Effect and Mechanism of Total Saponins from Panax Japonicus on White Adipose Tissue Browning/Brown Adipose Tissue Activation in High-fat Diet-induced Mice / 中国实验方剂学杂志

ObjectiveTo investigate the effect and mechanism of total saponins from Panax japonicus （TSPJ） on white adipose tissue （WAT） browning/brown adipose tissue （BAT） activation in C57BL6/J male mice fed on a high-fat diet （HFD）. MethodThirty-two C57BL6/J male mice （8-week-old） were randomly divided into a normal group， a model group， a low-dose TSPJ group， and a high-dose TSPJ group. The mice in the low-dose and high-dose TSPJ groups were given TSPJ for four months by gavage at 25， 75 mg·kg-1·d-1， respectively， and those in the other groups were given 0.5% sodium carboxymethyl cellulose （CMC-Na） accordingly. After four months of feeding， all mice were placed at 4 ℃ for acute cold exposure， and the core body temperature was monitored. Subsequently， all mice were sacrificed， and BAT and inguinal WAT （iWAT） were separated rapidly to detect the corresponding indexes. Hematoxylin-eosin （HE） staining was used to observe the morphological changes in each group. The effect of TSPJ on the mRNA expression of uncoupling protein 1 （UCP1）， fatty acid-binding protein 4 （FABP4）， cytochrome C （CytC）， PR domain-containing protein 16 （PRDM16）， elongation of very long chain fatty acids protein 3 （ELOVL3）， peroxisome proliferator-activated receptor γ （PPARγ）， and peroxisome proliferator-activated receptor-γ coactivator-1α （PGC-1α） in iWAT and BAT was detected by Real-time polymerase chain reaction （Real-time PCR）. Western blot was also used to detect the protein expression of UCP1， PRDM16， PPARγ， and PGC-1α in BAT and iWAT of each group. The effect of TSPJ on UCP1 expression in BAT and iWAT was detected by immunohistochemistry. Result① Compared with the model group， TSPJ could decrease the body weight and proportions of iWAT and BAT in the HFD-induced mice （P<0.05， P<0.01）. ② The body temperature of mice in the model group decreased compared with that in the normal group in the acute cold exposure tolerance test （P<0.05）. The body temperature in the high-dose TSPJ group increased compared with that in the model group （P<0.01）. ③ Compared with the normal group， the model group showed increased adipocyte diameter in iWAT and BAT and decreased number of adipocytes per unit area. Compared with the model group， the TSPJ groups showed significantly reduced cell diameter and increased number of cells per unit area， especially in the high-dose TSPJ group. ④ Compared with the normal group， the model group showed decreased mRNA expression of FABP4， UCP1， CytC， PRDM16， ELOVL3， PGC-1α， and PPARγ in adipose tissues of mice （P<0.05， P<0.01）. Compared with the model group， after intervention with TSPJ， the mRNA expression was significantly up-regulated （P<0.05， P<0.01）. ⑤ Compared with the normal group， the model group showed decreased protein expression of UCP1， PRDM16， PPARγ， and PGC-1α in adipose tissues of mice （P<0.05， P<0.01）. Compared with the model group， after intervention with TSPJ， the protein expression increased significantly （P<0.05， P<0.01）. ConclusionTSPJ could induce the browning of iWAT/BAT activation and enhance adaptive thermogenesis in obese mice induced by HFD. The underlying mechanism may be attributed to the activation of the PPARγ/PGC-1α 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.

Pathologically Elevated-Cyclic Stretch Suppressed Vascular Smooth Muscle Cell Mitochondrial Biogenesis by Down-regulating PGC1α Expression / 医用生物力学

Objective To investigate the effect of pathologically elevated-cyclic stretch induced by hypertension on mitochondrial biogenesis of vascular smooth muscle cells (VSMCs), and the role of PGC1α in this process. Methods The Flexcell-5000T stretch loading system in vitro was applied to VSMCs with a frequency of 1. 25 Hz and an amplitude of 5% or 15% to simulate the mechanical environment under normal physiological or hypertensive pathological conditions respectively. Western blotting and qPCR were used to detect the expression of PGC1α, citrate synthase and mitochondrial DNA (mtDNA) copy number in VSMCs under normal physiological or hypertensive pathological conditions. VSMCs were treated with PGC1α specific activator ZLN005 to promote PGC1α expression or specific interfering fragment siRNA to inhibit PGC1α expression in order to detect the effect on citrate synthase and mtDNA copy number. Results Compared with 5% physiological cyclic stretch, 15% pathologically elevated-cyclic stretch significantly suppressed the expression of PGC1α, citrate synthase and mtDNA copy number in VSMCs. Compared with control group, the protein expression of PGC1α was significantly decreased and increased respectively. When VSMCs transfected with PGC1α siRNA or incubated PGC1α activator ZLN005, the expression of citrate synthase and mtDNA copy number were also significantly down regulated and up-regulated in VSMCs accordingly. Under physiological cyclic stretch conditions, the protein level of PGC1α was significantly down-regulated by PGC1α siRNA, which also significantly down-regulated citrate synthase expression and mtDNA copy number. The protein expression of PGC1α was significantly up-regulated by ZLN005, which also enhanced the expression of citrate synthase and mtDNA copy number. Conclusions The pathological cyclic stretch induced by hypertension significantly down-regulated the expression of citrate synthase and mtDNA copy number via suppressing the expression of PGC1α, resulting in mitochondrial dysfunction of VSMCs. PGC1α may be a potential therapeutic target molecule to alleviate the progression of hypertension.

