Skeletal Muscle Resistance Research Articles

Fetal hyperglycemia acutely induces persistent insulin resistance in skeletal muscle.

Offspring exposed in utero to maternal diabetes exhibit long-lasting insulin resistance, though the initiating mechanisms have received minimal experimental attention. Herein, we show that rat fetuses develop insulin resistance after only 2-day continuous exposure to isolated hyperglycemia starting on gestational day 18. Hyperglycemia-induced reductions in insulin-induced AKT phosphorylation localized primarily to fetal skeletal muscle. The skeletal muscle of hyperglycemia-exposed fetuses also exhibited impaired in vivo glucose uptake. To address longer term impacts of this short hyperglycemic exposure, neonates were cross-fostered and examined at 21 days postnatal age. Offspring formerly exposed to 2 days late gestation hyperglycemia exhibited mild glucose intolerance with insulin signaling defects localized only to skeletal muscle. Fetal hyperglycemic exposure has downstream consequences which include hyperinsulinemia and relative uteroplacental insufficiency. To determine whether these accounted for induction of insulin resistance, we examined fetuses exposed to late gestational isolated hyperinsulinemia or uterine artery ligation. Importantly, 2 days of fetal hyperinsulinemia did not impair insulin signaling in murine fetal tissues and 21-day-old offspring exposed to fetal hyperinsulinemia had normal glucose tolerance. Similarly, fetal exposure to 2-day uteroplacental insufficiency did not perturb insulin-stimulated AKT phosphorylation in fetal rats. We conclude that fetal exposure to hyperglycemia acutely produces insulin resistance. As hyperinsulinemia and placental insufficiency have no such impact, this occurs likely via direct tissue effects of hyperglycemia. Furthermore, these findings show that skeletal muscle is uniquely susceptible to immediate and persistent insulin resistance induced by hyperglycemia.

310-OR: Integrating Islet and Skeletal Muscle Enhancer RNA Signatures with Diverse Omics Data Enables Fine-Mapping of Diabetes and Related Trait GWAS Loci

Type 2 diabetes (T2D) results from interaction between genetic variation and environmental exposures that influence insulin secretion in pancreatic islets and insulin resistance in skeletal muscle and other peripheral tissues. Genome-wide association studies (GWAS) have identified hundreds of loci that modulate risk for T2D and related insulin traits. These loci mostly occur in non protein-coding regions which makes identification of causal regulatory single nucleotide polymorphisms (SNPs) and their associated mechanisms difficult. Previous studies show that many active enhancers are transcribed into enhancer RNA (eRNA). To identify actively transcribed regulatory elements with high genomic resolution and prioritize SNPs, we generated eRNA profiles using cap analysis of gene expression (CAGE) across 57 islet and 146 skeletal muscle samples and 15 replicates of the human beta cell line EndoC-BH1. We computationally integrated these CAGE profiles with ChIP-seq profiles for five histone marks along with open chromatin ATAC-seq profiles to generate high-resolution cross-tissue chromatin states. We identify multiple CAGE-enriched chromatin states that correspond to actively transcribed promoter or enhancer regions, which are supported by diverse patterns of chromatin activity across an orthogonal set of 127 human epigenome maps. We find 11,856 CAGE+ promoter regions across islets and muscle and 54% are shared across the two tissues, whereas we observe 1,712 CAGE+ eRNA regions and only 7% are shared, indicating the cell-identity nature of these eRNA states. We find significant enrichment for GWAS and eQTL SNPs to overlap these CAGE+ chromatin states. Finally, at a subset of loci, we validate these findings using a modified high-throughput STARR-seq assay where we observe allelic activity. These results provide the highest-resolution epigenome maps for islets and muscle to date, and we are using them to fine-map GWAS SNPs. Disclosure S. Parker: None. A. Varshney: None. Funding American Diabetes Association/Pathway to Stop Diabetes (1-14-INI-07 to S.P.); National Institutes of Health (1R01DK117960)

291-LB: Long Noncoding RNA H19 Improves Insulin Resistance in Skeletal Muscle by Regulating Heterogeneous Nuclear Ribonucleoprotein A1

