calculated； the pathological changes of renal tissue were observed and the pathological score was calculated； the apoptotic rate of cells in renal tissue and the expression levels of Caspase-3 and Bcl-2 associated X protein （Bax）， as well as the phosphorylation levels of silent information regulator of transcription 1 （SIRT1） and AMP-activated protein kinase （AMPK） were detected. RESULTS Compared with the model control group， the levels of SCr， BUN， UP and MDA， the renal index， the pathological score of renal tissue， the apoptotic rate of cells in renal tissue， the protein expression levels of Caspase-3 and Bax in renal tissue of rats in each administration group were decreased significantly； SOD and GSH-Px levels， phosphorylation levels of SIRT1 and AMPK protein were increased significantly （P＜0.05）， and the effect of proanthocyanidins was in a dose-dependent manner （P＜0.05）. There were no significant differences in the above indexes between proanthocyanidins 200 mg/kg group and benazepril hydrochloride 5 mg/kg group （P＞0.05）. CONCLUSIONS The improvement effect of proanthocyanidins on AKI rats may be related to the activation of SIRT1/AMPK signaling pathway to inhibit oxidative stress.

Neuroprotective effect of sodium aescinate on rats with Parkinson’s disease by regulating SIRT1/NF-κB signaling pathway / 中国药房

OBJECTIVE To explore the neuroprotective effect of sodium aescinate on rats with Parkinson’s disease by regulating the silent information regulator 1 （SIRT1）/nuclear factor-κB （NF-κB） signaling pathway. METHODS The Parkinson’s disease rat model was constructed by using 6-hydroxydopamine injection method. Forty-eight rats successfully modeled were randomly divided into model group， sodium aescinate low-dose group （1.8 mg/kg）， sodium aescinate high-dose group （3.6 mg/kg）， sodium aescinate+EX527 （sodium aescinate 3.6 mg/kg+SIRT1 inhibitor EX527 5 mg/kg） group， with 12 rats in each group. Another 12 healthy rats were selected as the sham operation group. Each group was injected with the corresponding drug solution intraperitoneally， once a day， for 21 consecutive days. Twenty-four hours after the end of the last administration， the motor and cognitive functions of rats were detected， and the morphology of neurons in the substantia nigra and CA1 region of hippocampal tissue were observed. The content of dopamine （DA） in the nigrostriatal and the expression levels of tyrosine hydroxylase （TH） and α-synuclein （α-Syn） in the substantia nigra were detected. The serum levels of pro-inflammatory factor [interleukin-6 （IL-6）， IL-18]， anti-inflammatory factor （IL-10）， and the expression levels of SIRT1， phosphorylated NF-κB p65 （p-NF-κB p65） and NF- κB p65 protein in nigrostriatal were detected. RESULTS Compared with sham operation group， the neurons in the substantia nigra and CA1 region of hippocampal tissue were seriously damaged in model group； the number of rotations， escape latency， the expression levels of α-Syn in substantia nigra， the levels of serum pro-inflammatory factors， the relative expression ratio of p-NF- κB p65 and NF-κB p65 protein in nigrostriatal were increased or prolonged significantly （P＜0.05）； the target quadrant residence time， the content of DA in nigrostriatal， the expression level of TH in substantia nigra， the serum level of anti-inflammatory factor， and the expression level of SIRT1 protein in substantia nigra striatum were significantly decreased or shortened （P＜0.05）. Compared with model group， the damage degrees of neuron in sodium aescinate groups were alleviated， and the quantitative indicators were significantly improved， which were more significant in the high-dose group （P＜0.05）； EX527 could reverse the improvement effect of high-dose sodium aescinate （P＜0.05）. CONCLUSIONS Sodium aescinate can inhibit the activation of NF-κB signal by up-regulating the protein expression of SIRT1， thereby reducing the neuroinflammation of rats with Parkinson’s disease， improving the motor and cognitive dysfunctions， and finally playing a neuroprotective role.

