Cell-based multiwell assays for the detection of substrate accumulation and oxidation

Wensaas, Andreas J.; Rustan, Arild C.; Lövstedt, K.; Kull, Bengt; Wikström, Sverre; Drevon, C. A.; Hallén, Stefan

doi:10.1194/jlr.d600047-jlr200

Cited by 86 publications

(94 citation statements)

References 17 publications

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“…GW501516 also increased mRNA levels for carnitine palmitoyltransferase-1 (CPT-1). Additionally, a dose-dependent increase in oleate oxidation to CO2 was observed in myotubes pretreated for three days with this compound during differentiation of the cells (Wensaas, et al, 2007). The PPARδ agonist was also shown to increase basal and insulin-mediated glucose transport and lead to phosphorylation of AMPK and p38 mitogen-activated protein kinase (MAPK) in cultured human myotubes (Kramer, et al, 2007).…”

Section: Strategies To Improve the Human Myotube Modelmentioning

confidence: 91%

Are cultured human myotubes far from home?

et al. 2013

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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.

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Section: Strategies To Improve the Human Myotube Modelmentioning

confidence: 91%

Are cultured human myotubes far from home?

et al. 2013

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“…Glycogen synthesis Glycogen synthesis was assayed as described previously [40]. Myotubes were glucose/serumstarved for 1.5 h, incubated for 3 h in DMEM containing 5.5 mmol/l glucose and 18.5 kBq/ml 2-deoxy[ 3 H]glucose with or without 100 nmol/l insulin, and then washed and collected.…”

Section: Exercise Intervention and [ 18 F]fluorodeoxyglucose Petmentioning

confidence: 99%

Naturally occurring R225W mutation of the gene encoding AMP-activated protein kinase (AMPK)γ3 results in increased oxidative capacity and glucose uptake in human primary myotubes

et al. 2010

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Aims/hypothesis AMP-activated protein kinase (AMPK) has a broad role in the regulation of glucose and lipid metabolism making it a promising target in the treatment of type 2 diabetes mellitus. We therefore sought to characterise for the first time the effects of chronic AMPK activation on skeletal muscle carbohydrate metabolism in carriers of the rare gain-of-function mutation of the gene encoding AMPKγ 3 subunit, PRKAG3 R225W.Methods Aspects of fuel metabolism were studied in vitro in myocytes isolated from vastus lateralis of PRKAG3 R225W carriers and matched control participants. In vivo, muscular strength and fatigue were evaluated by isokinetic dynamometer and surface electromyography, respectively. Glucose uptake in exercising quadriceps was determined using [ 18 F]fluorodeoxyglucose and positron emission tomography. Results Myotubes from PRKAG3 R225W carriers had threefold higher mitochondrial content (p < 0.01) and oxidative capacity, higher leak-dependent respiration (1.6-fold, p<0.05), higher basal glucose uptake (twofold, p<0.01) and higher glycogen synthesis rates (twofold, p<0.05) than control myotubes. They also had higher levels of intracellular glycogen (p<0.01) and a trend for lower intramuscular triacylglycerol stores. R225W carriers showed remarkable resistance to muscular fatigue and a trend for increased glucose uptake in exercising muscle in vivo. Conclusions/interpretation Through the enhancement of skeletal muscle glucose uptake and increased mitochondrial content, the R225W mutation may significantly enhance exercise performance. These findings are also consistent with the hypothesis that the γ 3 subunit of AMPK is a promising tissue-specific target for the treatment of type 2 diabetes mellitus, a condition in which glucose uptake and mitochondrial function are impaired.

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“…Radioactive 14 CO 2 was trapped in a filter plate for 4 h before myotubes were placed on ice, washed twice with PBS (150 μl/well) and 200 μl 0.05 mol/l NaOH added. Lysed cell fractions (50 μl) and filter plates were quantified by liquid scintillation (Wallac MicroBeta Trilux, PerkinElmer, Oslo, Norway), as previously described [37]. Cells were later assayed for protein.…”

Section: Methodsmentioning

confidence: 99%

Liver X receptor antagonist reduces lipid formation and increases glucose metabolism in myotubes from lean, obese and type 2 diabetic individuals

et al. 2007

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Aims/hypothesis Liver X receptors (LXRs) play important roles in lipid and carbohydrate metabolism. The purpose of the present study was to evaluate effects of the endogenous LXR agonist 22-R-hydroxycholesterol (22-R-HC) and its stereoisomer 22-S-hydroxycholesterol (22-S-HC), in comparison with the synthetic agonist T0901317 on lipid and glucose metabolism in human skeletal muscle cells (myotubes). Methods Myotubes established from lean and obese control volunteers and from obese type 2 diabetic volunteers were treated with LXR ligands for 4 days. Lipid and glucose metabolisms were studied with labelled precursors, and gene expression was analysed using real-time PCR.

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Cell-based multiwell assays for the detection of substrate accumulation and oxidation

Abstract: We describe multiwell assays for detecting the accumulation as well as the subsequent oxidation of 14

Cited by 86 publications

References 17 publications

Are cultured human myotubes far from home?

Are cultured human myotubes far from home?

Naturally occurring R225W mutation of the gene encoding AMP-activated protein kinase (AMPK)γ3 results in increased oxidative capacity and glucose uptake in human primary myotubes

Liver X receptor antagonist reduces lipid formation and increases glucose metabolism in myotubes from lean, obese and type 2 diabetic individuals

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