Skeletal muscles play a pivotal role in glucose and lipid metabolism, and it has become a consensus that impaired lipid metabolism leads to ectopic lipid accumulation in skeletal muscles, which ultimately causes systematic insulin resistant. Previous reports have shown that down-regulation of long non-coding RNA(lncRNA) H19 in skeletal muscle impairs glucose metabolism. To better understand the impact of H19 on insulin resistance, here we focused on the relationships between H19 and fatty acid metabolism in skeletal muscle. In our research, decreased H19 expression was found in skeletal muscle of db/db mice compared with db/m mice. In vivo overexpression of H19 via injection of recombinant H19-adenovirus by tail-vein in db/db mice significantly improved glucose tolerance, lowered serum insulin, TG and skeletal muscle TG levels, which were consistent with improved skeletal muscle H&E and oil red staining contrast to vehicle-treated db/db mice, indicating that overexpression of H19 in db/db mice ameliorated ectopic lipid accumulation in skeletal muscle. As expected, similar effects were observed in vitro. Overexpression of H19 in muscle cells treated with palmitate acid reduced lipid contents,promoted mitochondrial synthesis and ATP productions,and increased the expression of fatty acid oxidation related genes and mitochondrial respiratory chain complexes.Finally, it was drawn from RNA pull-down and RIP that heterogeneous nuclear ribonucleoprotein A1(hnRNPA1)is the target gene of H19, whose function in mitochondria has not been reported yet. Furthermore, we identified PGC1a and CPT1b were among the target genes of hnRNPA1 by searching widely used bioinformatics databases and preliminary RIP validation. Based on these results, we put forth our hypothesis that lncRNA H19 could improve insulin resistance by promoting mitochondrial biosynthesis and fatty acid oxidation in skeletal muscle, through regulating hnRNPA1. Disclosure X. Lin: None. W. Gui: None. H. Li: None. Funding National Natural Science Foundation of China (81873653)

33-OR: Intact Muscle Insulin Signaling in Patients with a Gain-of-Function LRP5 Mutation Despite Decreased Insulin Sensitivity

Type 2 diabetes (T2D) is characterized by insulin resistance in skeletal muscle. GWAS studies have shown a link between T2D and Wnt signalling, and there is evidence that mutations in LRP5, a co-receptor in the canonical Wnt signaling pathway, may alter insulin sensitivity. Here, we investigated whether a gain-of-function (GoF) LRP5 mutation causing high bone mass is accompanied by changes in insulin sensitivity and abnormalities in the insulin signalling pathway in skeletal muscle. Skeletal muscle biopsies were obtained before and after a euglycemic-hyperinsulinemic clamp (3-h, 40 mU/min/m2) from 13 patients with GoF LRP5 mutation (T253I) and 14 age, and weight-matched controls. Protein abundance/phosphorylation of key mediators of insulin signalling to glucose transport and glycogen synthesis, such as Akt, TBC1D4, glycogen synthase kinase 3 (GSK3α and ß) and Erk1/2 were investigated. Insulin sensitivity measured as insulin-stimulated glucose disposal (Rd) was reduced (25%) in GoF LRP5 mutation carriers (p<0.05). However, the insulin-stimulated phosphorylation of Akt at Ser473 and Thr308, TBC1D4 at Thr642 and Erk1/2 was fully intact in skeletal muscle of GoF LRP5 mutation carriers. Moreover, both the basal and the insulin-mediated inhibitory phosphorylation of GSK3 (α and ß) were unaltered in muscle of LRP5 mutation carriers compared with controls. To further investigate the molecular consequences of this GoF LRP5 mutation, we are currently investigating key regulators of Wnt signalling (activity of ß-catenin and CaMKII) and performing RNA-sequencing in muscle. In conclusion, our data demonstrate that reduced insulin sensitivity in GoF LRP5 mutation carriers is not explained by abnormalities in the classical components of the insulin signalling cascade in skeletal muscle. This suggests that the GOF LRP5 mutation negatively affects insulin sensitivity by other mechanisms possible involving over-activation in Wnt signalling. Disclosure R. Sabaratnam: None. J. Lauterlein: None. M.F. Nielsen: None. K. Hojlund: None.

294-LB: Transcriptome-Wide Signals Converge Upon Mitochondrial Function in Developmental Stages of Type 2 Diabetes in Human Skeletal Muscle