Protective effect of SRT1720 on acute liver injury induced by acetaminophen in mice and its mechanism / 中国生物制品学杂志

@#Objective To evaluate the protective effect of the activator of silent information regulator 2-related enzymes 1(SIRT1),SRT1720,on liver injury induced by acetaminophen(APAP)in mice and explore its mechanism. Methods Forty male C57BL/6J mice were randomly divided into normal control group,SRT1720 treatment group,APAP treatment group,and APAP + SRT1720 treatment group,with 10 mice in each group. Mice in SRT1720 and APAP + SRT1720 groups were given SRT1720(30 mg/kg body mass)by intragastric administration,while normal saline of equal volume was given by intragastric administration in control and APAP groups,once a day for 5 days;On the 6th day,mice in APAP and APAP + SRT1720 groups were injected i. p. with APAP(325 mg/kg body mass),while those in control and SRT1720 groups with equal volume of normal saline. After 24 hours,the peripheral blood was taken and the serum was separated,which were detected for the levels of alanine aminotransferase(ALT)and aspartate aminotransferase(AST)by the corresponding kits;The liver tissue of mice was taken aseptically,observed for the pathological changes by HE staining,detected for the mRNA transcription levels of GRP78,PERK,eIF2 α,ATF4 and CHOP genes related to PERK-eIF2α-CHOP signaling pathway by RT-qPCR and detected for the relative expression levels of these corresponding proteins and Caspase12 protein by Western blot. Results Compared with normal control group,the serum ALT and AST levels of mice in APAP group significantly increased(t = 55. 21 and34. 29 respectively,each P < 0. 01);significant necrosis of hepatocytes was observed in liver tissue,the structure of hepatic lobules changed significantly,and the swelling and deformation of hepatocytes in some areas were serious;the mRNA transcription and relative protein expression levels of GRP78,PERK,eIFα,ATF4 and CHOP genes increased significantly(t = 9. 85～33. 89,each P < 0. 05)and the relative expression level of Caspase12 protein increased significantly(t = 11. 78,P < 0. 01). Compared with APAP group,the serum ALT and AST levels of mice in APAP + SRT1720 group decreased significantly(t = 42. 92 and 18. 02 respectively,each P < 0. 01);the degree of hepatocyte injury was obviously reduced and the number of swollen and deformed cells also significantly decreased;the mRNA transcription and relative protein expression levels of GRP78,PERK,eIF2α,ATF4 and CHOP decreased significantly(t = 6. 19～22. 43,each P < 0. 05)and the expression level of Caspase12 protein showed no significant decrease(t = 0. 34,P > 0. 05). Conclusion SRT1720improved APAP-induced liver injury in mice,possibly by inhibiting PERK-eIF2α-CHOP signaling pathway in endoplasmic reticulum stress(ERS).

SIRT1 is transcriptionally repressed by YY1 and suppresses ferroptosis in rheumatoid arthritis

Abstract Background Sirtuin 1 (SIRT1) is reported downregulated in rheumatoid arthritis (RA), and the protective effects of SIRT1 on tissue damage and organ failure may be related to cellular ferroptosis. However, the exact mechanism by which SIRT1 regulates RA remains unclear. Methods Quantitative real-time PCR (qPCR) and western blot assays were performed to explore the expressions of SIRT1 and Yin Yang 1 (YY1). CCK-8 assay was used for cytoactive detection. The interaction between SIRT1 and YY1 was validated by dual-luciferase reporter gene assay and chromatin immunoprecipitation (ChIP). DCFH-DA assay and iron assay were applied to detect the reactive oxygen species (ROS) and iron ion levels. Results In the serum of RA patients, SIRT1 was downregulated, but YY1 was upregulated. In LPS-induced synoviocytes, SIRT1 could increase cell viability and decrease ROS and iron levels. Mechanistically, YY1 downregulated the expression of SIRT1 by inhibiting its transcription. YY1 overexpression partly revised the effects of SIRT1 on ferroptosis in synoviocytes. Conclusion SIRT1 is transcriptionally repressed by YY1 and inhibits the ferroptosis of synoviocytes induced by LPS, so as to relieve the pathological process of RA. Therefore, SIRT1 might be a new diagnosis and therapeutic target of RA. Highlights Combining SIRT1 with synoviocytes ferroptosis in rheumatoid arthritis for the first time. The transcription factor YY1 combined to the SIRT1 promoter in synovial cells and inhibited its expression and functional roles. The inhibition of SIRT1 with YY1 decreased the ferroptosis in synoviocytes.

