Acute Exercise Induced Mitochondrial H 2

Exercise induced skeletal muscle phenotype change involves a complex interplay between signaling pathways and downstream regulators. This study aims to investigate the effect of acute exercise on mitochondrial H2O2 production and its association with p66Shc, FOXO3a, and antioxidant enzymes. Male ICR/CD-1 mice were subjected to an acute exercise. Muscle tissues (gastrocnemius and quadriceps femoris) were taken after exercise to measure mitochondrial H2O2 content, expression of p66Shc and FOXO3a, and the activity of antioxidant enzymes. The results showed that acute exercise significantly increased mitochondrial H2O2 content and expressions of p66Shc and FOXO3a in a time-dependent manner, with a linear correlation between the increase in H2O2 content and p66Shc or FOXO3a expression. The activity of mitochondrial catalase was slightly reduced in the 90 min exercise group, but it was significantly higher in groups with 120 and 150 min exercise compared to that of 90 min exercise group. The activity of SOD was not significantly affected. The results indicate that acute exercise increases mitochondrial H2O2 production in the skeletal muscle, which is associated with the upregulation of p66Shc and FOXO3a. The association of p66Shc and FOXO3a signaling with exercise induced H2O2 generation may play a role in regulating cellular oxidative stress during acute exercise

Antioxidant Supplement Inhibits Skeletal Muscle Constitutive Autophagy rather than Fasting-Induced Autophagy in Mice

In this study, we tested the hypothesis that NAC administration leads to reduced oxidative stress and thus to decreased expression of autophagy markers in young mice. Our results reveal that NAC administration results in reduced muscle mRNA levels of several autophagy markers, including Beclin-1, Atg7, LC3, Atg9, and LAMP2. However, NAC supplement fails to block the activation of skeletal muscle autophagy in response to fasting, because fasting significantly increases the mRNA level of several autophagy markers and LC3 lipidation. We further examined the effects of NAC administration on mitochondrial antioxidant capacity in fed and 24-hour fasted mice. Our results clearly show that NAC administration depresses the expression of manganese superoxide dismutase (MnSOD) and TP53-induced glycolysis and apoptosis regulator (TIGAR), both of which play a predominant antioxidant role in mitochondria by reducing ROS level. In addition, we found no beneficial effect of NAC supplement on muscle mass but it can protect from muscle loss in response to fasting. Collectively, our findings indicate that ROS is required for skeletal muscle constitutive autophagy, rather than starvation-induced autophagy, and that antioxidant NAC inhibits constitutive autophagy by the regulation of mitochondrial ROS production and antioxidant capacity

PO-062 Acute exercise intervention combined with metformin’s influences on glucose homeostasis in T2D mice