Skeletal muscle is a major contributor to overall glucose homeostasis in the body and together with the liver is chiefly responsible for the whole-body insulin sensitivity. However, few studies have investigated the transcriptional signature of insulin resistance in skeletal muscles, at different stages of T2D development. Thus, the aim of this study was to identify the transcriptional signature of dysregulated glucose metabolism in skeletal muscle in humans by profiling the transcriptome with next-generation sequencing (NGS). We selected 40 sedentary males, aged 40-65 years, with BMI 26-33 kg/m2, diagnosed into three groups: 18 normoglycemics, 14 prediabetics and 8 subjects with type 2 diabetes. mRNA was extracted from biopsied Vastus lateralis muscles and sequenced with Illumina HiSeq 4000.Comparing normoglycemic and diabetic subjects revealed 4404 significantly differentially expressed (DE) genes, which represents around 30% of all mapped genes in studied samples. For the comparison between type 2 diabetics and prediabetics, the number of DE genes was 4055 genes. Interestingly we found no significantly DE genes between normoglycemics and prediabetics. KEGG enrichment analysis revealed that DE pathways between diabetics vs. nondiabetics were enriched for pathways related to oxidative phosphorylation, thermogenesis, nonalcoholic fatty liver disease and neurological diseases, which share the same underlying mechanism: mitochondrial dysfunction.These results indicate that skeletal muscle contributes most to overt T2D, with a possible indication that the liver contributes most to early stages the disease development. Differences observed between type 2 diabetics and the two remaining groups, occurring primarily in oxidative phosphorylation and electron transport chain pathways suggests that mitochondrial dysfunction plays the predominant role in the pathogenesis of T2D. Disclosure L. Szczerbinski: None. M. Niemira: None. K. Szczerbinski: None. U. Puchta: None. E. Siewiec: None. A. Citko: None. J. Zapolska: None. M. Taylor: None. S. Larsen: None. M. Gorska: None. A. Kretowski: None.

1777-P: Acute Hyperbaric Oxygen (HBO) Treatment Increases Hepatic Insulin Sensitivity and Muscle Reactive Oxygen Species in Type 2 Diabetes

Diabetes mellitus type 2 (T2D) is characterized by insulin resistance of liver and skeletal muscle, which is at least partly due to impaired muscle mitochondrial function. Long-term HBO therapy, as applied for treating the diabetic foot syndrome, has been shown to improve blood glucose concentrations. To study the underlying mechanisms, we examined the short-term effect of HBO treatment on insulin sensitivity and mitochondrial oxidative capacity and production of reactive oxygen species (ROS) in a randomized, double blinded, placebo-controlled trial. Volunteers with type 2 diabetes (n=4; age 61±4 years; body mass index 30.2±2.2 kg/m2; HbA1c 7.0±0.7%) underwent two 2-hours sessions in a hyperbaric chamber of once with application of 100% oxygen (1.4 bar over atmospheric pressure [barO] for 20 min; HBO) and once with ambient air as placebo (21% oxygen; 1.4 barO; PLC). Tissue-specific insulin sensitivity was assessed by hyperinsulinemic-euglycemic clamps with stable isotope dilution, while muscle oxygen fluxes were measured with high resolution respirometry. After HBO treatment, hepatic insulin sensitivity was higher (insulin-mediated suppression of endogenous glucose production: 69.7±0.1% vs. PLC 55.3±3.2%, p<0.05), whereas peripheral insulin sensitivity was comparable (2.0±0.3 vs. 2.2±0.3 mg*kg-1*min-1, p=0.24). Muscle ROS production markedly increased after HBO (increase in ROS: 73±3% vs. PLC -14±5%; p<0.005), while mitochondrial oxidative capacity only tended to be lower after HBO. In conclusion, a single session of HBO treatment already improves hepatic insulin resistance and stimulates muscle ROS release, which may activate antioxidative defense, both of which could contribute to improved glycaemia upon chronic treatment. Disclosure T. Sarabhai: None. E. Anadol-Schmitz: None. H. Katsuyama: None. D.F. Markgraf: Research Support; Self; Sanofi. L. Busch: None. R. Guthoff: None. J. Schneppendahl: None. M. Roden: Advisory Panel; Self; Boehringer Ingelheim Pharmaceuticals, Inc., Poxel SA, Servier. Board Member; Self; Eli Lilly and Company. Research Support; Self; Boehringer Ingelheim Pharmaceuticals, Inc., Sanofi. Speaker's Bureau; Self; Novo Nordisk Inc.

Exercise Induction of Key Transcriptional Regulators of Metabolic Adaptation in Muscle Is Preserved in Type 2 Diabetes.

Type 2 diabetes (T2D) is characterized by insulin resistance in skeletal muscle. Regular exercise improves insulin sensitivity, mitochondrial function, and energy metabolism. Thus, an impaired response to exercise may contribute to insulin resistance. We hypothesized that key transcriptional regulators of metabolic adaptation to exercise show an attenuated response in skeletal muscle in T2D. Skeletal muscle biopsies were obtained from 13 patients with T2D and 14 age- and weight-matched controls before, immediately after 1 hour acute exercise (70% maximal pulmonary oxygen uptake), and 3 hours into recovery to examine mRNA expression of key transcription factors and downstream targets and activity of key upstream kinases underlying the metabolic adaptation to exercise. Acute exercise increased gene expression of the nuclear hormone receptor 4A (NR4A) subfamily (∼4- to 36-fold) and other key transcription factors, including ATF3, EGR1, JUNB, SIK1, PPARA, and PPARG (∼1.5- to 12-fold), but with no differences between groups. The expression of NR4A1 (approximately eightfold) and NR4A3 (∼75-fold) was further increased 3 hours into recovery, whereas most muscle transcripts sustained elevated or returned to basal levels, again with no differences between groups. Muscle expression of HKII and SLC2A4 and hexokinase II protein content were reduced in patients with T2D. The phosphorylation of p38 MAPK, Erk1/2, Ca2+/calmodulin-dependent kinase II, and cAMP-responsive element-binding protein was equally increased in response to exercise and/or recovery in both groups. Acute exercise elicits a pronounced and overall similar increase in expression of key transcription factors and activation of key upstream kinases involved in muscle metabolic adaptation to exercise in patients with T2D and weight-matched controls.

