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.
Feng YZ, Nikolić N, Bakke SS, Kase ET, Guderud K, Hjelmesaeth J, Aas V, Rustan AC, Thoresen GH. Myotubes from lean and severely obese subjects with and without type 2 diabetes respond differently to an in vitro model of exercise.
Cultured human myotubes have a low mitochondrial oxidative potential. This study aims to remodel energy metabolism in myotubes by replacing glucose with galactose during growth and differentiation to ultimately examine the consequences for fatty acid and glucose metabolism. Exposure to galactose showed an increased [14C]oleic acid oxidation, whereas cellular uptake of oleic acid uptake was unchanged. On the other hand, both cellular uptake and oxidation of [14C]glucose increased in myotubes exposed to galactose. In the presence of the mitochondrial uncoupler carbonylcyanide p-trifluormethoxy-phenylhydrazone (FCCP) the reserve capacity for glucose oxidation was increased in cells grown with galactose. Staining and live imaging of the cells showed that myotubes exposed to galactose had a significant increase in mitochondrial and neutral lipid content. Suppressibility of fatty acid oxidation by acute addition of glucose was increased compared to cells grown in presence of glucose. In summary, we show that cells grown in galactose were more oxidative, had increased oxidative capacity and higher mitochondrial content, and showed an increased glucose handling. Interestingly, cells exposed to galactose showed an increased suppressibility of fatty acid metabolism. Thus, galactose improved glucose metabolism and metabolic switching of myotubes, representing a cell model that may be valuable for metabolic studies related to insulin resistance and disorders involving mitochondrial impairments.
We investigated the effects of PGC-1α (peroxisome proliferator-activated receptor γ coactivator-1α) overexpression on the oxidative capacity of human skeletal muscle cells ex vivo. PGC-1α overexpression increased the oxidation rate of palmitic acid and mRNA expression of genes regulating lipid metabolism, mitochondrial biogenesis, and function in human myotubes. Basal and insulin-stimulated deoxyglucose uptake were decreased, possibly due to upregulation of PDK4 mRNA. Expression of fast fiber-type gene marker (MHCIIa) was decreased. Compared to skeletal muscle in vivo, PGC-1α overexpression increased expression of several genes, which were downregulated during the process of cell isolation and culturing. In conclusion, PGC-1α overexpression increased oxidative capacity of cultured myotubes by improving lipid metabolism, increasing expression of genes involved in regulation of mitochondrial function and biogenesis, and decreasing expression of MHCIIa. These results suggest that therapies aimed at increasing PGC-1α expression may have utility in treatment of obesity and obesity-related diseases.
(2)Fast growing woody plants represent effective tools for cadmium (Cd) extraction during remediation of low to medium Cd contaminated soils. Poplars are good candidates for this task because of their rapid growth rate, high biomass yield, and adaptability, as well as the availability of well-characterized clones/ genotypes with various anatomical and physiological traits. The present study evaluates the potential of Populus deltoides (clone B-81) and Populus × euramericana (clone Pannonia) for phytoremediation of Cd contamination in soil. Poplar clones were analyzed for (1) plant growth response to Cd contamination, (2) Cd accumulation, translocation, and partitioning between plant organs, and (3) morphological, anatomical and physiological responses to Cd stress as a function of biomass production. Plants were cultivated in soil moderately contaminated with Cd (8.14 mg kg -1 soil) under semi-controlled conditions for six weeks. Our results suggest that P. × euramericana and P. deltoides clones respond differently to Cd contamination. Biomass production and morphological characteristics were more negatively affected in P. × euramericana than in P. deltoides plants. However, most examined leaf structural parameters were not significantly affected by Cd. In most cases, photosynthetic characteristics and gas exchange parameters were affected by Cd treatment, but the levels and patterns of changes depended on the clone. High tolerance to applied Cd levels, as estimated by the tolerance index, was observed in both clones, but was higher in P. deltoides than P. × euramericana (82.2 vs. 66.5, respectively). We suspect that the higher tolerance to Cd toxicity observed in P. deltoides could be related to unchanged proline content and undisturbed nitrogen metabolism. Following treatment, 58.0 and 46.7% of the total Cd content was accumulated in the roots of P. × euramericana and P. deltoides, respectively, with the remainder in the stems (18.2 and 39.9%) and leaves (23.8 and 13.4%). In summary, P. deltoides displayed better phytoextraction performance under Cd exposure than P. × euramericana, suggesting its potential not only for Cd phytostabilization, but also phytoextraction projects.
Obesity and weight gain are serious concerns after solid organ transplantation (Tx); however, no unbiased comparison regarding body weight parameter evolution across organ groups has yet been performed. Using data from the prospective nationwide Swiss Transplant Cohort Study, we compared the evolution of weight parameters up to 3 years post-Tx in 1359 adult kidney (58.3%), liver (21.7%), lung (11.6%), and heart (8.4%) recipients transplanted between May 2008 and May 2012. Changes in mean weight and body mass index (BMI) category were compared to reference values from 6 months post-Tx. At 3 years post-Tx, compared to other organ groups, liver Tx recipients showed the greatest weight gain (mean 4.8±10.4 kg), 57.4% gained >5% body weight, and they had the highest incidence of obesity (38.1%). After 3 years, based on their BMI categories at 6 months, normal weight and obese liver Tx patients, as well as underweight kidney, lung and heart Tx patients had the highest weight gains. Judged against international Tx patient data, the majority of our Swiss Tx recipients' experienced lower post-Tx weight gain. However, our findings show weight gain pattern differences, both within and across organ Tx groups that call for preventive measures.
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