Objective Type 2 diabetes mellitus is a common chronic diseases prevailing in the world and the amount of diabetic and pre-diabetic patients is increasing gradually. Exercise combined with hypoglycemic drug is the first recommended therapy to treat type 2 diabetes. Metformin was found from galegine in 1957 and has been used now as the first cheap and effective hypoglycemic guanidines. Our study aims to explore the effects of different ways of acute exercise intervention combined with high dose of metformin on glucose homeostasis and its relative molecular mechanisms in type 2 diabetic mice. Methods Adopt 4-week high fat diet (HFD, 45% fat content) and one-time STZ (Streptozocin, 100mg/kg) intraperitoneal injection to build type 2 diabetic mice. There are 84 mice in total, 24 mice were divided into three groups: normal control (NC) group, normal acute resistance training (NCR) group and normal acute endurance training (NCE) group, N=8 each group, they were fed normal chow. The rest 60 mice were fed HFD as T2D modeling group. 48 mice were developing type 2 diabetes and they were divided into 6 groups: diabetic control (DC) group, diabetic acute resistance training (DCR) group, diabetic acute endurance training (DCE) group, high dose of metformin control (HMC) group, high dose of metformin combined with acute resistance training (HMR) group and high dose of metformin combined with acute endurance training (HME) group, N=8 each group. Acute resistance training is climbing 1 meter ladder from down to up, 5 times a group, 3 groups in total, monitoring the glucose change with extracting mouse tail vein blood during each group, using ACCU-CHEK monitor. Acute endurance training is running at the speed of 18 m/min on the platform for 50 minutes and blood glucose change was monitored every 10 minutes by extracting mouse tail vein blood. HMC, HMR and HME group mice were intraperitoneally injected high dose of metformin (200mg/kg) one hour before the acute exercise intervention. Comparatively, NC, NCR, NCE, DC, DCR, DCE group mice were intraperitoneally injected 0.9% saline one hour before the acute exercise intervention. ELISA, RT-PCR and Western Blot were used to evaluate relative serum indicators, mRNA and protein expression of regulating blood glucose homeostasis. Results 1) 4-week high fat diet and one time 100mg/kg Streptozocin intraperitoneal injection induces mice to develop type 2 diabetes. The fasting blood glucose, IPGTT, ITT, glucose AUC and insulin AUC of T2D group mice are significantly higher than NC group. 2) Compared with DCR group, the blood glucose value and fluctuation of HMR group mice are both significantly decreased, but the blood glucose value of DCR and HMR group mice are significantly higher than NCR group. In the same way, the blood glucose value and fluctuation of HME group mice is lower than DCE group and the whole blood glucose level of both group are higher than NCE group. Acute resistance training and acute endurance training combined with high dose of metformin have not affected the weight of type 2 diabetic mice. Hence compared with HMC group, the eWAT (epididymal white adipose tissue) of HMR and HME group mice is significantly declined. 3) Compared with NC group, the indicators of serum glucose, GSP (glycosylated serum protein), serum TG and serum T-CHO of DC group are notably increased, further reflect that the success model of type 2 diabetic mice. Compared with HMC group, the indicators of serum glucose, GSP, serum TG and serum T-CHO of HMR group mice are notably decreased, in the mean time, the indicators of serum glucose and serum TG of HME group mice are significantly declined. Interestingly, the serum insulin of HME group mice is notably lower than HMR group. 4) Compared with DC group, the indicators of mRNA expression about hepatic gluconeogenesis key rate-limiting enzymes PEPCK and G6pase of HMC group are significantly declined, but mRNA expression of regulating hepatic glucose homeostasis GLUT2 of HMC group is notably raised. Compared with HMC group, G6Pase mRNA expression of HMR and HME group is significantly escalated and Fbp mRNA expression of both groups are significantly declined. Compared with HMC, the indicators of mRNA expression about regulating hepatic glucose homeostasis GLUT2 and Gck of HMR and HME group mice show opposite trend, the former is down and the latter is up. Compared with HMC group, PEPCK mRNA expression of HMR group mice is notably escalated. Compared with HMR group, PEPCK and G6Pase mRNA expression of HME group mice are notably raised. 5) In the liver, there is a signaling pathway of AMPKα-PGC-1α-CREB to regulate glucose homeostasis and hepatic gluconeogenesis. Our study find that compared with HMC group, AMPKα2, PGC-1α and CREB mRNA expression of HMR and HME group mice are notably increased and only AMPKα1 mRNA expression of HMR group mice is significantly increased. Conclusions 1) Acute resistance training and acute endurance training combined with high dose of metformin can effectively reduce glucose fluctuation during exercise in type 2 diabetic mice, therefore these two way can both improve glucose homeostasis during acute exercise intervention in type 2 diabetic mice. 2) Acute resistance training and acute endurance training combined with high dose of metformin can improve serum glucose and lipid metabolism in type 2 diabetic mice, but acute resistance training combined with high dose of metformin are better to improve serum lipid metabolism. 3) Acute exercise intervention combined with high dose of metformin can comparatively increase hepatic gluconeogenesis key rate-limiting enzymes PEPCK and G6Pase and regulating hepatic glucose transport Gck mRNA expression. In the opposite, these two ways inhibit the other hepatic gluconeogenesis key rate-limiting enzyme Fbp and regulating hepatic glucose transport GLUT2 mRNA expression. 4) Compared with acute endurance training combined with high dose of metformin, acute resistance training combined with high dose of metformin can better improve glucose homeostasis and hepatic gluconeogenesis in type 2 diabetic mice via the signaling pathway of AMPKα-PGC-1α-CREB

OR-008 ERK-BAX signaling is involved in GLP-1-mediated antidepressant effects of metformin and exercise in CUMS mice