Proinflammatory NFkB signalling promotes mitochondrial dysfunction in skeletal muscle in response to cellular fuel overloading

Sustained nutrient (fuel) excess, as occurs during obesity and diabetes, has been linked to increased inflammation, impaired mitochondrial homeostasis, lipotoxicity, and insulin resistance in skeletal muscle. Precisely how mitochondrial dysfunction is initiated and whether it contributes to insulin resistance in this tissue remains a poorly resolved issue. Herein, we examine the contribution that an increase in proinflammatory NFkB signalling makes towards regulation of mitochondrial bioenergetics, morphology, and dynamics and its impact upon insulin action in skeletal muscle cells subject to chronic fuel (glucose and palmitate) overloading. We show sustained nutrient excess of L6 myotubes promotes activation of the IKKβ-NFkB pathway (as judged by a six-fold increase in IL-6 mRNA expression; an NFkB target gene) and that this was associated with a marked reduction in mitochondrial respiratory capacity (>50%), a three-fold increase in mitochondrial fragmentation and 2.5-fold increase in mitophagy. Under these circumstances, we also noted a reduction in the mRNA and protein abundance of PGC1α and that of key mitochondrial components (SDHA, ANT-1, UCP3, and MFN2) as well as an increase in cellular ROS and impaired insulin action in myotubes. Strikingly, pharmacological or genetic repression of NFkB activity ameliorated disturbances in mitochondrial respiratory function/morphology, attenuated loss of SDHA, ANT-1, UCP3, and MFN2 and mitigated the increase in ROS and the associated reduction in myotube insulin sensitivity. Our findings indicate that sustained oversupply of metabolic fuel to skeletal muscle cells induces heightened NFkB signalling and that this serves as a critical driver for disturbances in mitochondrial function and morphology, redox status, and insulin signalling.

TFAM Enhances Fat Oxidation and Attenuates High-Fat Diet-Induced Insulin Resistance in Skeletal Muscle.

Diet-induced insulin resistance (IR) adversely affects human health and life span. We show that muscle-specific overexpression of human mitochondrial transcription factor A (TFAM) attenuates high-fat diet (HFD)-induced fat gain and IR in mice in conjunction with increased energy expenditure and reduced oxidative stress. These TFAM effects on muscle are shown to be exerted by molecular changes that are beyond its direct effect on mitochondrial DNA replication and transcription. TFAM augmented the muscle tricarboxylic acid cycle and citrate synthase facilitating energy expenditure. TFAM enhanced muscle glucose uptake despite increased fatty acid (FA) oxidation in concert with higher β-oxidation capacity to reduce the accumulation of IR-related carnitines and ceramides. TFAM also increased pAMPK expression, explaining enhanced PGC1α and PPARβ, and reversing HFD-induced GLUT4 and pAKT reductions. TFAM-induced mild uncoupling is shown to protect mitochondrial membrane potential against FA-induced uncontrolled depolarization. These coordinated changes conferred protection to TFAM mice against HFD-induced obesity and IR while reducing oxidative stress with potential translational opportunities.

Mitochondrial (Dys)function and Insulin Resistance: From Pathophysiological Molecular Mechanisms to the Impact of Diet.

Mitochondrial dysfunction has been implicated in the pathogenesis of insulin resistance, the hallmark of type 2 diabetes mellitus (T2DM). However, the cause-effect relationship remains to be fully elucidated. Compelling evidence suggests that boosting mitochondrial function may represent a valuable therapeutic tool to improve insulin sensitivity. Mitochondria are highly dynamic organelles, which adapt to short- and long-term metabolic perturbations by undergoing fusion and fission cycles, spatial rearrangement of the electron transport chain complexes into supercomplexes and biogenesis governed by peroxisome proliferator-activated receptor γ co-activator 1α (PGC 1α). However, these processes appear to be dysregulated in type 2 diabetic individuals. Herein, we describe the mechanistic link between mitochondrial dysfunction and insulin resistance in skeletal muscle alongside the intracellular pathways orchestrating mitochondrial bioenergetics. We then review current evidence on nutritional tools, including fatty acids, amino acids, caloric restriction and food bioactive derivatives, which may enhance insulin sensitivity by therapeutically targeting mitochondrial function and biogenesis.

