IL-15 improves skeletal muscle oxidative metabolism and glucose uptake in association with increased respiratory chain supercomplex formation and AMPK pathway activation

Background

IL-15 is believed to play a role in the beneficial impact of exercise on muscle energy metabolism. However, previous studies have generally used supraphysiological levels of IL-15 that do not represent contraction-induced IL-15 secretion.

Palmitoylation is a post-translational modification of Alix regulating the membrane organization of exosome-like small extracellular vesicles

Background

Virtually all cell types have the capacity to secrete nanometer-sized extracellular vesicles, which have emerged in recent years as potent signal transducers and cell-cell communicators. The multifunctional protein Alix is a bona fide exosomal regulator and skeletal muscle cells can release Alix-positive nano-sized extracellular vesicles, offering a new paradigm for understanding how myofibers communicate within skeletal muscle and with other organs. S-palmitoylation is a reversible lipid post-translational modification, involved in different biological processes, such as the trafficking of membrane proteins, achievement of stable protein conformations, and stabilization of protein interactions.

The absence of dysferlin induces the expression of functional connexin-based hemichannels in human myotubes

Background

Mutations in the gene encoding for dysferlin cause recessive autosomal muscular dystrophies called dysferlinopathies. These mutations induce several alterations in skeletal muscles, including, inflammation, increased membrane permeability and cell death. Despite the fact that the etiology of dysferlinopathies is known, the mechanism that explains the aforementioned alterations is still elusive. Therefore, we have now evaluated the potential involvement of connexin based hemichannels in the pathophysiology of dysferlinopathies.

Results

Human deltoid muscle biopsies of 5 Chilean dysferlinopathy patients exhibited the presence of muscular connexins (Cx40.1, Cx43 and Cx45). The presence of these connexins was also observed in human myotubes derived from immortalized myoblasts derived from other patients with mutated forms of dysferlin. In addition to the aforementioned connexins, these myotubes expressed functional connexin based hemichannels, evaluated by ethidium uptake assays, as opposed to myotubes obtained from a normal human muscle cell line, RCMH. This response was reproduced in a knock-down model of dysferlin, by treating RCMH cell line with small hairpin RNA specific for dysferlin (RCMH-sh Dysferlin). Also, the presence of P2X₇ receptor and the transient receptor potential channel, TRPV2, another Ca²⁺ permeable channels, was detected in the myotubes expressing mutated dysferlin, and an elevated resting intracellular Ca²⁺ level was found in the latter myotubes, which was in turn reduced to control levels in the presence of the molecule D4, a selective Cx HCs inhibitor.

Conclusions

The data suggests that dysferlin deficiency, caused by mutation or downregulation of dysferlin, promotes the expression of Cx HCs. Then, the de novo expression Cx HC causes a dysregulation of intracellular free Ca²⁺ levels, which could underlie muscular damage associated to dysferlin mutations. This mechanism could constitute a potential therapeutical target in dysferlinopathies.

     

HMGB1/autophagy pathway mediates the atrophic effect of TGF-β1 in denervated skeletal muscle

Background

Transforming growth factor beta 1 (TGF-β1) is a classical modulator of skeletal muscle and regulates several processes, such as myogenesis, regeneration and muscle function in skeletal muscle diseases. Skeletal muscle atrophy, characterized by the loss of muscle strength and mass, is one of the pathological conditions regulated by TGF-β1, but the underlying mechanism involved in the atrophic effects of TGF-β1 is not fully understood.

Methods

Mice sciatic nerve transection model was created and gastrocnemius were analysed by western blot, immunofluorescence staining and fibre diameter quantification after 2 weeks. Exogenous TGF-β1 was administrated and high-mobility group box-1 (HMGB1), autophagy were blocked by siRNA and chloroquine (CQ) respectively to explore the mechanism of the atrophic effect of TGF-β1 in denervated muscle. Similar methods were performed in C2C12 cells.

Results

We found that TGF-β1 was induced in denervated muscle and it could promote atrophy of skeletal muscle both in vivo and in vitro, up-regulated HMGB1 and increased autophagy activity were also detected in denervated muscle and were further promoted by exogenous TGF-β1. The atrophic effect of TGF-β1 could be inhibited when HMGB1/autophagy pathway was blocked.

Conclusions

Thus, our data revealed that TGF-β1 is a vital regulatory factor in denervated skeletal muscle in which HMGB1/ autophagy pathway mediates the atrophic effect of TGF-β1. Our findings confirmed a new pathway in denervation-induced skeletal muscle atrophy and it may be a novel therapeutic target for patients with muscle atrophy after peripheral nerve injury.