Mechanism of Salvianolate in Inducing Autophagy in Podocytes of Rats with Membranous Nephropathy via AMPK/SIRT1/PGC-1α Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo observe the effect of salvianolate on the protein expressions of adenosine monophosphate （AMP）-activated protein kinase （AMPK）， silent information regulator 1 （SIRT1） and peroxisome proliferator-activated receptor-γ coactivator-1α （PGC-1α）， autophagy and apoptosis in kidney tissue of rats with membranous nephropathy （MN）， and to explore its possible molecular mechanism against MN. MethodEighty male SD rats were randomly divided into normal group， model group， benazepril hydrochloride group （10 mg·kg-1）， and salvianolate low-， medium-， and high-dose groups （16.7， 33.3 and 66.7 mg·kg-1）. The rats were modeled by injection of cationized bovine serum albumin （C-BSA） into the tail vein. After successful modeling， rats in the administration groups were given corresponding doses of drugs for 4 consecutive weeks， and then 24-hour urine， serum and kidney tissue were collected for the detection of 24-hour urinary protein （UTP）， blood urea nitrogen （BUN）， serum creatinine （SCr）， interleukin-6 （IL-6）， tumor necrosis factor-α （TNF-α）， C reactive protein （CRP）， glutathione peroxidase （GSH-Px）， superoxide dismutase （SOD）， and malondialdehyde （MDA）. The pathological changes of kidneys were observed by light microscope， electron microscope and immunofluorescence. Western blot was used to detect the protein expressions of phospho-AMPK （p-AMPK）， AMPK， phospho-SIRT1 （p-SIRT1）， SIRT1 and PGC-1α in rat kidney tissue. The protein expressions of autophagy-specific gene （Beclin-1）， microtubule-associated protein 1 light chain 3 （LC3） Ⅱ， ubiquitin-binding protein （p62）， B cell lymphoma （Bcl-2）， Bcl-2-associated X （Bax）， and cysteine aspartic protease-7 （Caspase-7） in rat kidney tissue were determined by immunohistochemistry （IHC）. ResultCompared with the conditions in the normal group， the levels of UTP， IL-6， TNF-α， CRP and MDA in the model group were increased （P<0.05） while the levels of SOD and GSH-Px were decreased （P<0.05）， and there was no difference in BUN and SCr. Compared with the model group， the administration groups had lowered UTP， IL-6， TNF-α， CRP and MDA （P<0.05） while elevated SOD and GSH-Px （P<0.05）. It could be seen from hematoxylin and eosin （HE） staining， Masson staining， immunofluorescence and electron microscopy that the pathological damage of rat kidney tissue in the model group was significant， but after treatment with benazepril hydrochloride and salvianolate， the pathological damage of kidney cells was gradually improved. The expressions of p-AMPK/AMPK， p-SIRT1/SIRT1， PGC-1α， Bcl-2， Beclin-1 and LC3Ⅱ in rat kidney in the model group were lower than those in the normal group （P<0.05） while the expressions of Bax， Caspase-7 and p62 were higher （P<0.05）. Compared with the model group， benazepril hydrochloride group and salvianolate groups had an up-regulation in the expressions of p-AMPK/AMPK， p-SIRT1/SIRT1， PGC-1α， Bcl-2， Beclin-1 and LC3Ⅱ in the kidney （P<0.05） while a down-regulation in the expressions of Bax， Caspase-7 and p62 （P<0.05）. ConclusionThe protective effect of salvianolate on the kidneys of MN rats may be related to the activation of AMPK/SIRT1/PGC-1α signaling pathway， the up-regulation of autophagy and the reduction of apoptosis.

Study on the effect and mechanism of taxifolin on ameliorating cisplatin-induced acute kidney injury in mice through peroxisome proliferator-activated receptor γ coactivator-1α-mediated antioxidant pathway / 中华肾脏病杂志

Objective:To explore the effect and mechanism of taxifolin (TAX) on ameliorating cisplatin-induced renal oxidative damage.Methods:(1) Forty male C57BL/6 mice were divided into 4 groups: control group ( n=10), TAX group ( n=10), cisplatin group ( n=10) and cisplatin+TAX group ( n=10). The weight of mice in each group was measured. The level of serum creatinine (Scr) and blood urea nitrogen (BUN) was analyzed. Kidney histopathological change in mice was analyzed by HE staining. The pro-inflammatory cytokines levels of interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) were measured by enzyme linked immunosorbent assay. The levels of reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT) and glutathione (GSH) were measured by multifunctional microplate reader. The expression of inflammatory factors, antioxidant genes, and peroxisome proliferator-activated receptor γ coactivator-1α ( PGC- 1) mRNA were measured by real-time PCR. Evaluation of mitochondrial function by measuring ATP level and mtDNA content. Determination of AMP-activated protein kinase (AMPK) and phosphorylated AMPK protein expression by Western blotting. (2) Evaluate the effect of taxifolin on chemotherapy of cisplatin by establishing Lewis lung cancer transplantation tumor C57BL/6 mice model. Results:Compared with the control group, the weight of the mice in the TAX group was not significantly reduced ( P>0.05), and there was no obvious kidney damage ( P>0.05), indicating that oral TAX had good safety. Compared with the cisplatin group, TAX could significantly delay cisplatin-induced the weight loss of mice, reduce the levels of Scr and BUN, and alleviate the pathological changes of kidney tissue (all P<0.05). TAX could reduce the levels of serum inflammatory factors IL-6 and TNF-α and the expression of renal inflammatory factors IL- 6, TNF- α and IL- 1β mRNA induced by cisplatin in mice (all P<0.05). TAX could significantly reduce the levels of ROS and MDA, and increase the activities of SOD, CAT and GSH in cisplatin-induced acute kidney injury mice (all P<0.01). Meanwhile, TAX could up-regulate the mRNA expression of UCP2, SOD2, CAT antioxidant genes and PGC- 1α in the kidneys of mice with acute kidney injury induced by cisplatin, and increase the levels of ATP and mtDNA in cisplatin-induced acute kidney injury mice (all P<0.01). Western blotting results showed that TAX significantly promoted the expression of phosphorylated AMPK protein in cisplatin-induced acute kidney injury mice ( P<0.01). In addition, through the establishment of Lewis lung cancer transplantation tumor C57BL/6 mice model, it was found that TAX had no significant effect on the anti-tumor efficacy of cisplatin. Conclusions:TAX can ameliorate cisplatin-induced renal oxidative damage, and its mechanism may be related to the activation of AMPK/PGC-1α pathway.