Exercise training restores the myogenic response in skeletal muscle resistance arteries and corrects peripheral edema in rats with heart failure.

Impairment of the myogenic response can affect capillary hydrostatic pressure and contribute to peripheral edema and exercise intolerance, which are markers of heart failure (HF). The aim of this study was to assess the effects of exercise training (ET) on myogenic response in skeletal muscle resistance arteries and peripheral edema in HF rats, focusing on the potential signaling pathways involved in these adjustments. Male Wistar rats were submitted to either coronary artery occlusion or a sham-operated surgery. After 4 wk, an exercise test was performed, and the rats were divided into the following groups: untrained normal control (UNC) and untrained HF (UHF) and exercise- trained (on treadmill, 50-60% of maximal capacity) NC (TNC) and exercise-trained HF (THF). Caudal tibial artery (CTA) myogenic response was impaired in UHF compared with UNC, and ET restored this response in THF to NC levels and increased it in TNC. Rho kinase (ROCK) inhibitor abolished CTA myogenic response in the untrained and blunted it in exercise-trained groups. CTA-stored calcium (Ca2+) mobilization was higher in exercise-trained rats compared with untrained rats. The paw volume was higher in UHF rats, and ET decreased this response compared with UNC. Myogenic constriction was positively correlated with maximal running distance and negatively correlated with paw volume. The results demonstrate, for the first time, that HF impairs the myogenic response in skeletal muscle arteries, which contributes to peripheral edema in this syndrome. ET restores the myogenic response in skeletal muscle arteries improving Ca2+ sensitization and handling. Additionally, this paradigm also improves peripheral edema and exercise intolerance. NEW & NOTEWORTHY The novel and main finding of the present study is that moderate intensity exercise training restores the impaired myogenic response of skeletal muscle resistance arteries, exercise intolerance and peripheral edema in rats with heart failure. These results also show for the first time to our knowledge that exercise training improving calcium sensitization through the ROCK pathway and enhancing intracellular calcium handling could contribute to restoration of flow autoregulation to skeletal muscle in heart failure.

Inhibitory effect of 17β‑estradiol on triglyceride synthesis in skeletal muscle cells is dependent on ESR1 and not ESR2.

The present study aimed to investigate the inhibitory effects and the mechanisms underlying 17β-estradiol (E2) effects on triglyceride synthesis and insulin resistance in skeletal muscle tissues and cells. Ovariectomy (OVX) was performed on 6-month-old female rats treated with or without E2. Subsequently, various serum biochemical markers were measured. Additionally, pathological alterations of the uterus, liver and skeletal muscle were analyzed, and the content of triglycerides (TG) in muscle was detected. Differentiated myotubes formed by C2C12 cells were treated with palmitic acid (PA) or pretreated with E2, estrogen receptor (ESR) 1 agonist propylpyrazoletriol (PPT) and ESR2 agonist diarylpropionitrile (DPN). Subsequently, the mRNA or protein expression levels of ESR1/2, peroxisome proliferator activated receptor α (PPARα), CD36 molecule (CD36), fatty acid synthase (FASN), perilipin 2 (PLIN2), phosphorylated acetyl-CoA carboxylase α (p-ACACA), p-AKT serine/threonine kinase (p-AKT) and p-mitogen-activated protein kinase 8 (p-MAPK8) were analyzed in skeletal muscle or in C2C12 cells by reverse transcription-semi-quantitative polymerase chain reaction and western blotting. The present results suggested that treatment with E2 inhibited OVX-induced body weight gain, TG accumulation and insulin resistance. The protein or mRNA expression levels of ESR1, CD36, PPARα, p-ACACA and p-AKT were decreased, whereas the protein or mRNA expression levels of ESR2, PLIN2, FASN and p-MAPK8 were increased in the OVX group. Of note, treatment with E2 restored the expression levels of the aforementioned factors. In C2C12 cells, treatment with E2 or PPT reversed the alterations induced by treatment with PA. In contrast, pretreatment with DPN did not influence the effect of PA. Collectively, E2 was able to interact with ESR1, thus activating the CD36-PPARα pathway, decreasing the level of TG in the muscles and improving insulin resistance in skeletal muscles and C2C12 cells.