     

Inactivation of glycogen synthase kinase 3β (GSK-3β) enhances mitochondrial biogenesis during myogenesis

Background

Mitochondrial biogenesis is crucial for myogenic differentiation and regeneration of skeletal muscle tissue and is tightly controlled by the peroxisome proliferator-activated receptor-γ co-activator 1 (PGC-1) signaling network. In the present study, we hypothesized that inactivation of glycogen synthase kinase (GSK)-3β, previously suggested to interfere with PGC-1 in non-muscle cells, potentiates PGC-1 signaling and the development of mitochondrial biogenesis during myogenesis, ultimately resulting in an enhanced myotube oxidative capacity.

S-allyl cysteine inhibits TNFα-induced skeletal muscle wasting through suppressing proteolysis and expression of inflammatory molecules

Background

Elevated levels of inflammatory molecules are key players in muscle wasting/atrophy leading to human morbidity. TNFα is a well-known pro-inflammatory cytokine implicated in the pathogenesis of muscle wasting under diverse clinical settings. S-allyl cysteine (SAC), an active component of garlic (Allium sativum), has established anti-oxidant and anti-inflammatory effects in various cell types. However, the impact of SAC on skeletal muscle pathology remains unexplored. Owing to the known anti-inflammatory properties of SAC, we investigated whether pre-treatment with SAC has a protective role in TNFα-induced atrophy in cultured myotubes.

Stabilization of G-quadruplex structure on vascular endothelial growth factor gene promoter depends on CpG methylation site and cation type

Background

DNA methylation at the 5-position of cytosine is an epigenetic modification of CpG dinucleotides. In addition to CpG methylation, the G-quadruplex (G4) structure has been reported as a regulator of gene expression. The identification of G4 forming sequences in CpG islands suggests an involvement of CpG-methylated G4 structures in biological processes; however, few reports have addressed the effects of CpG methylation on G4 structure.

BMS309403 Stimulates Glucose Uptake in Myotubes through Activation of AMP-Activated Protein Kinase

BMS309403 is a biphenyl azole inhibitor against fatty acid binding protein 4 (FABP4) and regarded as a lead compound for effective treatment of obesity related cardio-metabolic diseases. Here we discovered an off-target activity of BMS309403 in that it stimulates glucose uptake in C2C12 myotubes in a temporal and dose dependent manner via activation of AMP-activated protein kinase (AMPK) signaling pathway but independent of FABPs. Further analysis indicated that BMS309403 activates AMPK through increasing the ratio of intracellular AMP:ATP while decreasing mitochondrial membrane potential. These findings provide mechanistic insights on the action of BMS309403.     

Pre-emptive pharmacological inhibition of fatty acid–binding protein 4 attenuates kidney fibrosis by reprogramming tubular lipid metabolism

Kidney fibrosis is a hallmark of chronic kidney disease (CKD) progression that is caused by tubular injury and dysregulated lipid metabolism. Genetic abolition fatty acid-binding protein 4 (FABP4), a key lipid transporter, has been reported to suppress kidney interstitial fibrosis. However, the role and underlying mechanism of chemical inhibition of FABP4 in fibrotic kidney have not been well-documented. Here, we examined preemptive the effect of a FABP4 inhibitor, BMS309403, on lipid metabolism of tubular epithelial cells (TECs) and progression of kidney fibrosis. The expression of FABP4 was significantly elevated, concomitated with the accumulation of lipid droplets in TECs during kidney fibrosis. Treatment with BMS309403 alleviated lipid deposition of TECs, as well as interstitial fibrotic responses both in unilateral ureteral obstruction (UUO)-engaged mice and TGF-β-induced TECs. Moreover, BMS309403 administration enhanced fatty acid oxidation (FAO) in TECs by regulating peroxisome proliferator-activated receptor γ (PPARγ) and restoring FAO-related enzyme activities; In addition, BMS309403 markedly reduced cell lipotoxicity, such as endoplasmic reticulum (ER) stress and apoptosis in fibrotic kidney. Taken together, our results suggest that preemptive pharmacological inhibition of FABP4 by BMS309403 rebalances abnormal lipid metabolism in TECs and attenuates the progression of kidney fibrosis, thus may hold therapeutic potential for the treatment of fibrotic kidney diseases.Subject terms: Metabolism, Chronic kidney disease     

The effects of 17 alpha-estradiol to inhibit inflammation in vitro

Background

17 Alpha-estradiol (17 α-E2) is a natural, non-feminizing stereoisomer of 17 beta-estradiol (17 β-E2). Whereas much is known about the physiological effects of 17 β-E2, much less is known about 17 α-E2. For example, 17 β-E2 exerts anti-inflammatory effects in neurons and adipocytes through binding and activation of estrogen receptor alpha (ERα); however, if 17 α-E2 has similar effects on inflammation is currently unknown.