Protective Effect of Combined Metoprolol and Atractylenolide I in Rats with Acute Myocardial Infarction via Modulation of the SIRT3/ß-CATENIN/PPAR-γ Signaling Pathway

ABSTRACT Herein, we examined the protective effect of metoprolol combined with atractylenolide I (Atr I) in acute myocardial infarction (AMI) by regulating the SIRT3 (silent information regulator 3)/ß-catenin/peroxisome proliferator-activated receptor gamma (PPAR-γ) signaling pathway. Briefly, 50 rats were randomly divided into the sham operation, model, metoprolol, Atr I, and combination metoprolol with Atr I groups (combined treatment group). The AMI model was established by ligating the left anterior descending coronary artery. After treatment, infarct size, histopathological changes, and cell apoptosis were examined using 2,3,5-triphenyltetrazolium chloride staining, hematoxylin-eosin staining, and the TUNEL assay. The left ventricular ejection fraction (LVEF), left ventricular fraction shortening (LVFS), and left ventricular mass index (LVMI) were detected by echocardiography. Endothelin-1 (ET-1), nitric oxide (NO), tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6) levels were detected using enzyme-linked immunosorbent assays. Furthermore, we measured lactate dehydrogenase (LDH), creatine kinase (CK) isoenzyme (CK-MB), and CK levels. Western blotting was performed to determine the expression of SIRT3, ß-catenin, and PPAR-γ. Herein, the combined treatment group exhibited increased levels of LVEF, LVFS, and NO, whereas LVMI, ET-1, TNF-α, IL-6, LDH, CK-MB, and CK levels were decreased. Importantly, the underlying mechanism may afford protection against AMI by increasing the expression levels of SIRT3, ß-catenin, and PPAR-γ

Pre-treatment of the beta3-adrenergic receptor agonist BRL37344 reduces in vivo myocardial ischemia/reperfusion injury by improving AMPK and SIRT1 activity and by suppressing mTOR and p70S6K signaling pathways

Abstract This study aimed to investigate the role and signaling pathways of β3-AR in myocardial ischemia/reperfusion (I/R) injury, which is one of the leading causes of death worldwide. 47 male rats were randomly divided into two main groups to evaluate infarct size and molecular parameters. Rats in both groups were randomly divided into 4 groups. Control (sham), I/R (30 min ischemia/120 min reperfusion), BRL37344 (BRL) (A) (5 µg/kg single-dose pre-treatment (preT) before I/R) and BRL (B) (5 µg/kg/day preT for 10 days before I/R). Infarct size was determined with triphenyltetrazolium chloride staining and analyzed with ImageJ program. The levels of AMPK, SIRT1, mTOR, and p70SK6 responsible for cellular energy and autophagy were evaluated by western blot. Infarct size increased in the I/R group (44.84 ± 1.47%) and reduced in the single-dose and 10-day BRL-treated groups (32.22 ± 1.57%, 29.65 ± 0.55%; respectively). AMPK and SIRT1 levels were decreased by I/R but improved in the treatment groups. While mTOR and p70S6K levels increased in the I/R group, they decreased with BRL preT. BRL preT protects the heart against I/R injury. These beneficial effects are mediated in part by activation of AMPK and SIRT1, inhibition of mTOR and p70S6K, and consequently protected autophagy.