Insulin-dependent GLUT4 trafficking is not regulated by protein SUMOylation in L6 myocytes

Type-II Diabetes Mellitus (T2DM) is one of the fastest growing public health issues today, consuming 12% of worldwide health budgets and affecting an estimated 400 million people. One of the key pathological traits of this disease is insulin resistance at ‘glucose sink’ tissues (mostly skeletal muscle), and this remains one of the features of this disease most intractable to therapeutic intervention. Several lines of evidence have implicated the post-translational modification, SUMOylation, in insulin signalling and insulin resistance in skeletal muscle. In this study, we examined this possibility by manipulation of cellular SUMOylation levels using multiple different tools, and assaying the effect on insulin-stimulated GLUT4 surface expression in differentiated L6 rat myocytes. Although insulin stimulation of L6 myocytes produced a robust decrease in total cellular SUMO1-ylation levels, manipulating cellular SUMOylation had no effect on insulin-responsive GLUT4 surface trafficking using any of the tools we employed. Whilst we cannot totally exclude the possibility that SUMOylation plays a role in the insulin signalling pathway in human health and disease, our data strongly argue that GLUT4 trafficking in response to insulin is not regulated by protein SUMOylation, and that SUMOylation does not therefore represent a viable therapeutic target for the treatment of insulin resistance.

Asprosin attenuates insulin signaling pathway through PKCδ-activated ER stress and inflammation in skeletal muscle.

It has been reported that asprosin is a novel adipokine which is augmented in mice and humans with type 2 diabetes (T2DM). Asprosin stimulates hepatic gluconeogenesis under fasting conditions. However, the roles of asprosin in inflammation, endoplasmic reticulum (ER) stress, and insulin resistance in skeletal muscle has not been studied. In the currents study, elevated levels of asprosin expression were observed in adipocytes under hyperlipidemic conditions. Treatment of C2C12 myocytes with asprosin-induced ER stress markers (phosphorylated inositol-requiring enzyme 1 and eukaryotic initiation factor 2, and CHOP expression) as well as inflammation markers (interleukin-6 expression, phosphorylated IκB, and nuclear translocated nuclear factor-κβ). Finally, asprosin treatment promoted exacerbation of insulin sensitivity as determined by levels of insulin receptor substrate 1 and Akt phosphorylation as well as glucose uptake. Moreover, treatment of asprosin augmented protein kinase C-δ (PKCδ) phosphorylation and nuclear translocation, but suppressed messenger RNA expression of sarcoplasmic reticulum Ca2+ ATPase 2b in both C2C12 myocytes and in mouse soleus skeletal muscle. These asprosin-induced effects were markedly decreased in small interfering (si) RNA-mediated PKCδ-knockdown in C2C12 myocytes. These results suggest that asprosin results in impairment of insulin sensitivity in skeletal muscle through PKCδ-associated ER stress/inflammation pathways and may be a valuable strategy for management of insulin resistance and T2DM.

Carbenoxolone ameliorates insulin sensitivity in obese mice induced by high fat diet via regulating the IκB-α/NF-κB pathway and NLRP3 inflammasome

The characteristic feature of obesity and insulin resistance is chronic low-grade inflammation. Nod-Like Receptor Pyrin 3 (NLRP3) inflammasome plays a central role in obesity-induced insulin resistance. However, how does Carbenoxolone (CBX) play its role in ameliorating insulin resistance in peripheral tissues of obese mice induced by high-fat diet (HFD) remains unknown. In our study, we explored the molecular mechanism of CBX in improving insulin resistance in liver and skeletal muscle in mice induced by the HFD. Our results revealed that in the CBX group, a significant decrease in fasting blood glucose, insulin and HOMA-IR score were observed. CBX could attenuate intracellular lipid accumulation and inflammation aggravation in liver and skeletal muscle. Besides, treatment with CBX could significantly reduce expressions of p-IκB-α, p-NF-κB, p-IRS-1, NLRP3 and inflammatory factors, increase expressions of p-PI3K and p-AKT. Therefore, CBX could dramatically improve insulin resistance in liver and skeletal muscle in mice induced by the high-fat diet. In conclusions, we demonstrate that CBX has a significant protective effect on diet-induced obesity in mice. The potential mechanisms include inhibiting IκB-α/NF-κB pathway, restricting the production of NLRP3 inflammasome and other inflammatory factors, reducing the expression of p-IRS-1, increasing the expressions of p-PI3K and p-AKT, thus ameliorating insulin resistance in liver and skeletal muscle of high-fat diet mice. Therefore CBX is an active agent against diet-induced obesity and is given the opportunity for the treatment of obesity related diseases.

Maternal Roux-en-Y gastric bypass impairs insulin action and endocrine pancreatic function in male F1 offspring.

