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Title: A new cell-based assay to evaluate myogenesis in mouse myoblast C2C12 cells

The development of the efficient screening system of detecting compounds that promote myogenesis and prevent muscle atrophy is important. Mouse C2C12 cells are widely used to evaluate myogenesis but the procedures of the assay are not simple and the quantification is not easy. We established C2C12 cells expressing the N-terminal green fluorescence protein (GFP) and the C-terminal GFP (GFP1–10 and GFP11 cells). GFP1–10 and GFP11 cells do not exhibit GFP signals until they are fused. The signal intensity correlates with the expression of myogenic markers and myofusion. Myogenesis-promoting reagents, such as insulin-like growth factor-1 (IGF1) and β-guanidinopropionic acid (GPA), enhance the signals, whereas the poly-caspase inhibitor, z-VAD-FMK, suppresses it. GFP signals are observed when myotubes formed by GFP1–10 cells are fused with single nuclear GFP11 cells, and enhanced by IGF1, GPA, and IBS008738, a recently-reported myogenesis-promoting reagent. Fusion between myotubes formed by GFP1–10 and GFP11 cells is associated with the appearance of GFP signals. IGF1 and GPA augment these signals, whereas NSC23766, Rac inhibitor, decreases them. The conditioned medium of cancer cells suppresses GFP signals during myogenesis and reduces the width of GFP-positive myotubes after differentiation. Thus the novel split GFP-based assay will provide the useful method for the studymore » of myogenesis, myofusion, and atrophy. - Highlights: • C2C12 cells expressing split GFP proteins show GFP signals when mix-cultured. • The GFP signals correlate with myogenesis and myofusion. • The GFP signals attenuate under the condition that muscle atrophy is induced.« less
Authors:
 [1] ;  [1] ;  [2] ; ;  [1] ;  [1] ;  [2] ; ;  [1] ;  [3] ;  [4] ;  [1] ;  [5] ;  [6] ;  [5] ;  [7] ;
  1. Department of Medical Biochemistry, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo (Japan)
  2. (China)
  3. Department of Breast Surgery, The Second Affiliated Hospital of Wenzhou Medical University, Wenzhou (China)
  4. Department of Chemistry, School of Science, The University of Tokyo, Tokyo (Japan)
  5. Chemical Biology Screening Center, Tokyo Medical and Dental University, Tokyo (Japan)
  6. Institute of Biomaterials and Bioengineering, Tokyo Medical and Dental University, Tokyo (Japan)
  7. (Japan)
Publication Date:
OSTI Identifier:
22462316
Resource Type:
Journal Article
Resource Relation:
Journal Name: Experimental Cell Research; Journal Volume: 336; Journal Issue: 2; Other Information: Copyright (c) 2015 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; ATROPHY; CARCINOMAS; FINGERS; FLUORESCENCE; GROWTH FACTORS; INSULIN; LUNGS; MICE; MUSCLES; MYOSIN; REAGENTS; RINGS; SCREENING; SIGNALS; WIDTH