Inhibitory Effect of Paeonol on Vascular Smooth Muscle Cell Senescence by Reducing SIRT6/PARP1-mediated DNA Damage / 中国实验方剂学杂志

ObjectiveTo investigate whether the effects of paeonol （Pae） on angiotensin Ⅱ （AngⅡ）-induced senescence in vascular smooth muscle cells （VSMCs） were related to angiotensinogen of silencing regulatory information factor 6 （SIRT6）/adenosine diphosphate ribose polymerase 1 （PARP1） signaling pathway in VSMCs. MethodThe model of VSMC-stress aging induced by AngⅡ （100 nmol·L-1） was established. The rats were divided into normal group， model group， low， medium， and high-concentration Pae groups （30， 60， 120 μmol·L-1）. The positive rate of cell senescence was detected by SA-β-Gal staining， the ability of cell proliferation was detected by the cell counting kit-8 （CCK-8） method， the expression of SIRT6， PARP1， p16， p21， p53， proliferating cell nuclear antigen （PCNA）， deoxyribonucleic acid （DNA）-damaged protein γ-H2AX was detected by Western blot， and VSMC proliferation was detected by EdU staining. The silenced VSMCs were prepared by siRNA-SIRT6 transfection， and the protein expressions of SIRT6， PARP1， p16， and γ-H2AX in VSMCs silenced by SIRT6 were observed. ResultThe results of SA-β-Gal staining showed that the senescence positive rate of SA-β-Gal staining in the model group was higher than that in the normal group （P<0.01）， and the positive rate of SA-β-Gal staining in the Pae group was significantly lower than that in the model group （P<0.05， P<0.01）. The results of Western blot showed that as compared with the normal group， the expression of PCNA， SIRT6， and PARP1 in the model group was down-regulated， and the expression of aging-related proteins p16， p21， p53， and γ-H2AX was up-regulated in the model group （P<0.05， P<0.01）. Compared with the model group， Pae promoted the protein expression of PCNA， SIRT6， and PARP1 and inhibited the protein expression of p16， p21， p53， and γ-H2AX in a dose-dependent manner （P<0.05， P<0.01）. The results of EdU staining showed that the number of EdU positive cells in the model group was lower than that in the normal group （P<0.01）， and the number of EdU positive cells in Pae groups was significantly higher than that in the model group （P<0.05， P<0.01）. After SIRT6 silencing， the effects of Pae on promoting SIRT6 and PARP1 and inhibiting P16 were reversed （P<0.05， P<0.01）. In addition， the addition of SIRT6 inhibitor （IN-1） promoted the occurrence of cell senescence induced by AngⅡ （P<0.05， P<0.01）. ConclusionPae can effectively inhibit the aging of VSMCs， and its mechanism may be related to the regulation of SIRT6/PARP1 signal pathway.

Construction of a prediction model for postoperative prognosis in patients with resectable cholangiocarcinoma based on SIRT2 expression / 浙江大学学报·医学版