Protective Effect of Wenxin Prescription on Mitochondrial Energy Metabolism in Rats with Myocardial Ischemia-reperfusion Injury / 中国实验方剂学杂志

ObjectiveTo reveal the effect of Wenxin prescription on mitochondrial energy metabolism and silent information regulator 1 （SIRT1）/peroxisome proliferator-activated receptor-γ coactivator-1α （PGC-1α）/recombinant estrogen-related receptor α （ERRα） signaling pathway in rats with myocardial ischemia-reperfusion injury. MethodTotally 90 male Wistar rats of SPF grade were randomly assigned into a sham operation group， a model group， and low-， medium-， and high-dose Wenxin prescription groups， with 18 rats in each group. The rats in low-， medium-， and high-dose Wenxin prescription groups were administrated with 0.99， 1.98， and 3.96 g·kg-1 granules by gavage， respectively， and those in the sham operation group and model group with the same amount of normal saline. Twenty-one days after pre-administration， the rat model of myocardial ischemia-reperfusion injury was established by ligation of the left anterior descending coronary artery for 30 min and reperfusion for 2 h， and the rats in the sham operation group were only threaded without ligation. Myocardial infarction area was observed through 2，3，5-triphenyl-2h-tetrazolium chloride （TTC） staining， and the myocardial histopathology through hematoxylin-eosin （HE） staining. The levels of creatine kinase-MB （CK-MB） and lactate dehydrogenase （LDH） in serum， cytochrome C oxidase （CCO） and succinate dehydrogenase （SDH） in mitochondrion， and ATP in myocardial tissue were detected according to kit instructions. The mRNA and protein levels of SIRT1， PGC-1α， ERRα， and mitochondrial transcription factor A （TFAM） in myocardial tissue were determined by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot， respectively. ResultCompared with the sham operation group， the model group showed broken and disordered myocardial fibers， cytoplasmic edema， and pyknosis and deviation of nuclei. Moreover， the modeling increased the levels of CK-MB and LDH （P<0.05， P<0.01）， lowered the levels of ATP， CCO， and SDH （P<0.05， P<0.01）， and down-regulated the mRNA and protein levels of SIRT1， PGC-1α， ERRα， and TFAM in myocardial tissue （P<0.05， P<0.01）. Compared with the model group， Wenxin prescription reduced the myocardial infarction area （especially in the high-dose group， P<0.01）， restored the pathological changes， lowered the levels of CK-MB and LDH （P<0.05， P<0.01）， increased the levels of ATP， CCO， and SDH （especially in the high-dose group， P<0.01）， and up-regulated the mRNA and protein levels of SIRT1， PGC-1α， ERRα， and TFAM in myocardial tissue （P<0.05， P<0.01）. ConclusionWenxin prescription can protect rats from myocardial ischemia-reperfusion injury by regulating myocardial mitochondrial energy metabolism via the SIRT1/PGC-1α/ERRα signaling pathway.

Establishment of Drug-Resistant Cell Lines of Acute Myeloid Leukemia and Correlation of Sirt1 and PGC-1α Expression Levels with Drug Resistance / 中国实验血液学杂志

OBJECTIVE@#To construct cytarabine-resistant acute myeloid leukemia (AML) cell lines, and explore the correlation between Sirt1, PGC-1α expression levels and drug resistance.@*METHODS@#Human acute promyelocytic leukemia Kasumi-1 cells were induced by the method of gradually increasing the concentration of Ara-C drug. The IC50 value of Kasumi-1 cells before and after drug addition was detected by CCK-8 method, so as to construct Ara-C resistant cell lines. The expression levels of Sirt1 and PGC-1α mRNA in Kasumi-1 drug-resistant cell lines and their parental cell lines were detected by real-time fluorescence quantitative PCR, and the expression levels of Sirt1 and PGC-1α protein in kasumi-1 drug-resistant cell lines and their parental cell lines were detected by Western blot.@*RESULTS@#The constructed Kasumi-1 cell line had common morphological characteristics of drug-resistant cell lines under microscope, and the drug resistance index was greater than 5, indicating that Kasumi-1 drug-resistant cells had good drug resistance after the construction. The RT-qPCR and Western blot assays showed that the expression levels of Sirt1 and PGC-1α mRNA and protein in the drug-resistant cell lines were higher than those of the parental cell lines (P<0.001).@*CONCLUSION@#AML cell lines resistant to Ara-C can be successfully induced by the method of gradually increasing the concentration, and the co-high expression of Sirt1 and PGC-1α may mediate the drug resistance of AML cells.

Mechanism of Zexie Decoction in improvement of nonalcoholic fatty liver disease based on LKB1/AMPK/PGC-1α pathway / 中国中药杂志

The present study investigated the pharmaceutical effect and underlying mechanism of Zexie Decoction(ZXD) on nonalcoholic fatty liver disease(NAFLD) in vitro and in vivo via the LKB1/AMPK/PGC-1α pathway based on palmitic acid(PA)-induced lipid accumulation model and high-fat diet(HFD)-induced NAFLD model in mice. As revealed by the MTT assay, ZXD had no effect on HepG2 activity, but dose-dependently down-regulated alanine aminotransferase(ALT) and aspartate aminotransferase(AST) in the liver cell medium induced by PA, and decreased the plasma levels of ALT and AST, and total cholesterol(TC) and triglyceride(TG) levels in the liver. Nile red staining showed PA-induced intracellular lipid accumulation, significantly increased lipid accumulation of hepatocytes induced by PA, suggesting that the lipid accumulation model in vitro was properly induced. ZXD could effectively improve the lipid accumulation of hepatocytes induced by PA. Oil red O staining also demonstrated that ZXD improved the lipid accumulation in the liver of HFD mice. JC-1 staining for mitochondrial membrane potential indicated that ZXD effectively reversed the decrease in mitochondrial membrane potential caused by hepatocyte injury induced by PA, activated PGC-1α, and up-regulated the expression of its target genes, such as ACADS, CPT-1α, CPT-1β, UCP-1, ACSL-1, and NRF-1. In addition, as revealed by the Western blot and immunohistochemistry, ZXD up-regulated the protein expression levels of LKB1, p-AMPK, p-ACC, and PGC-1α in vivo and in vitro. In conclusion, ZXD can improve NAFLD and its mechanism may be related to the regulation of the LKB1/AMPK/PGC-1α pathway.

