Background and AimsPhysical exercise leads to substantial adaptive responses in skeletal muscles and plays a central role in a healthy life style. Since exercise induces major systemic responses, underlying cellular mechanisms are difficult to study in vivo. It was therefore desirable to develop an in vitro model that would resemble training in cultured human myotubes.MethodsElectrical pulse stimulation (EPS) was applied to adherent human myotubes. Cellular contents of ATP, phosphocreatine (PCr) and lactate were determined. Glucose and oleic acid metabolism were studied using radio-labeled substrates, and gene expression was analyzed using real-time RT-PCR. Mitochondrial content and function were measured by live imaging and determination of citrate synthase activity, respectively. Protein expression was assessed by electrophoresis and immunoblotting.ResultsHigh-frequency, acute EPS increased deoxyglucose uptake and lactate production, while cell contents of both ATP and PCr decreased. Chronic, low-frequency EPS increased oxidative capacity of cultured myotubes by increasing glucose metabolism (uptake and oxidation) and complete fatty acid oxidation. mRNA expression level of pyruvate dehydrogenase complex 4 (PDK4) was significantly increased in EPS-treated cells, while mRNA expressions of interleukin 6 (IL-6), cytochrome C and carnitin palmitoyl transferase b (CPT1b) also tended to increase. Intensity of MitoTracker®Red FM was doubled after 48 h of chronic, low-frequency EPS. Protein expression of a slow fiber type marker (MHCI) was increased in EPS-treated cells.ConclusionsOur results imply that in vitro EPS (acute, high-frequent as well as chronic, low-frequent) of human myotubes may be used to study effects of exercise.

The beneficial health-related effects of exercise are well recognized, and numerous studies have investigated underlying mechanism using various in vivo and in vitro models. Although electrical pulse stimulation (EPS) for the induction of muscle contraction has been used for quite some time, its application on cultured skeletal muscle cells of animal or human origin as a model of in vitro exercise is a more recent development. In this review, we compare in vivo exercise and in vitro EPS with regard to effects on signalling, expression level and metabolism. We provide a comprehensive overview of different EPS protocols and their applications, discuss technical aspects of this model including critical controls and the importance of a proper maintenance procedure and finally discuss the limitations of the EPS model.

IntroductionSatellite cells can be isolated from skeletal muscle biopsies, activated to proliferating myoblast and differentiated into multinuclear myotubes in culture. These cell cultures represent an essential model system to intact human skeletal muscle, which can be modulated ex vivo. Advantages of this system include; having the most relevant genetic background to study human disease (as opposed to rodent cell cultures), the extracellular environment can be precisely controlled and the cells are not immortalized, thereby offering the possibility of studying innate characteristics of the donor. This review will focus on how human myotubes can be used as a tool to study metabolism in skeletal muscles, with a special attention to changes in muscle energy metabolism in obesity and type 2 diabetes.Limitations of this cell system and possible approaches to improve the current model will also be discussed.

This study was conducted to evaluate the chronic effects of eicosapentaenoic acid (EPA) on fatty acid and glucose metabolism in human skeletal muscle cells. Uptake of [ 14 C]oleate was increased .2-fold after preincubation of myotubes with 0.6 mM EPA for 24 h, and incorporation into various lipid classes showed that cellular triacylgycerol (TAG) and phospholipids were increased 2-to 3-fold compared with control cells. After exposure to oleic acid (OA), TAG was increased 2-fold. Insulin (100 nM) further increased the incorporation of [ 14 C]oleate into all lipid classes for EPA-treated myotubes. Fatty acid b-oxidation was unchanged, and complete oxidation (CO 2 ) decreased in EPA-treated cells. Basal glucose transport and oxidation (CO 2 ) were increased 2-fold after EPA, and insulin (100 nM) stimulated glucose transport and oxidation similarly in control and EPA-treated myotubes, whereas these responses to insulin were abolished after OA treatment. Lower concentrations of EPA (0.1 mM) also increased fatty acid and glucose uptake. CD36/FAT (fatty acid transporter) mRNA expression was increased after EPA and OA treatment compared with control cells. Moreover, GLUT1 expression was increased 2.5-fold by EPA, whereas GLUT4 expression was unchanged, and activities of the mitogen-activated protein kinase p38 and extracellular signal-regulated kinase were decreased after treatment with OA compared with EPA. Together, our data show that chronic exposure of myotubes to EPA promotes increased uptake and oxidation of glucose despite a markedly increased fatty acid uptake and synthesis of complex lipids.-Aas, V

Liver X receptors (LXRs) are important regulators of cholesterol and lipid metabolism and are also involved in glucose metabolism. However, the functional role of LXRs in human skeletal muscle is at present unknown. This study demonstrates that chronic ligand activation of LXRs by a synthetic LXR agonist increases the uptake, distribution into complex cellular lipids, and oxidation of palmitate as well as the uptake and oxidation of glucose in cultured human skeletal muscle cells. Furthermore, the effect of the LXR agonist was additive to acute effects of insulin on palmitate uptake and metabolism. Consistently, activation of LXRs induced the expression of relevant genes: fatty acid translocase (CD36/FAT), glucose transporters (GLUT1 and -4), sterol regulatory element-binding protein-1c, peroxisome proliferator-activated receptor-␥, carnitine palmitoyltransferase-1, and uncoupling protein 2 and 3. Interestingly, in response to activation of LXRs, myotubes from patients with type 2 diabetes showed an elevated uptake and incorporation of palmitate into complex lipids but an absence of palmitate oxidation to CO 2 . These results provide evidence for a functional role of LXRs in both lipid and glucose metabolism and energy uncoupling in human myotubes. Furthermore, these data suggest that increased intramyocellular lipid content in type 2 diabetic patients may involve an altered response to activation of components in the LXR pathway.

PPARα is one of three members of the soluble nuclear receptor family called peroxisome proliferator-activated receptor (PPAR). It is a sensor for changes in levels of fatty acids and their derivatives that responds to ligand binding with PPAR target gene transcription, inasmuch as it can influence physiological homeostasis, including lipid and carbohydrate metabolism in various tissues. In this paper we summarize the involvement of PPARα in the metabolically active tissues liver and skeletal muscle and provide an overview of the risks and benefits of ligand activation of PPARα, with particular consideration to interspecies differences.

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