OBJECTIVES@#To develop a prediction model for postoperative prognosis in patients with cholangiocarcinoma (CCA) based on the expression of silence information regulator 2 (SIRT2).@*METHODS@#The differential expression of SIRT2 between CCA and normal tissues was analyzed using TCGA and GEO databases. Gene set enrichment analysis (GSEA) was used to explore potential mechanisms of SIRT2 in CCA. The expression of SIRT2 protein in CCA tissues and normal tissues (including 44 pairs of specimens) was also detected by immunohistochemistry (IHC) staining in 89 resectable CCA patients who underwent surgical treatment in The First Affiliated Hospital of Bengbu Medical College between January 2016 and December 2021. The relationship between SIRT2 expression and clinicopathological characteristics and prognosis of CCA patients was analyzed. A survival prediction model for patients with resectable CCA was constructed with COX regression results, the calibration curve and the time-dependent receiver operating characteristic curve (ROC) were used to evaluate the performance of the constructed model, and the predictive power between this model and the AJCC/TNM staging system (8th Edition) was compared.@*RESULTS@#SIRT2 mRNA was overexpressed in CCA tissues as shown in TCGA and GEO databases. IHC staining showed that SIRT2 protein expression in CCA tissues was significantly higher than that in adjacent non-tumor tissues. GSEA results showed that elevated SIRT2 expression may be involved in multiple metabolism-related signaling pathway, such as fatty acid metabolism, oxidative phosphorylation, amino acid metabolism, etc. SIRT2 expression level was related to serum triglycerides level, tumor size and lymph node metastasis (all P<0.05). The survival analysis results showed that the patients with higher SIRT2 expression had a significant lower overall survival (OS) than patients with lower SIRT2 expression (P<0.05). Univariate COX regression analysis suggested that pathological differentiation, clinical stage, postoperative treatment and SIRT2 expression level were associated with the prognosis of CCA patients (all P<0.05). Multivariate regression analysis confirmed that clinical stage and SIRT2 expression level were independent predictors of OS in postoperative CCA patients (both P<0.05). A nomogram based on SIRT2 for prediction of survival in postoperative CCA patients was constructed. The C-index of the model was 0.675, and the area under the time-dependent ROC curve (AUC) for predicting survival in the first, second, and third years was 0.879, 0.778, and 0.953, respectively, which were superior to those of AJCC/TNM staging system (8th Edition).@*CONCLUSIONS@#SIRT2 is highly expressed in CCA tissues, which is associated with poor prognosis in patients with resectable CCA. The nomogram developed based on SIRT2 may have better predictive power than the AJCC/TNM staging system (8th Edition) in prediction of survival of postoperative CCA patients.

Liraglutide ameliorates high glucose-induced oxidative stress injury in rat H9c2 cells through modulation of SIRT1 / 中华内分泌代谢杂志

Objective:To investigate the effect of liraglutide(LRG) on high glucose-induced oxidative stress injury in(H9c2) cardiomyocytes and its underlying mechanisms.Methods:A high glucose treatment was applied to H9c2 cells for 24 hours to establish an in vitro model of myocardial cell injury. Different concentrations of liraglutide(10, 100, 1000 nmol/L) were administered for intervention. Cell viability was evaluated using the CCK-8 assay, and changes in cell morphology were observed under an inverted microscope. After 24 hours of liraglutide(100 nmol/L) intervention following high glucose treatment, the levels of lactate dehydrogenase(LDH), superoxide dismutase(SOD), and malondialdehyde(MDA) in the cell supernatant were measured. RT-PCR and Western blotting were used to detect the mRNA and protein levels of silent information regulator factor 1(SIRT1) and forkhead box protein O1(FOXO1). Western blotting was also used to assess the acetylation level of FOXO1 protein. Small interfering RNA(siRNA) technology was employed to silence SIRT1 in H9c2 cells to confirm its role in the study. Results:Compared to the control group, the high glucose group showed decreased cell viability, cell structure damage, increased levels of LDH and MDA in the cell supernatant, decreased SOD levels, aggravated oxidative stress, decreased SIRT1 expression, and increased acetylation level of FOXO1(all P<0.05). Compared to the high glucose group, liraglutide intervention resulted in increased cell viability, improved cardiac cell morphology, reduced oxidative stress levels, increased SIRT1 expression, and decreased acetylation level of FOXO1(all P<0.05). When SIRT1 was downregulated, the protective effects of liraglutide were weakened(all P<0.05). Conclusions:Liraglutide has a protective effect against high glucose-induced oxidative stress injury in H9c2 cells, which may be associated with the upregulation of SIRT1 expression.