Study on Protective Mechanism of Dracocephalum moldavica Total Flavonoids against Myocardial Ischemia- reperfusion Injury in Rats Based on AMPK/SIRT 1/PGC-1α Signaling Pathway / 中国药房

OBJECTIVE：To s tudy the effects of Dracocephalum moldavica total flavonoids （TFDM）on AMPK/SIRT 1/PGC-1α signaling pathway ，and to explore the mechanism of its protective effect on myocardial ischemia reperfusion injury （MIRI）rats. METHODS：Totally 50 healthy male SD rats were randomly divided into sham operation group ，model group ，TFDM group [ 60 mg/（kg·d），by extract] ，Compound C+TFDM group [ig administration of 60 mg/（kg·d）TFDM+intravenous injection of 250 μg/kg Compound C（AMPK inhibitor ）via tail vein 15 min before reperfusion] ，EX-527+TFDM group [ig administration of 60 mg/ （kg·d）TFDM+ip injection of 5 mg/kg EX- 527（SIRT1 inhibitor）20 min before reperfusion] ，with 10 rats in each group. They were given relevant medicine intragastrically ，once a day ，for consecutive 7 days. After last ig administration ，sham operation group underwent sham operation ，other 4 groups were established MIRI model by ligating left anterior descending coronary artery ， ischemia for 30 min and reperfusion for 2 h. After reperfusion ，the myocardial histopathological changes were observed by HE staining；RP-HPLC method was used to determine the contents of ATP ，ADP，AMP and NAD + in cardiac tissue. mRNA expressions of AMPK ，SIRT1 and PGC- 1α were detected by quantitative real-time PCR assay. Western blotting assay was the expressions of SIRT 1 and PGC- 1α protein in myocardium. RESULTS： Compared with sham operation group ， model group showed myocardial fib ers arranged disorder and horizontal stripes disappearance ，cell swelling burst and necrosis ，and nuclei deformation displacement ；the contents of ATP and NAD+，mRNA expression of AMPK ，SIRT1 and PGC- 1α，protein expression of SIRT 1 and PGC- 1α in cardiac tissue were decreased significantly （P＜0.05 or P＜0.01）；the contents of ADP and AMP ，the phosphorylation level of AMPK protein were increased significantly （P＜0.01）. Compared with model group ，myocardial pathological morphology were improved significantly in TFDM group ；the contents of ATP and NAD + in cardiac tissue ，mRNA expression of AMPK ，SIRT1 and PGC- 1α，the phosphorylation level of AMPK protein ，the protein expression of SIRT 1 and PGC- 1α were increased significantly（P＜0.05 or P＜ 0.01），while the contents of ADP and AMP were decreased significantly （P＜0.01）. Compared with TFDM group ，improvement effects of Compound C + TFDM group and EX- 527 + TFDM group on above indexes were reversed （P＜0.05 or P＜0.01）. CONCLUSIONS：TFDM may play a protective role on myocardium by activating AMPK/SIRT 1/PGC-1α signaling pathway and regulating energy metabolism.

Da Chaihutang Protects Liver Mitochondrial Function of Nutritionally Obese Rats via CREB/PGC-1α Pathway / 中国实验方剂学杂志

Objective:To observe the effects of Da Chaihutang on Cyclic adenosine monophosphate （cAMP）-response element binding protein （CREB）/peroxisome proliferator-activated receptor-gamma coactivator-1 alpha （PGC-1<italic>α</italic>） pathway in nutritionally obese rats and the protective mechanism on liver mitochondria. Method:A total of 120 8-week-old male SD rats were randomly divided into a control group （<italic>n</italic>=20） and an experimental group （<italic>n</italic>=100）. The rats in the control group were fed on a normal diet， while those in the experimental group were administered with a high-fat feed. Successfully modeled rats were randomly divided into a model group， a positive drug （metformin） group， and low-， medium- and high-dose Da Chaihutang groups （4.25， 8.5， and 17 g∙kg^-1， respectively）， with 20 rats in each group. After treatment with Da Chaihutang， the body weight， Lee's index， liver mitochondrial membrane potential and mitochondrial ultrastructure， PGC-1<italic>α </italic>expression and CREB phosphorylation of each group were measured and compared. Result:Compared with the control group， the model group showed increased body weight and Lee's index （<italic>P</italic><0.01）， whereas decreased mitochondrial membrane potential， PGC-1<italic>α</italic> expression， and CREB phosphorylation level （<italic>P</italic><0.01）. As compared with the model group， Da Chaihutang significantly reduced the body weight and Lee's index of obese rats （<italic>P</italic><0.05， <italic>P</italic><0.01）， enhanced liver mitochondrial membrane potential （<italic>P</italic><0.05， <italic>P</italic><0.01） to protect the integrity of mitochondrial structure， up-regulated PGC-1<italic>α</italic> expression and promoted CREB phosphorylation （<italic>P</italic><0.05， <italic>P</italic><0.01）. Conclusion:Da Chaihutang protects the structure and function of mitochondria and inhibits weight gain in obese rats by activating the CREB/PGC-1<italic>α</italic> pathway.