Methods

To begin to address this, we analyzed the ability of 17 α-E2 and 17 β-E2 to suppress lipopolysaccharide (LPS)-induced inflammation in vitro using embryonic fibroblast cells (MEF) from wild type and total body ERα (ERKO) male and female mice. Additionally, we further probed if there were sex differences with respect to the effects of E2s using primary pre-adipocyte cells from C57BL/6J male and female mice. Also, we probed mechanistically the effects of E2s in fully differentiated 3T3-L1 cells.

Results

Both E2s decreased LPS-induced markers of inflammation Tnf-α and Il-6, and increased the anti-inflammatory markers Il-4 and IL-6 receptor (Il-6ra) in MEF cells. To begin to understand the mechanisms by which both E2’s mediate their anti-inflammatory effects, we probed the role of ERα using two methods. First, we used MEF cells from ERKO mice and found reductions in ERα diminished the ability of 17 α-E2 to suppress Tnf-α in female but not in male cells, demonstrating a sexual dimorphism in regard to the role of ERα to mediate 17 α-E2’s effects. Second, we selectively reduced the expression of ERα in 3T3-L1 cells using siRNA and found reductions in ERα diminished the ability of both E2s to suppress Tnf-α and Il-6 expression. Lastly, to determine the mechanisms by which E2s reduce inflammation, we explored the role of NFκB-p65 and found both E2s decreased NFκB-p65 expression.

Conclusions

In conclusion, we demonstrate for the first time that 17 α-E2, as well as 17 β-E2, suppresses inflammation through their effects on ERα and NFκB-p65.

     

Effect of everolimus on the glucose metabolic pathway in mouse skeletal muscle cells (C2C12)

Introduction

Everolimus selectively inhibits mammalian target of rapamycin complex 1 (mTORC1) and exerts an antineoplastic effect. Metabolic disturbance has emerged as a common and unique side effect of everolimus.

Objectives

We used targeted metabolomic analysis to investigate the effects of everolimus on the intracellular glycometabolic pathway.

Methods

Mouse skeletal muscle cells (C2C12) were exposed to everolimus for 48 h, and changes in intracellular metabolites were determined by capillary electrophoresis time-of-flight mass spectrometry. mRNA abundance, protein expression and activity were measured for enzymes involved in glycometabolism and related pathways.

Results

Both extracellular and intracellular glucose levels increased with exposure to everolimus. Most intracellular glycometabolites were decreased by everolimus, including those involved in glycolysis and the pentose phosphate pathway, whereas no changes were observed in the tricarboxylic acid cycle. Everolimus suppressed mRNA expression of enzymes related to glycolysis, downstream of mTOR signaling enzymes and adenosine 5′-monophosphate protein kinases. The activity of key enzymes involved in glycolysis and the pentose phosphate pathway were decreased by everolimus. These results show that everolimus impairs glucose utilization in intracellular metabolism.

Conclusions

The present metabolomic analysis indicates that everolimus impairs glucose metabolism in muscle cells by lowering the activities of glycolysis and the pentose phosphate pathway.

     

Fructose ingestion impairs expression of genes involved in skeletal muscle’s adaptive response to aerobic exercise

Background

The inverse relationship between exercise capacity and its variation over time and both cardiovascular and all-cause mortality suggests the existence of an etiological nexus between cardiometabolic diseases and the molecular regulators of exercise capacity. Coordinated adaptive responses elicited by physical training enhance exercise performance and metabolic efficiency and possibly mediate the health benefits of physical exercise. In contrast, impaired expression of genes involved in mitochondrial biogenesis or protein turnover in skeletal muscle—key biological processes involved in adaptation to physical training—leads to insulin resistance and obesity. Ingestion of fructose has been shown to suppress the exercise-induced GLUT4 response in rat skeletal muscle. To evaluate in greater detail how fructose ingestion might blunt the benefits of physical training, we investigated the effects of fructose ingestion on exercise induction of genes that participate in regulation of mitochondrial biogenesis and protein turnover in rat’s skeletal muscle.

Methods

Eight-week-old Wistar rats were randomly assigned to sedentary (C), exercise (treadmill running)-only (E), fructose-only (F), and fructose + exercise (FE) groups and treated accordingly for 8 weeks. Blood and quadriceps femoris were collected for biochemistry, serum insulin, and gene expression analysis. Expression of genes involved in regulation of mitochondrial biogenesis and autophagy, GLUT4, and ubiquitin E3 ligases MuRF-1, and MAFbx/Atrogin-1 were assayed with quantitative real-time polymerase chain reaction.

