C2 and C2C12 murine skeletal myoblast models of atrophic and hypertrophic potential: relevance to disease and ageing?
Reduced muscle mass and increased susceptibility to TNF-induced degradation accompany inflamed ageing and chronic diseases. Furthermore, C(2) myoblasts display diminished differentiation and increased susceptibility to TNF-alpha-induced cell death versus subcloned C(2)C(12) cells, providing relevant models to assess: differentiation (creatine kinase), growth (protein), death (trypan-blue) and anabolic/catabolic parameters (RT-PCR) over 72 h +/- TNF-alpha (20 ng ml(-1)). At 48 and 72 h, respectively, larger myotubes and significantly higher CK activity (320.26 +/- 6.82 vs. 30.71 +/- 2.5, P < 0.05; 544.94 +/- 27.7 vs. 39.4 +/- 3.37 mU mg ml(-1), P < 0.05), fold increases in myoD (21.45 +/- 3.12 vs. 3.97 +/- 1.76, P < 0.05; 31.07 +/- 3.1 vs. 6.82 +/- 1.93, P < 0.05) and myogenin mRNA (241.8 +/- 40 vs. 36.80 +/- 19.3, P < 0.05; 440 +/- 100.5 vs. 201.1 +/- 86, P < 0.05) were detected in C(2)C(12) versus C(2). C(2)C(12) showed significant increases in IGF-I mRNA (243.05 +/- 3.87 vs. 105.75 +/- 21.95, P < 0.05), reduced proliferation and significantly lower protein expression (1.21 +/- 0.28 vs. 1.79 +/- 0.29 mg ml(-1), P < 0.05) at 72 h versus C(2) cells. Significant temporal reductions in C(2)C(12) IGFBP2 mRNA (28.02 +/- 15.44, 13.82 +/- 8.07, 6.92 +/- 4.37, P < 0.05) contrasted increases in C(2)s (4.31 +/- 3.31, 13.02 +/- 9.92, 82.9 +/- 58.9, P < 0.05) at 0, 48 and 72 h, respectively. TNF-alpha increased cell death in C(2)s (2.67 +/- 1.54%, 34.42 +/- 5.39%, 29.71 +/- 5.79% (0, 48, 72 h), P < 0.05), yet was without effect in C(2)C(12)s at 48 h but caused a small significant increase at 72 h (9.88 +/- 4.02% (TNF-alpha) vs. 6.17 +/- 0.749% (DM), 72 h). TNF-alpha and TNFRI mRNA were unchanged; however, larger reductions in IGF-I (8.2- and 7.5-fold vs. 4.5- and 4.1-fold (48, 72 h)), IGF-IR (2-fold vs. no-significant reduction (72 h)) and IGFBP5 (3.24 vs. 1.38 (48 h) and 2.21 vs. 1.71 (72 h), P < 0.05) mRNA were observed in C(2) versus C(2)C(12) with TNF-alpha. This investigation provides insight into regulators of altered basal hypertrophy and TNF-induced atrophy, providing a model for future investigation into therapeutic initiatives for ageing/wasting disorders.
Sharples
Adam P
AP
Faculty of Science and Engineering, Institute for Biomedical Research into Human Movement and Health, Manchester Metropolitan University, Manchester, UK. a.p.sharples@mmu.ac.uk
Al-Shanti
Nasser
N
Stewart
Claire E
CE
D000375:Aging / Q000502:physiology*; D000818:Animals; D001284:Atrophy / Q000378:metabolism* / Q000473:pathology; D016923:Cell Death / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D003402:Creatine Kinase / Q000378:metabolism; D006984:Hypertrophy / Q000378:metabolism* / Q000473:pathology; D018971:Insulin-Like Growth Factor Binding Protein 2 / Q000235:genetics / Q000378:metabolism; D018975:Insulin-Like Growth Factor Binding Protein 5 / Q000235:genetics / Q000378:metabolism; D007334:Insulin-Like Growth Factor I / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D017570:MyoD Protein / Q000235:genetics / Q000378:metabolism; D032446:Myoblasts / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D018008:Myogenin / Q000235:genetics / Q000378:metabolism; D017526:Receptor, IGF Type 1 / Q000235:genetics / Q000378:metabolism; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000235:genetics / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D018971:Insulin-Like Growth Factor Binding Protein 2; D018975:Insulin-Like Growth Factor Binding Protein 5; D017570:MyoD Protein; C057363:MyoD1 myogenic differentiation protein; D018008:Myogenin; D047888:Receptors, Tumor Necrosis Factor, Type I; C506236:Tnfrsf1a protein, mouse; D014409:Tumor Necrosis Factor-alpha; D007334:Insulin-Like Growth Factor I; D017526:Receptor, IGF Type 1; D003402:Creatine Kinase