Obesity is predominant in women of reproductive age. Roux-en-Y gastric bypass (RYGB) is the most common bariatric procedure that is performed in obese women for weight loss and metabolic improvement. However, some studies suggest that this procedure negatively affects offspring. Herein, using Western diet (WD)-obese female rats, we investigated the effects of maternal RYGB on postnatal body development, glucose tolerance, insulin secretion and action in their adult male F1 offspring. Female Wistar rats consumed a Western diet (WD) for 18weeks, before being submitted to RYGB (WD-RYGB) or SHAM (WD-SHAM) operations. After 5weeks, WD-RYGB and WD-SHAM females were mated with control male breeders, and the F1 offspring were identified as: WD-RYGB-F1 and WD-SHAM-F1. The male F1 offspring of WD-RYGB dams exhibited decreased BW, but enhanced total nasoanal length gain. At 120days of age, WD-RYGB-F1 rats displayed normal fasting glycemia and glucose tolerance but demonstrated reduced insulinemia and higher glucose disappearance after insulin stimulus. In addition, these rodents presented insulin resistance in the gastrocnemius muscle and retroperitoneal fat, as judged by lower Akt phosphorylation after insulin administration, but an increase in this protein in the liver. Finally, the islets from WD-RYGB-F1 rats secreted less insulin in response to glucose and displayed increased β-cell area and mass. RYGB in WD dams negatively affected their F1 offspring, leading to catch-up growth, insulin resistance in skeletal muscle and white fat, and β-cell dysfunction. Therefore, our data are the first to demonstrate that the RYGB in female rats may aggravate the metabolic imprinting induced by maternal WD consumption, in their male F1 descendants. However, since we only used male F1 rats, further studies are necessary to demonstrate if such effect may also occur in female F1 offspring from dams that underwent RYGB operation.

Molecular regulation of skeletal muscle mass and the contribution of nitric oxide: A review.

A variety of internal and external factors such as exercise, nutrition, inflammation, and cancer‐associated cachexia affect the regulation of skeletal muscle mass. Because skeletal muscle functions as a crucial regulator of whole body metabolism, rather than just as a motor for locomotion, the enhancement and maintenance of muscle mass and function are required to maintain health and reduce the morbidity and mortality associated with diseases involving muscle wasting. Recent studies in this field have made tremendous progress; therefore, identification of the mechanisms that regulate skeletal muscle mass is necessary for the physical and nutritional management of both athletes and patients with muscle wasting disease. In this review, we present an overall picture of the interactions regulating skeletal muscle mass, particularly focusing on the insulin‐like growth factor‐I (IGF‐I)/insulin‐Akt‐mammalian target of rapamycin (mTOR) pathway, skeletal muscle inactivity, and endurance and resistance exercise. We also discuss the contribution of nitric oxide (NO) to the regulation of skeletal muscle mass based on the current knowledge of the novel role of NO in these processes.

Age‐Related Changes in Skeletal Muscle and Small Mesenteric Arterial Function in Spontaneously Hypertensive Rats

Essential hypertension is a chronic idiopathic medical condition that is characterized by elevated arterial blood pressure and is associated with increased risk of cardiovascular disease. Endothelial dysfunction is associated with increased age and a casual factor for the development of hypertension. Therefore, we hypothesized that endothelial dysfunction would be exacerbated by age to a greater extent in spontaneously hypertensive rats (SHRs) compared to Wistar‐Kyoto rats (WKY) control rats. Vascular function was assessed using pressure myography in skeletal muscle and mesenteric resistance arteries isolated from SHR and WKY across three age groups: young (16–17 weeks, n = 4/group), aged (62–64 weeks, n = 4/group) and old (88–100 weeks, n = 5/group). Phenylephrine‐induced vasoconstriction responses in either of the resistance arteries were not significantly different between WKY and SHR at any of the age groups. Endothelial‐dependent vasodilation in mesenteric arteries was not significantly different between WKY and SHR across age groups either. However, endothelium‐dependent vasodilation in the skeletal muscle resistance arteries was significantly enhanced in the young SHR compared to WKY, but not in the aged or old rats. Moreover, endothelial‐independent vasodilation in mesenteric arteries was significantly enhanced in SHR compared to WKY across all age groups, while this difference was only observed in the skeletal muscle resistance arteries of the young rats. Contrary to our hypothesis, these data suggest that endothelial dysfunction is not exacerbated by age in the resistance arteries of SHR.Support or Funding InformationResearch support: National Institute of Health grants: R01 HL137769 (JP), R01 HL088105 (LM‐L).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