Results

Aerobic training improved exercise capacity in both E and FE groups. A main effect of fructose ingestion on body weight and fasting serum triglyceride concentration was detected. Fructose ingestion impaired the expression of PGC-1α, FNDC5, NR4A3, GLUT4, Atg9, Lamp2, Ctsl, Murf-1, and MAFBx/Atrogin-1 in skeletal muscle of both sedentary and exercised animals while expression of Errα and Pparδ was impaired only in exercised rats.

Conclusions

Our results show that fructose ingestion impairs the expression of genes involved in biological processes relevant to exercise-induced remodeling of skeletal muscle. This might provide novel insight on how a dietary factor contributes to the genesis of disorders of glucose metabolism.

     

Nitrate enhances skeletal muscle fatty acid oxidation via a nitric oxide-cGMP-PPAR-mediated mechanism

Background

Insulin sensitivity in skeletal muscle is associated with metabolic flexibility, including a high capacity to increase fatty acid (FA) oxidation in response to increased lipid supply. Lipid overload, however, can result in incomplete FA oxidation and accumulation of potentially harmful intermediates where mitochondrial tricarboxylic acid cycle capacity cannot keep pace with rates of β-oxidation. Enhancement of muscle FA oxidation in combination with mitochondrial biogenesis is therefore emerging as a strategy to treat metabolic disease. Dietary inorganic nitrate was recently shown to reverse aspects of the metabolic syndrome in rodents by as yet incompletely defined mechanisms.

Results

Herein, we report that nitrate enhances skeletal muscle FA oxidation in rodents in a dose-dependent manner. We show that nitrate induces FA oxidation through a soluble guanylate cyclase (sGC)/cGMP-mediated PPARβ/δ- and PPARα-dependent mechanism. Enhanced PPARβ/δ and PPARα expression and DNA binding induces expression of FA oxidation enzymes, increasing muscle carnitine and lowering tissue malonyl-CoA concentrations, thereby supporting intra-mitochondrial pathways of FA oxidation and enhancing mitochondrial respiration. At higher doses, nitrate induces mitochondrial biogenesis, further increasing FA oxidation and lowering long-chain FA concentrations. Meanwhile, nitrate did not affect mitochondrial FA oxidation in PPARα^?/? mice. In C2C12 myotubes, nitrate increased expression of the PPARα targets Cpt1b, Acadl, Hadh and Ucp3, and enhanced oxidative phosphorylation rates with palmitoyl-carnitine; however, these changes in gene expression and respiration were prevented by inhibition of either sGC or protein kinase G. Elevation of cGMP, via the inhibition of phosphodiesterase 5 by sildenafil, also increased expression of Cpt1b, Acadl and Ucp3, as well as CPT1B protein levels, and further enhanced the effect of nitrate supplementation.

Conclusions

Nitrate may therefore be effective in the treatment of metabolic disease by inducing FA oxidation in muscle.

     

Selenium unmasks protective iron armor: A possible defense against cutaneous inflammation and cancer

Background

A link between selenium deficiency and inflammatory skin diseases have been noted by many, but this link is still not well understood. We have previously studied the efficacy of ceramide analogs, based on the fire ant venom Solenopsin A, against our psoriasis animal model. Treatment of animals with solenopsin analogs resulted in significantly improved skin as well as in a coordinate downregulation of selenoproteins, namely Glutathione Peroxidase 4 (GPX4). We thus hypothesize that ferroptosis may be a physiologic process that may protect the skin from both inflammatory and neoplastic processes.

INFLAM – INFLAMmation in Brain and Vessels with Iron Nanoparticles and Cell Trafficking: A Multiscale Approach of Tissue Microenvironment,Iron Nanostructure and Iron Biotransformation

Background

Inflammation is a share process in atherosclerosis and stroke and is thought to be a key player in the evolution of these diseases. Ten years ago, inflammation imaging with magnetic resonance imaging (MRI) was considered very promising for both pre-clinical and clinical studies of atherosclerosis and stroke.

Plasma N-glycome composition associates with chronic low back pain

Background

Low back pain (LBP) is the symptom of a group of syndromes with heterogeneous underlying mechanisms and molecular pathologies, making treatment selection and patient prognosis very challenging. Moreover, symptoms and prognosis of LBP are influenced by age, gender, occupation, habits, and psychological factors. LBP may be characterized by an underlying inflammatory process. Previous studies indicated a connection between inflammatory response and total plasma N-glycosylation. We wanted to identify potential changes in total plasma N-glycosylation pattern connected with chronic low back pain (CLBP), which could give an insight into the pathogenic mechanisms of the disease.