Cultured slow vs. fast skeletal muscle cells differ in physiology and responsiveness to stimulation

University of Dundee, Dundee, Scotland, United Kingdom
AJP Cell Physiology (Impact Factor: 3.78). 08/2006; 291(1):C11-7. DOI: 10.1152/ajpcell.00366.2005
Source: PubMed


In vitro studies have used protein markers to distinguish between myogenic cells isolated from fast and slow skeletal muscles. The protein markers provide some support for the hypothesis that satellite cells from fast and slow muscles are different, but the data are equivocal. To test this hypothesis directly, three-dimensional skeletal muscle constructs were engineered from myogenic cells isolated from fast tibialis anterior (TA) and slow soleus (SOL) muscles of rats and functionality was tested. Time to peak twitch tension (TPT) and half relaxation time (RT(1/2)) were approximately 30% slower in constructs from the SOL. The slower contraction and relaxation times for the SOL constructs resulted in left shift of the force-frequency curve compared with those from the TA. Western blot analysis showed a 60% greater quantity of fast myosin heavy chain in the TA constructs. 14 days of chronic low-frequency electrical stimulation resulted in a 15% slower TPT and a 14% slower RT(1/2), but no change in absolute force production in the TA constructs. In SOL constructs, slow electrical stimulation resulted in an 80% increase in absolute force production with no change in TPT or RT(1/2). The addition of cyclosporine A did not prevent the increase in force in SOL constructs after chronic low-frequency electrical stimulation, suggesting that calcineurin is not responsible for the increase in force. We conclude that myogenic cells associated with a slow muscle are imprinted to produce muscle that contracts and relaxes slowly and that calcineurin activity cannot explain the response to a slow pattern of electrical stimulation.