Sevoflurane inhibits proliferation and invasion of breast cancer cells by promoting SIRT2 expression and inducing ACLY deacetylation / 中华内分泌外科杂志

Objective:To explore the effects of sevoflurane (Sev) on proliferation and invasion of breast cancer cells.Methods:Normal human breast epithelial cell line MCF10A and human breast cancer cell line MCF7 were purchased. The expression level of sirtuin 2 (SIRT2) , ATP citrate lyase (ACLY) in breast cancer cells and acetylation level of ACLY were measured. Breast cancer cells were divided into the following groups: Control group, 2% SEV group, 4% SEV group, si-NC group, si-SIRT2 group, 4% SEV+si-NC group, 4% SEV+si-SIRT2 group, SIRT2 group, SIRT2+ACLY-WT group, SIRT2+ACLY-3KQ group, SIRT2+ACLY-3KQ+4% SEV group, si-ACLY group, si-ACLY+ACLY-WT group, si-ACLY+ACLY-3KQ group. MTT and Transwell assay were used to detect cell proliferation and invasion.Results:Compared with MCF-10A cells (1.00±0.15) , SIRT2 was low expressed in Control group cells (0.43±0.03) ( q=11.98, P<0.001) , SEV could induce the expression of SIRT2 ( F=88.71, P<0.001) . In addition, ACLY and ACLY-3K acetylation level were up-regulated in breast cancer cells (all P<0.05) . Knockdown of SIRT2 or overexpression of ACLY and ACLY-3KQ could promote the proliferation and invasion of MCF7 cells (all P<0.05) , while SEV, overexpression of SIRT2 or knockdown of ACLY showed the opposite effects (all P<0.05) . Conclusion:Sev may inhibit the proliferation and invasion of breast cancer cells through SIRT2, which may be related to the regulation of ACLY deacetylation.

Puerarin alleviates lipopolysaccharide-induced acute kidney injury in mice by modulating the SIRT1/NF-κB pathway / 南方医科大学学报

OBJECTIVE@#To investigate the role of the SIRT1/NF-κB pathway in mediating the effect of puerarin against lipopolysaccharide (LPS)-induced acute kidney injury (AKI).@*METHODS@#Fifteen BALB/C mice were randomized into control group, LPS group and puerarin treatment group, and in the latter two groups, the mice were given an intraperitoneal injection of LPS (5 mg/kg), followed by daily injection of normal saline for 3 days or injection of puerarin (25 mg/kg) given 1 h later and then on a daily basis for 3 days. On day 5 after modeling, the kidney tissues were taken for histological observation and detection of cell apoptosis. The renal function indexes including urea nitrogen (BUN), serum creatinine (Scr) and kidney injury molecule 1 (KIM-1) and the levels of tumor necrosis factor (TNF-α) and interleukin 1β (IL-1β) were measured, and the expressions of SIRT1 and NF-κB-p65(acetyl K310) in the renal tissues were detected.@*RESULTS@#Intraperitoneal injection of LPS caused obvious glomerular capillary dilatation, hyperemia, renal interstitial edema, and renal tubular epithelial cell swelling and deformation in the mice. The mouse models of LPS-induced AKI also showed significantly increased renal tubular injury score and renal cell apoptosis (P < 0.01) with increased serum levels of BUN, Scr, KIM-1, TNF-α and IL-1β (P < 0.01), enhanced renal expressions of TNF-α, IL-1β and NF-κB p65(acetyl K310) (P < 0.01) and lowered renal expression of SIRT1 (P < 0.05). Treatment with puerarin effectively alleviated LPS-induced renal interstitial edema and renal tubular epithelial cell shedding, lowered renal tubular injury score (P < 0.01) and renal cell apoptosis rate (P < 0.01), and decreased serum levels of BUN, Scr, KIM, TNF-α and IL-1β (P < 0.01). Puerarin treatment significantly reduced TNF-α, IL-1β and NF-κB p65 (acetyl K310) expression in the renal tissue (P < 0.05) and increased SIRT1 expression by 17% (P < 0.05) in the mouse models.@*CONCLUSION@#Puerarin can effectively alleviate LPS-induced AKI in mice possibly by modulating the SIRT1/NF-κB signaling pathway.