Objective Both depression itself and antidepressant medication have been reported to be significantly related to the risk of type 2 diabetes mellitus (T2DM). Glucagon-like peptide-1 (GLP-1), a treatment target for T2DM, has a neuroprotective effect. As an enhancer and sensitiser of GLP-1, metformin has been reported to be safe for the neurodevelopment. The present study aimed to determine whether and how GLP-1 mediates antidepressant effects of metformin and exercise in mice. Methods Male C57BL/6 mice were exposed to chronic unpredictable mild stress (CUMS) for 8 weeks. From the 4th week, CUMS mice were subjected to oral metformin treatment and/or treadmill running. A videocomputerized tracking system was used to record behaviors of mice for a 5-min session. ELISA, western blotting and immunohistochemistry were used to examine gene expression in mouse serum or hippocampus. Results Our results supported the validity of metformin as a useful antidepressant; moreover, treadmill running favored metformin effects on exploratory behaviors and serum corticosterone levels. CUMS reduced GLP-1 protein levels and phosphorylation levels of extracellular signal-regulated kinase 1/2 (ERK1/2), but increased protein levels of B-cell lymphoma 2-associated X-protein (BAX) in mice hippocampus. All these changes were restored by both single and combined treatment with metformin and exercise. Conclusions Our findings have demonstrated that ERK-BAX signaling is involved in GLP-1-mediated antidepressant effects of metformin and exercise, which may provide a novel topic for future clinical research

Physical Exercise Regulates p53 Activity Targeting SCO2 and Increases Mitochondrial COX Biogenesis in Cardiac Muscle with Age

The purpose of this study was to outline the timelines of mitochondrial function, oxidative stress and cytochrome c oxidase complex (COX) biogenesis in cardiac muscle with age, and to evaluate whether and how these age-related changes were attenuated by exercise. ICR/CD-1 mice were treated with pifithrin-μ (PFTμ), sacrificed and studied at different ages; ICR/CD-1 mice at younger or older ages were randomized to endurance treadmill running and sedentary conditions. The results showed that mRNA expression of p53 and its protein levels in mitochondria increased with age in cardiac muscle, accompanied by increased mitochondrial oxidative stress, reduced expression of COX subunits and assembly proteins, and decreased expression of most markers in mitochondrial biogenesis. Most of these age-related changes including p53 activity targeting cytochrome oxidase deficient homolog 2 (SCO2), p53 translocation to mitochondria and COX biogenesis were attenuated by exercise in older mice. PFTμ, an inhibitor blocking p53 translocation to mitochondria, increased COX biogenesis in older mice, but not in young mice. Our data suggest that physical exercise attenuates age-related changes in mitochondrial COX biogenesis and p53 activity targeting SCO2 and mitochondria, and thereby induces antisenescent and protective effects in cardiac muscle

PL-010 Chronic mild stress improves glucose homeostasis via myonectin-mediated suppression of sympathetic activity in high-fat diet-fed mice

Objective Recent studies suggest that chronic stress exposure can ameliorate the progression of diet-induced prediabetic disease, by inhibiting an increase in weight gain, caloric intake and efficiency and insulin resistance. To determine the underlying mechanism by which chronic stress improves the progression of type 2 diabetes, we developed a model of chronic mild stress in high-fat diet(HFD)-fed mice which are resistant to obesity and exhibit a healthy-like metabolic phenotype. Methods High-fat diet (HFD): 45% kcal derived from fat (Research Diets, Inc.).Mice experienced one stressor during the day and a different stressor during the night. Stressors were randomly chosen from the following list : cage tilt on a 45° angle for 1 to 16 h; food deprivation for 12 to 16 h; white noise for 1 to 16 h; strobe light illumination for 1 to 16 h; crowded housing; light cycle (continuous illumination) for 24 to 36 h; dark cycle (continuous darkness) for 24 to 36 h; water deprivation for 12 to 16 h; damp bedding (200 ml water poured into sawdust bedding) for 12 to 16 h.Recombinant adeno-associated virus (AAV): AAV9 vectors encoding myonectin under the control of the ubiquitous CMV promoter (AAV9-CMV-Vip) or an equal dose of the AAV9-CMV-null vector were delivered to C57BL/6 mice by the tail vein. Mice were deprived of food for 16 h and then subjected to test 7 days after AAV injection. Results Chronic stress improved glucose intolerance and sympathetic overactivity in HFD-fed mice. Chronic stress attenuated epinephrine(EPI)-stimulated glycerol release into blood in vivo and accelerated glycerol release from white adipose tissue followed by in vitro incubation with EPI. Chronic stress reduced plasma triglyceride but increased the levels of plasma insulin and myonectin. We further found that adeno-associated virus 9 (AAV9)-mediated myonectin overexpression improved glucose homeostasis and reduced epinephrine sensitivity. Myonectin overexpression reduced plasma norepinephrine, EPI and leptin levels, and increased insulin sensitivity in brown and white adipose tissue. Intense sympathetic activity with high-intensity running inhibited myonectin expression in skeletal muscle, whereas medium and low-intensity exercise running increased myonectin expression. Conclusions These findings suggest that chronic mild stress can improve glucose homeostasis via myonectin-mediated suppression of sympathetic activity in high-fat diet-fed mice