Effect of AMPK/PGC1 alpha on improving skeletal muscle atrophy in type 2 diabetic rats by aerobic exercise / 中国组织工程研究

BACKGROUND: The regulation of mitochondrial energy metabolism by adenylate activated protein kinase (AMPK) is an important cause of fat accumulation in obese and type 2 diabetic patients. Chronic inflammation will further induce skeletal muscle atrophy. Aerobic exercise can increase the activity of AMPK and regulate energy metabolism, but the mechanism of aerobic exercise in improving skeletal muscle atrophy in type 2 diabetes by increasing AMPK is unclear. OBJECTIVE: To explore the effect of aerobic exercise on skeletal muscle atrophy in type 2 diabetic rats and the role of AMPK. METHODS: The model of type 2 diabetic rats was established by high fat feeding and streptozotocin injection, and the rats were divided into four groups: control group (n=6), exercise group (n=9), diabetic control group (n=8) and diabetic exercise group (n=12). The control group and the diabetic control group were kept for 4 weeks, and the exercise group and the diabetic exercise group were given aerobic exercise intervention for 4 weeks. After 4 weeks of aerobic exercise (running speed 16 m/min, 60 min/d, 5 days/week), the muscle atrophy of soleus was observed by immunohistochemical staining. The expression levels of AMPK, PGC-1 α, MAFbx and MuRF1 were detected by western blot assay. The study protocol was approved by the Ethical Committee of School of Sport Science, Beijing Sport University in China on June 25, 2016, with approval No. 2016014. RESULTS AND CONCLUSION: Blood glucose of type 2 diabetes rats was significantly increased, and body weight and insulin levels of type 2 diabetes rats were significantly decreased (P < 0.01). The mean cross sectional area of soleus fiber in the diabetic group was significantly lower than that in the control group (P < 0.01), and the cross sectional area of soleus muscle fiber in the diabetic exercise group was significantly higher than that in the diabetic group (P < 0.01). The expression levels of AMPK and PGC-1 α in the soleus muscle of diabetic rats were significantly lower than those in the control group, and the expression levels of MAFbx and MuRF1 were significantly higher than those in the control group (P < 0.01). The expression levels of AMPK, MAFbx and MuRF1 in the diabetic exercise group were significantly higher than those in the diabetic group (P < 0.01). These results suggest that aerobic exercise can improve mitochondrial function, inhibit the expression of MAFbx and MuRF1, improve skeletal muscle atrophy and restore the metabolic balance of type 2 diabetes mellitus to some extent by activating AMPK/PGC-1α signaling pathway.

Effect of Xiao Jianzhongtang on AMPK/PGC1-α Signal Pathway in Skeletal Muscle of Exercise Fatigue Mice / 中国实验方剂学杂志

Objective:To observe the effect of Xiao Jianzhongtang on Adenylate-activated protein kinase/peroxidase proliferation-activated receptor coactivator 1-α （AMPK/PGC1-α） signaling pathway in skeletal muscle of exercise fatigue mice.Method:Forty Kunming mice were randomly divided into normal group， model group， Buzhong Yiqitang group and Xiao Jianzhongtang group， with 10 mice in each group. The model group， Buzhong Yiqitang group and Xiao Jianzhongtang group were trained on the treadmill to establish a fatigue model， and the normal group did not apply any intervention. At the same time as the treadmill training， the model group was given the same amount of normal saline. Xiao Jianzhongtang was administered with 5 g·kg-1 of medicine， and Buzhong Yiqitang was administered with 2.8 g·kg-1 of medicine for 6 days. After the experiment， the weight of each group of mice and the time of running out of exhaustion were measured，the colorimetric method was used to detect the serum urea （UREA）， lactate dehydrogenase （LDH）， muscle glycogen （MG）， and skeletal muscle of each group of mice Na+-K+-ATPase， Ca2+-Mg2+-ATPase content， pathological changes of skeletal muscle of each group were observed by hematoxylin-eosin （HE） staining， Western blot was used to detect the protein expression of AMPK and PGC1-α in skeletal muscle of each group .Result:Compared with normal group， the body weight of model group significantly decreased （P<0.01）， and the contents of Na+-K+-ATPase， Ca2+-Mg2+-ATPase， LDH， and MG significantly decreased （P<0.05，P<0.01）. The content of UREA increased significantly （P<0.01）， and the expression of AMPK and PGC1-α protein increased significantly （P<0.01）. Compared with model group， the mice in the Xiao Jianzhongtang group had significantly increased body weight （P<0.05）， significantly increased the time spent on treadmill exhaustion（P<0.01）， Na+-K+-ATPase， Ca2+-Mg2+-ATPase， LDH， and MG. The content increased significantly（P<0.05， P<0.01）， the content of UREA decreased significantly （P<0.01）， and the expression of AMPK and PGC1-α protein increased significantly （P<0.01）.Conclusion:Xiao Jianzhongtang has an anti-exercise fatigue effect， which may be related to enhancing skeletal muscle AMPK/PGC1-α pathway，enhancing mitochondrial oxidative phosphorylation，reducing accumulation of metabolites，slowing down glycogen consumption and decomposition，and enhancing skeletal muscle energy synthesis.