Effects of SIRT1 on morphine-induced conditioned place preference in rats through CREB/BDNF pathway / 中国药理学通报

Aim To investigate the effect of microinjection of EX527, a selective SIRT1 antagonist, into the ventrolateral orbital cortex (VLO) on morphine-induced conditioned place preference (CPP), and to explore the role of CREB/BDNF in it. Methods The cannulas were implanted bilaterally in the VLO of rats by brain stereotaxis surgery, and the model of morphine-induced CPP was established. The behavioral experiment consisted of four stages:habituation (d 1), pre-test (d 2-4), conditioning training (d 5-14) and test (d 15). At the stage of conditioning training, EX527 (1 μL, 5 g·L

SuanZaoRen decoction amliorates mitochondrial dysfunction of APP/PS1 mice via activating AMPK/SIRT1/PGC-1α signaling pathway / 中国药理学通报

Aim To explore the effect of Suanzaoren decoction(SZRD) on mitochondrial dysfunction in AD model of APP/PS1 mice via AMPK/SIRT1/PGC-1α signaling pathway and to reveal the possible mechanism. Methods Thirty APP/PS1 mice were randomly divided into app /PS1 group, low-dose SZRD group(L-SZRD) and high-dose SZRD group(H-SZRD). Ten C57BL/6JNju mice were set as control group(WT). Morris water maze test was used to detect the learning and memory ability of mice. Thioflavin T staining was used to observe senile plaques hippocampus. Immunohistochemistry was performed to detect the expression level of Aβ in hippocampus. Transmission electron microscope was used to observe the mitochondrial morph hology in hippocampus. Kits were employed to detect the contents of ATP and ROS in hippocampus; Western blot was employed to detect the expression levels of AMPK, p-AMPKThrK172, SIRT1, PGC-1α, NRF1, NRF2 and TFAM in hippocampus. Results Compared to the APP/PS1 group, L-SZRD and H-SZRD induced mouse cognitive impairment, reduced the deposition of senile plaques, inhibited the expression of Aβ, improved the damage of mitochondrial structure, increased the content of ATP in the hippocampus, reduced the expression level of ROS in hippocampus and increased the expression of p-AMPK-ThrK172, SIRT1, PGC-1α, NRF1, NRF2, TFAM Conclusions SZRD could improve the cognitive impairment, senile plaque deposition and mitochondrial dysfunction of AD mice, and its mechanism may be involved in the up-regulation of the expression of AMPK/SIRT1/PGC-1α signaling pathway.Reduced the Deposition of Senile Plaques, Inhibited the Expression of Aβ, Improved The Damage of Mitochondric Structure, Increased the Content of At in TH. E hippocampus, Reduced the Expression level of Ros in Hippocampus and Increased The Expression of P-Ampk-Thrk172, SIRT1, SIRT1 PGC-1α, NRF1, NRF2, TFAM. Conclusions SZRD could improve the cognitive impairment, senile plaque deposition and mitochondrial dysfunction of AD mice, and its mechanism may be involved in the up-regulation of the expression of AMPK/SIRT1/PGC-1α signaling pathway.Reduced the Deposition of Senile Plaques, Inhibited the Expression of Aβ, Improved The Damage of Mitochondric Structure, Increased the Content of At in TH. E hippocampus, Reduced the Expression level of Ros in Hippocampus and Increased The Expression of P-Ampk-Thrk172, SIRT1, SIRT1 PGC-1α, NRF1, NRF2, TFAM. Conclusions SZRD could improve the cognitive impairment, senile plaque deposition and mitochondrial dysfunction of AD mice, and its mechanism may be involved in the up-regulation of the expression of AMPK/SIRT1/PGC-1α signaling pathway.Senile plaque deposition and mitochondrial dysfunction of AD mice, and its mechanism may be involved in the up-regulation of the expression of AMPK/SIRT1/PGC-1α signaling pathway.Senile plaque deposition and mitochondrial dysfunction of AD mice, and its mechanism may be involved in the up-regulation of the expression of AMPK/SIRT1/PGC-1α signaling pathway.