Increased Insulin Sensitivity and Distorted Mitochondrial Adaptations during Muscle Unloading

We aimed to further investigate mitochondrial adaptations to muscle disuse and the consequent metabolic disorders. Male rats were submitted to hindlimb unloading (HU) for three weeks. Interestingly, HU increased insulin sensitivity index (ISI) and decreased blood level of triglyceride and insulin. In skeletal muscle, HU decreased expression of pyruvate dehydrogenase kinase 4 (PDK4) and its protein level in mitochondria. HU decreased mtDNA content and mitochondrial biogenesis biomarkers. Dynamin-related protein (Drp1) in mitochondria and Mfn2 mRNA level were decreased significantly by HU. Our findings provide more extensive insight into mitochondrial adaptations to muscle disuse, involving the shift of fuel utilization towards glucose, the decreased mitochondrial biogenesis and the distorted mitochondrial dynamics

Understanding the Role of Exercise in Nonalcoholic Fatty Liver Disease: ERS-Linked Molecular Pathways

Nonalcoholic fatty liver disease (NAFLD) is globally prevalent and characterized by abnormal lipid accumulation in the liver, frequently accompanied by insulin resistance (IR), enhanced hepatic inflammation, and apoptosis. Recent studies showed that endoplasmic reticulum stress (ERS) at the subcellular level underlies these featured pathologies in the development of NAFLD. As an effective treatment, exercise significantly reduces hepatic lipid accumulation and thus alleviates NAFLD. Confusingly, these benefits of exercise are associated with increased or decreased ERS in the liver. Further, the interaction between diet, medication, exercise types, and intensity in ERS regulation is more confusing, though most studies have confirmed the benefits of exercise. In this review, we focus on understanding the role of exercise-modulated ERS in NAFLD and ERS-linked molecular pathways. Moderate ERS is an essential signaling for hepatic lipid homeostasis. Higher ERS may lead to increased inflammation and apoptosis in the liver, while lower ERS may lead to the accumulation of misfolded proteins. Therefore, exercise acts like an igniter or extinguisher to keep ERS at an appropriate level by turning it up or down, which depends on diet, medications, exercise intensity, etc. Exercise not only enhances hepatic tolerance to ERS but also prevents the malignant development of steatosis due to excessive ERS

Transcriptional Regulation by Nuclear Corepressors and PGC-1α: Implications for Mitochondrial Quality Control and Insulin Sensitivity

The peroxisome proliferator-activated receptors (PPARs) and estrogen-related receptor (ERRα) are ligand-activated nuclear receptors that coordinately regulate gene expression. Recent evidence suggests that nuclear corepressors, NCoR, RIP140, and SMRT, repress nuclear receptors-mediated transcriptional activity on specific promoters, and thus regulate insulin sensitivity, adipogenesis, mitochondrial number, and activity in vivo. Moreover, the coactivator PGC-1α that increases mitochondrial biogenesis during exercise and calorie restriction directly regulates autophagy in skeletal muscle and mitophagy in the pathogenesis of Parkinson's disease. In this paper, we discuss the PGC-1α’s novel role in mitochondrial quality control and the role of nuclear corepressors in regulating insulin sensitivity and interacting with PGC-1α