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Available from: Keith Baar, Jul 08, 2014
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    • "We utilized a 24hr pre-conditioning stimulus of a continuous 10Hz train of 0.4sec followed by a 3.6 second rest that we have used previously (Donnelly et al., 2010; Khodabukus and Baar, 2012), but found that this is not required for the cells to adapt to the adult-like stimulus (data not shown). Myoblasts of different origins have different ranges of plasticity in response to electrical stimulation (Huang et al., 2006; Wehrle et al., 1994). C2C12s originate from mouse thigh muscle which is a mixed phenotype muscle (Blau et al., 1985). "
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    ABSTRACT: The role of factors such as frequency, contraction duration and active time in the adaptation to chronic low-frequency electrical stimulation (CLFS) is widely disputed. In this study we explore the ability of contraction duration (0.6, 6, 60 and 600 secs) to induce a fast-to-slow shift in engineered muscle while using a stimulation frequency of 10Hz and keeping active time constant at 60%. We found that all contraction durations induced similar slowing of time-to-peak tension. Despite similar increases in total myosin heavy (MHC) levels with stimulation, increasing contraction duration resulted in progressive decreases in total fast myosin. With contraction durations of 60 and 600 secs, MHC IIx levels decreased and MHC IIa levels increased. All contraction durations resulted in fast-to-slow shifts in TnT and TnC but increased both fast and slow TnI levels. Half-relaxation slowed to a greater extent with contraction durations of 60 and 600 secs despite similar changes in the calcium sequestering proteins calsequestrin and parvalbumin and the calcium uptake protein SERCA. All CLFS groups resulted in greater fatigue resistance than control. Similar increases in GLUT4, mitochondrial enzymes (SDH and ATPsynthase), the fatty acid transporter CPT-1 and the metabolic regulators PGC-1α and MEF2 were found with all contraction durations. However, the mitochondrial enzymes cytochrome C and citrate synthase were increased to greater levels with contraction durations of 60 and 600 seconds. These results demonstrate that contraction duration plays a pivotal role in dictating the level of CLFS-induced contractile and metabolic adaptations in tissue engineered skeletal muscle. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    Journal of Cellular Physiology 04/2015; 230(10). DOI:10.1002/jcp.24985 · 3.84 Impact Factor
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    • "Muscles engineered from fast and slow rat skeletal muscle retain in vitro the contractile dynamics (i.e. TPT and 1/2RT) of the muscle from which they were derived (Huang, Dennis et al. 2006). However, rat muscles also display more distinct fiber types than mouse (Schiaffino 2010), as best exemplified in the rat soleus muscle (96% Type I, 4% Type IIa) (Bloemberg and Quadrilatero 2012) compared to the mouse soleus (31% Type I, 51% IIa, 15% IIx and 3% IIb) (Bloemberg and Quadrilatero 2012). "
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    ABSTRACT: Satellite cells derived from fast and slow muscles have been shown to adopt contractile and metabolic properties of their parent muscle. Mouse muscle shows less distinctive fiber-type profiles than rat or rabbit muscle. Therefore, in this study we sought to determine whether three-dimensional muscle constructs engineered from slow soleus (SOL) and fast tibialis anterior (TA) from mice would adopt the contractile and metabolic properties of their parent muscle. Time-to-peak tension (TPT) and half-relaxation time (1/2RT) was significantly slower in SOL constructs. In agreement with TPT, TA constructs contained significantly higher levels of fast myosin heavy chain (MHC) and fast troponin C, I, and T isoforms. Fast SERCA protein, both slow and fast calsequestrin isoforms and parvalbumin were found at higher levels in TA constructs. SOL constructs were more fatigue resistant and contained higher levels of the mitochondrial proteins SDH and ATP synthase and the fatty acid transporter CPT-1. SOL constructs contained lower levels of the glycolytic enzyme phosphofructokinase but higher levels of the β-oxidation enzymes LCAD and VLCAD suggesting greater fat oxidation. Despite no changes in PGC-1α protein, SOL constructs contained higher levels of SIRT1 and PRC. TA constructs contained higher levels of the slow-fiber program repressor SOX6 and the six transcriptional complex (STC) proteins Eya1and Six4 which may underlie the higher in fast-fiber and lower slow-fiber program proteins. Overall, we have found that muscles engineered from predominantly slow and fast mouse muscle retain contractile and metabolic properties of their native muscle. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
    Journal of Cellular Physiology 10/2014; 230(8). DOI:10.1002/jcp.24848 · 3.84 Impact Factor
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    • "(Pette and Staron, 1990; Schiaffino and Reggiani, 1996). Skeletal muscle phenotype is dependent on developmental state (Condon et al., 1990; Hallauer and Hastings, 2002; Kelly and Rubinstein, 1980) and cellular origin (DiMario and Stockdale, 1997; Huang et al., 2006; Kalhovde et al., 2005) but is primarily regulated by neural input in adult muscle (Eken and Gundersen, 1988; Salmons and Sreter, 1976; Wehrle et al., 1994). "
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    ABSTRACT: Cell culture conditions can vary between laboratories and have been optimised for 2D cell culture. In this study, engineered muscle was cultured in 5.5 mM low glucose (LG) or 25 mM high glucose (HG) and in the absence or presence (+S) of streptomycin and the effect on C2C12 tissue-engineered muscle function and metabolism was determined. Following 2 weeks differentiation, streptomycin (3-fold) and LG (0.5-fold) significantly decreased force generation. LG and/or streptomycin resulted in upward and leftward shifts in the force-frequency curve and slowed time-to-peak tension and half-relaxation time. Despite changes in contractile dynamics, no change in myosin isoform was detected. Instead, changes in troponin isoform, calcium sequestering proteins (CSQ and parvalbumin) and the calcium uptake protein SERCA predicted the changes in contractile dynamics. Culturing in LG and/or streptomycin resulted in increased fatigue resistance despite no change in the mitochondrial enzymes SDH, ATPsynthase and cytochrome C. However, LG resulted in increases in the β-oxidation enzymes LCAD and VLCAD and the fatty acid transporter CPT-1, indicative of a greater capacity for fat oxidation. In contrast, HG resulted in increased GLUT4 content and the glycolytic enzyme PFK, indicative of a more glycolytic phenotype. These data suggest that streptomycin has negative effects on force generation and that glucose can be used to shift engineered muscle phenotype via changes in calcium-handling and metabolic proteins. J. Cell. Physiol. © 2014 Wiley Periodicals, Inc.
    Journal of Cellular Physiology 10/2014; 230(6). DOI:10.1002/jcp.24857 · 3.84 Impact Factor
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