In Vivo Targeting of the Endothelium to Improve Vascular Function in a Rodent Model of Type 2 Diabetes

Endothelial dysfunction has been identified as an early event in the progression of cardiovascular complications (e.g. hypertension, cardiomyopathy, stroke) commonly observed in patients with Type 2 Diabetes (T2D). Importantly, reduction/reversal of endothelial dysfunction is expected to mitigate the cardiovascular deficits associated with T2D. Small‐ and intermediate‐conductance, Ca2+‐activated K+ channels (KCa2.3 and KCa3.1, respectively) are prominently expressed in the vascular endothelium, and pharmacologic activators of these channels (e.g. SKA‐31) induce robust vasodilation upon acute exposure in isolated arteries and intact animals. However, the effects of prolonged in vivo administration of KCa activators such as SKA‐31 have not been examined to date. In our current study, we have hypothesized that daily administration of SKA‐31 (10 and 30 mg/kg, I.P. injection) to male, Type 2 Diabetic Goto‐Kakizaki rats (T2D GK, 14 weeks of age) for 12 weeks would ameliorate diabetes‐related cardiovascular deficits. In isolated mesenteric resistance arteries examined by pressure myography, we observed that SKA‐31 treatment significantly improved endothelium‐dependent vasodilation in response to 0.3 μM acetylcholine (ACh) (percent of maximal dilation = 37.7 ± 7.3% and 33.6 ± 6.2% for 10 and 30 mg/kg treatments) compared with vehicle‐treated animals (20.7 ± 3.1%) (means ± S.D., n = 6 animals/group). Similar effects were observed for 0.3 μM bradykinin (BK) (evoked vasodilation = 54.1 ±7.8% and 47.9 ± 6.6% for 10 and 30 mg/kg SKA‐31, respectively) compared with vehicle (33.6 ±6.1%). SKA‐31 administration also reduced alpha1‐adrenergic (phenylephrine, 1 μM) stimulated constriction in 10 and 30 mg/kg treated T2D GK rats (percent of maximal contraction = 37.1 ± 5.0% and 36.1 ± 3.7%, respectively) compared with vehicle treated animals (46.6 ± 4.5%, n = 6 per group). Western blot analyses further revealed that SKA‐31 administration increased the expression of mesenteric KCa2.3 and KCa3.1 channels and associated cell‐signaling components (i.e. type 1 IP3 receptor and the SERCA2 Ca2+‐ATPase) that contribute to endothelium‐dependent, agonist‐evoked vasodilation. In contrast, SKA‐31 treatment did not augment ACh and BK evoked vasodilation in myogenically active, cremaster skeletal muscle resistance arteries from the same animals, compared with the vehicle treated group. Collectively, these data demonstrate that in vivo administration of an endothelial KCa channel activator can selectively improve vascular function in the setting of T2D.Support or Funding InformationThis study was supported by research funding to APB from the Canadian Institutes of Health Research (MOP‐142467) and the Natural Sciences and Engineering Research Council of Canada (RGPIN‐2017‐04116), and to HW from the National Institutes of Health (R21 NS101876).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Integrin‐mediated Vasoconstrictor Function Declines with Age in Skeletal Muscle Resistance Arteries

IntroductionVasoconstrictor responses decline with age in skeletal muscle resistance arteries. The mechanism for functional regulation of resistance artery contractility is not fully understood but has been proposed to involve impaired integrin signaling in vascular smooth muscle.ObjectiveThe purpose of this study was to test the hypothesis that RGD binding integrin‐mediated constrictor responses decline with age in skeletal muscle resistance arteries.MethodsSoleus muscle feed arteries (SFA) were isolated from young (4 mo) and old (24 mo) male Fischer 344 rats. SFA were cannulated with glass micropipettes and pressurized to 90 cm H2O for assessment of vasoconstrictor function. Vasoconstrictor responses were assessed using an a1 selective adrenergic agonist (phenylephrine; PE). To test the role of integrin signaling, constrictor responses were assessed in the absence or presence of an integrinblocking peptide (RGD) or a control peptide (RGE).ResultsConstrictor responses to PE were significantly impaired in old SFA relative to young SFA. In the presence of the RGD soluble peptide, constrictor responses to PE were significantly reduced in young SFA such that the young SFA responded like control (no treatment) old SFA. In the presence of the RGD peptide, constrictor responses to PE were further reduced in old SFA. Constrictor responses to PE were not altered by RGE in young or old SFA.ConclusionThe results of this study indicate that RGD binding integrin‐mediated constrictor function declines with age in SFA.Support or Funding InformationResearch support: National Institute on Aging, Sydney and J.L. Huffines Institute of Sports Medicine and Human Performance.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.