Effect of electroacupuncture combined with treadmill exercise on body weight and expression of PGC-1α， Irisin and AMPK in skeletal muscle of diet-induced obesity rats / 针刺研究

OBJECTIVE: To investigate the effect of electroacupuncture (EA) plus treadmill exercise on the expression of peroxisome proliferator activated receptor γ coactivator 1α(PGC-1α), Irisin, AMP-activated protein kinase (AMPK) in skeletal muscle of diet-induced obesity (DIO) rats, so as to explore its mechanism underlying body reduction promotion. METHODS: Forty-two male SD rats were divided into normal diet (control, n＝10), high fat diet (model), EA, treadmill exercise and EA plus treadmill exercise (combination) groups (n＝8 in each of the latter 4 groups). The obesity model was established by feeding the rats with high fat diet. EA (2 Hz/15 Hz, 1 mA) was applied to bilateral "Zusanli" (ST36) and "Tianshu" (ST25) for 30 min, 5 times per week for a total of 8 weeks. Rats of the treadmill exercise group were forced to perform exercise on a treadmill (16 m/min) for 30 min, 5 times per week for a total of 8 weeks. Rats in the combination group received the above-mentioned two methods. During the treatment, rats in the control group were fed with normal fodder, rats in other groups were fed with high fat fodder, and their body weight was measured once a week. The expression levels of PGC-1α， fibronectin type Ⅲ domain containing 5 (FNDC5), AMPK mRNA and protein of skeletal muscle were measured by quantitative real-time PCR and Western blot，respectively. RESULTS: After modeling, the body weight was significantly increased (P<0.05), and the expression levels of PGC-1α and FNDC5 mRNA and protein, AMPK mRNA and phosphorylated AMPK (p-AMPK) protein in the skeletal muscle were considerably decreased in the model group relevant to the control group (P<0.05). Following the treatment, the body weight was significantly down-regulated, while the expression levels of PGC-1α and FNDC5 mRNAs and proteins, AMPK mRNA and p-AMPK protein were obviously up-regulated in the EA, treadmill exercise and combination groups relevant to the model group (P<0.05). The therapeutic effect of EA plus treadmill exercise was significantly superior to those of both simple EA and simple treadmill exercise in down-regulating the body weight, as well as in up-regulating the expression of PGC-1α and FNDC5 mRNAs and proteins, AMPK mRNA, and p-AMPK protein (P<0.05). CONCLUSION: Both EA and treadmill exercise can significantly increase the expression of PGC-1α， FNDC5 and p-AMPK in skeletal muscle of DIO rats, suggesting their efficacy in restoring fatty acid oxidation in skeletal muscle cells and improving mitochondrial function, which may contribute to their function in body reduction. The therapeutic effect of EA plus treadmill exercise is better than that of simple EA and simple treadmill exercise.

Effects of sevoflurane postconditioning on oxidative stress and expression of SIRTl/PGC-1α of hippocampus in a rat model of hemorrhagic shock and resuscitation / 中国药理学通报

Aim To investigate the effects of sevoflurane postconditioning on oxidative stress and the expression of silent information regulation (SIRT1) and peroxisome proliferator-activated receptor γ coactivator la (PGC-1α) in hippocampus of rats subjected to hemorrhagic shock and resuscitation. Methods Male SD rats were randomly divided into sham surgery group (Sham group), shock and resuscitation group (Shock group) and 2.4% sevoflurane postconditioning group (Sevo group). The rats in Sevo group were inhaled 2.4% sevoflurane when received resuscitation after hemorrhagic shock, while rats in Sham and Shock group were treated with 95% O2 and 5% CO2 in the corresponding period. MAP and arterial blood gases were measured at TO (start bleeding), Tl (30 min after bleeding),T2 (start resuscitation), and T3 (30 min after resuscitation). After 24h of surgery,rats with successful model were chosen for the detection of various indexes. The content of malonaldehyde (MDA) in hippocampus and the activity of superoxide dismutase (SOD) in mitochondria isolated from hippocampal tissue were detected. Western blot was used to analyze the protein relative expression levels of SIRT1 and PGC-la in hippocampus. Results Compared with Sham group, the content of MDA increased, the activity of SOD decreased, and the expression of SIRT1 and PGC-1α a increased in Shock group (P < 0. 05). Compared with Shock group, the content of MDA decreased, the activity of SOD increased, and the expression of SIRT1 and PGC-1α increased in Sevo group (P < 0. 05). Conclusions Sevoflurane postconditioning can alleviate oxidative stress in hippocampus of a model rat of hemorrhagic shock and resuscitation, which may be correlated with the up-regulation of the protein relative expression levels of SIRT1 and PGC-1α.

Effects of phlegm-resolving and stasis-removing herbal drugs on PGC-1α mRNA expressions and insulin resistance in NAFLD rats / 国际中医中药杂志

Objective To explore the mechanism of phlegm-resolving and stasis- removing herbals on NAFLD by observing expressions of PGC1α mRNA and insulin resistance. Methods A total of 60 male SD rats were randomly divided into the normal group, model group, positive medication control group, high-dose, middle-dose and low-dose group. The rats were fed with high-fat forage for 8 weeks. The positive medication control group were gavaged with Dongbao-Gantai liquid (0.9 g/kg/day), the high-dose, middle-dose and low-dose group were gavaged with Xiaotan-Huayu liquid (43.34、32.50、21.67 g/kg/day), and normal group, model group were gavaged with equal volume of distilled water. The drugs were given by 1 ml/100 g and last for 8 weeks. The levels of TC, TG, FFA, ALT, AST, FBG, FINS, and HOMA-IR in serum, and levels of TC, TG, and PGC-1α mRNA and pathological morphological changes in hepatic tissue were observed after 8 weeks. Results The levels of TG (0.55 ± 0.10 mmol/L, 0.58 ± 0.09 mmol/L, 0.67 ± 0.11 mmol/L vs. 1.18 ± 0.15 mmol/L), TC (1.48 ± 0.24 mmol/L, 1.69 ± 0.27 mmol/L, 1.74 ± 0.27 mmol/L vs. 3.29 ± 0.26 mmol/L), FFA (251.08 ± 48.18 μmol/L, 277.53 ± 56.73 μmol/L, 291.82 ± 48.67 μmol/L vs. 432.19 ± 67.83 μmol/L), ALT (29.32 ± 4.17 U/L, 31.26 ± 4.74 U/L, 33.56 ± 5.18 U/L vs. 47.21 ± 8.67 U/L), AST (11.05 ± 2.18 U/L, 12.15 ± 2.67 U/L, 12.96 ± 2.93 U/L vs. 19.43 ± 3.68 U/L), FBG (5.68 ± 1.22 mmol/L, 6.86 ± 1.36 mmol/L, 7.94 ± 1.82 mmol/L vs. 11.88 ± 2.54 mmol/L), FINS (8.48 ± 1.22 mmol/L, 9.55 ± 1.95 mmol/L, 9.96 ± 1.74 mmol/L vs. 12.96 ± 2.67 mmol/L), HOMA-IR (1.91 ± 0.26, 2.91 ± 0.65, 3.52 ± 0.58 vs. 6.89 ± 1.21) in serum of high-dose, middle-dose and low-dose groups were decreased than model group. Levels of FFA (242.19 ± 35.13 μmol/L, 259.78 ± 29.33 μmol/L, 277.62 ± 34.29 μmol/L vs. 436.48 ± 52.15 μmol/L), TG (23.65 ± 3.28 mmol/L, 24.41 ± 3.15 mmol/L, 25.37 ± 3.59 mmol/L vs. 15.98 ± 2.37 mmol/L), TC (7.15 ± 0.82 mmol/L, 8.60 ± 0.95 mmol/L, 8.86 ± 1.04 mmol/L vs. 36.98 ± 4.28 mmol/L) were in hepatic tissue of high-dose, middle-dose and low-dose groups were significantly lower than the model group. The levels of PGC-1α mRNA (1.24 ± 0.06, 1.02 ± 0.07, 0.99 ± 0.08 vs. 0.43 ± 0.06) in hepatic tissue of high-dose, middle-dose and low-dose groups were significantly higher than model group. Conclusions The phlegm-resolving and stasis-removing herbals may improve lipid metabolism by regulating the expression of PGC-1α mRNA, inhibiting gluconeogenesis and liver sugar output, correcting disturbance of lipid metabolism and improving insulin resistance.

Electroacupuncture of "Zusanli" (ST 36) Raises Muscular Force by Adjusting AMPK/PGC-1 α Signaling in Rats with Chronic Fatigue Syndrome / 针刺研究

OBJECTIVE: To observe the effect of electroacupuncture (EA) of "Zusanli" (ST 36) on mitochondrial oxidative stress of skeletal muscle in rats with chronic fatigue syndrome (CFS) based on adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK)/ peroxlsome proliferator-activated receptor-γ coactivator-1 α (PGC-1 α) signaling, in order to reveal its mechanism underlying improvement of CFS. METHODS: Forty SD rats were randomly divided into normal control, CFS model, EA-Zusanli (ST 36) and EA-non-acupoint groups (n=10 rats in each group). The CFS model was established by forced exhausted load-bearing swimming (twice daily), chronic constraint (1 h) and sleep deprivation (20 h/day) for 14 days. Following modeling, EA (2 Hz/100 Hz, 2 V) was applied to bilateral Zusanli (ST 36) or non-acupoint (about 10-15 mm superior to the bilateral Iliac creast and about 20 mm lateral to the posterior median line) for 20 min, once a day for 10 days. The expression levels of ATP synthase, AMPK, phosphorylated (p)-AMPK, silent mating type information regulation 2 homolog-1 (SIRT 1) and PGC-1 α proteins, and ATP synthase, SIRT 1 and PGC-1 α mRNAs of the quadriceps femoris muscle were detected by Western blot and fluorescence quantitative PCR, respectively. The rats' grabbing force was detected by using a grabbing-force detector. RESULTS: Compared with the normal group, the grabbing force, and the expression levels of ATP synthase and PGC-1 α proteins and mRNAs were significantly decreased (P<0.05, P<0.01), while the expression of SIRT 1 protein was significantly up-regulated (P<0.05) in the CFS model group. Following EA intervention, the grabbing force and the expression levels of ATP synthase mRNA, SIRT 1 and PGC-1 α proteins and mRNAs, and p-AMPK/AMPK were significantly up-regulated in the EA-Zusanli (ST 36) group (P<0.05, P<0.01). CONCLUSION: EA of ST 36 can raise the grabbing force of CFS rats, which may be related to its effects in up-regulating the expression of ATP synthase mRNA, SIRT 1 and PGC-1 α proteins and mRNAs, and p-AMPK/AMPK to reduce mitochondrial oxidative stress reaction and in increasing ATP synthesis.

Electroacupuncture Relieves Obesity by Up-regulating PGC-1 α/UCP-1 Signaling in White Adipose Tissue in Diet-induced Obesity Rats / 针刺研究

OBJECTIVE: To observe the effect of electroacupuncture (EA) on the activities of peroxisome proliferator-activated receptor gamma coactivator-1 alpha/uncoupling protein-1 (PGC-1 α/UCP-1) signaling pathway in white adipose tissue(WAT)of diet-induced obesity (DIO) rats, so as to reveal its underlying mechanism in body weight loss. METHODS: Thirty-four male Wistar rats were randomly divided into normal diet (control, n＝10), high fat diet (model), sham EA-acupoint and EA groups (n＝8 in each of the latter 3 groups). The obesity model was established by feeding the rats with high fat diet containing lard oil, sugar, albumen powder, cholesterol, salt and sodium cholate for 12 weeks. EA (2 Hz/15 Hz, 1 mA) was applied to bilateral "Zusanli" (ST 36) and "Tianshu" (ST 25) or sham acupoints (about 5 mm beside ST 36 and ST 25) for 30 min, once daily, 5 times per week for a total of 8 weeks. During the treatment, all rats were fed with normal diet, and their body weight and length were measured once a week for calculating the Lee's index. The contents of serum total cholesterol (TC) and triglyceride (TG) were measured by using biochemical methods. The immunoactivity of PGC-1 α and UCP-1 in the abdominal WAT was detected by immunohistochemistry. RESULTS: After modeling, the Lee's index, serum TC and TG contents were significantly increased, and the levels of serum HDL-C, and PGC-1 α and UCP-1 immunoactivity in WAT considerably decreased in the model group relevant to the control group (P<0.05). Following the treatment, the Lee's index, TC and TG contents were significantly down-regulated while HDL-C and PGC-1 α and UCP-1 immunoactivity were obviously up-regulated in the EA-acupoint group relevant to the model group (P<0.05). CONCLUSION: EA can effectively reduce the body weight and adipose content in obesity rats, which may be closely related to its effect in up-regulating PGC-1 α/UCP-1 signaling in WAT, suggesting an efficacy of EA in promoting the browning of WAT.

Protective effects of puerarin on high glucose-induced oxidative injury in HUVEC-12 cells / 中成药

AIM To explore the action mechanism of puerarin's protective effects against oxidative stress of HUVEC-12 cells induced by high glucose.METHODS HUVEC-12 cells cultured with 100 mmol/L glucose medium and 10,25,50 μmoL/L puerarin for 36 h had the cell proliferation,the levels of lactate dehydrogenase (LDH),intracellular reactive oxygen species (ROS),the activities of caspase-3,superoxide dismutase (SOD)and catalase (CAT),and the contents of malondialdehyde (MDA) and glutathione (GSH) measured.The mRNA expressions of SIRT1 and PGC-1α were detected by real-time flu orescence quantitative PCR,and the protein contents of SIRT1 and PGC-1 α were determined by enzyme-linked immunosorbent assay.RESULTS The puerarin treatment to HUVEC-12 cells resulted in markedly lowered LDH level,caspase-3 activity,intracellular levels of MDA and ROS,and notable improvement of the cell viability,the activities of SOD and CAT,GSH content,the mRNA expressions and the protein contents of SIRT1 and PGC-1α as well.CONCLUSION The protective effect of puerarin on high glucose-induced oxidative damage of HUVEC-12 cells may be attributed to the SIRT1/PGC-1α pathway activation.

Effects of salvianolic acid D on mitochondrial function and biosynthesis in SH-SY5Y cells after MPP+injury / 中国药理学通报

Aim To investigate the effects of salvianol-ic acid D ( SalD) on mitochondrial function and bio- synthesis in SH-SY5Y cells after MPP+injury and the possible mechanisms. Methods The cell model was established by MPP+injury in SH-SY5Y cells. The cytotoxicity of MPP+was detected by MTT assay. The effects of SalD on viability of SH-SY5Y cells were ex-amined by MTT and LDH assay. The apoptosis of SH-SY5Y cells was detected by AO/EB assay. The levels of ROS and mitochondrial superoxide were determined using DCFH-DA and MitoSOX probes, respectively. Mitochondrial function was examined by measuring ATP level and mitochondrial membrane potential. The levels of PGC-1α and its downstream regulatory genes NRF1 and TFAM mRNA were detected by qPCR. The protein levels of PGC-1α, NRF1 and TFAM in cells were detected by Western blot and immunofluorescence assays. Results MPP+injury resulted in a significant reduction of cell viability to 51.34%. 0.1, 1, 5 μmol ·L-1SalD and 5 mmol·L-1NAC could reduce MPP+-induced SH-SY5Y cell injury and LDH release. The cell viability increased to 67.98% , 71.79% , 76.91% and 77.55% , respectively. Moreover, SalD could reduce the increase of intracellular ROS and mi-tochondrial superoxide induced by MPP+, decrease mitochondrial membrane potential and improve mito-chondrial function. SalD also significantly increased both the transcription and expression levels of PGC-1α, NRF1 and TFAM. Conclusion SalD could in-hibit MPP+-induced SH-SY5Y cell injury and improve mitochondrial function and mitochondrial biosynthesis.

Effects of Aerobic and Resistance Exercises on Morphology and Activity of Brown Adipose Tissues in Rats / 中国运动医学杂志

Objective To compare the effect of aerobic and resistance exercise on brown adipose tissues in rats.Methods Sprague-Dawley rats were randomly assigned to a control(C)group,an aerobic exercise (T) group and a resistance exercise(L)group.The rats of group T performed treadmill exercise 5 days per week for 8 weeks,while those of group L climbed vertical ladders with progressively increased weights once in 3 days for 8 weeks.All rats were weighed before and after the intervention.Then the bilateral interscapular brown adipose tissue was isolated and weighed at 48h after the last exercise.The hematoxylin-eosin staining was used to observe the morphology of adipocytes,and the quantitative real-time PCR and Western blotting were employed to detect the mRNA and protein expression of browning genes [PR-domain-containing 16 (PRDM 16),PPAR gamma coactivator-1α (PGC-1 α) and uncoupling protein 1(UCP1)] respectively.Results After the intervention,the increment of weight of group T and group L were only 52％ and 70％ of group C(P＜0.01 and P＜0.05).The weight of BAT in both group T and L was lower than group C,but without significance(P＞0.05).The average size of lipid droplets in group T decreased significantly(P＜0.01) and slightly in group L(P＞0.05) compared with group C.The PRDM16,PGC-1α and UCP1 mRNA expression of group T was 1.78(P＜0.01),2.2(P＜ 0.01) and 1.3(P＞0.05) times of group C,while that of group L was 1.30(P＞0.05),1.25(P＞0.05) and 1.21(P＞0.05) times of group C.The PRDM16,PGC-1α and UCP1 protein level in group T was 1.46-fold(P＜0.05),1.56-fold(P＞0.05)and 1.20-fold(P＞0.05),while that of group L was 1.08-fold(P＞0.05),0.94-fold(P＞0.05) and 1.20-fold(P＞0.05) of group C.Conclusion Eight weeks of aerobic exercise can significantly make the lipid droplets of adipocyte smaller,increase the differentiation and metabolic activity of BAT,and weakly stimulate BAT thermogenesis.However,8 weeks of resistance exercise has no significant effect on BAT.