Response of the porcine MYH4-promoter and MYH4-expressing myotubes to known anabolic and catabolic agents in vitro

Brearley, Madelaine C.; Loczenski-Brown, David M.; Loughna, Paul T.; Parr, Tim; Brameld, John M.

Abstract

© 2021 The Authors Myosin heavy chain-IIB (MyHC-IIB; encoded by MYH4 or Myh4) expression is often associated with muscle hypertrophic growth. Unlike other large mammals, domestic pig breeds express MyHC-IIB at both the mRNA and protein level. Aim: To utilise a fluorescence-based promoter-reporter system to test the influence of anabolic and catabolic agents on increasing porcine MYH4-promoter activity and determine whether cell hypertrophy was subsequently induced. Methods: C2C12 myoblasts were co-transfected with porcine MYH4-promoter-driven ZsGreen and CMV-driven DsRed expression plasmids. At the onset of differentiation, treatments (dibutyryl cyclic-AMP (dbcAMP), Des(1–3) Insulin-Like Growth Factor-1 (IGF-I), triiodo-L-thyronine (T3) and dexamethasone (Dex)) or appropriate vehicle controls were added and cells maintained for up to four days. At day 4 of differentiation, measurements were collected for total fluorescence and average myotube diameter, as indicators of MYH4-promoter activity and cell hypertrophy respectively. Results: Porcine MYH4-promoter activity increased during C2C12 myogenic differentiation, with a marked increase between days 3 and 4. MYH4-promoter activity was further increased following four days of dbcAMP treatment and average myotube diameter was significantly increased by dbcAMP. Porcine MYH4-promoter activity also tended to be increased by T3 treatment, but there were no effects of Des(1–3) IGF-I or Dex treatment, whereas average myotube diameter was increased by Des(1–3) IGF-I, but not T3 or Dex. Conclusion: Porcine MYH4-promoter activity responded to dbcAMP, Des(1–3) IGF-I and T3 treatment in vitro as observed previously in reported in vivo studies. However, we report that increased MYH4-promoter activity was not always associated with muscle cell hypertrophy. The fluorescence-based reporter system offers a useful tool to study muscle cell hypertrophic growth.

Citation

Brearley, M. C., Loczenski-Brown, D. M., Loughna, P. T., Parr, T., & Brameld, J. M. (2021). Response of the porcine MYH4-promoter and MYH4-expressing myotubes to known anabolic and catabolic agents in vitro. Biochemistry and Biophysics Reports, 25, Article 100924. https://doi.org/10.1016/j.bbrep.2021.100924

• Journal Article Type: Article
• Acceptance Date: Jan 17, 2021
• Online Publication Date: Feb 2, 2021
• Publication Date: Mar 1, 2021
• Deposit Date: Feb 5, 2021
• Publicly Available Date: Feb 5, 2021
• Journal: Biochemistry and Biophysics Reports
• Electronic ISSN: 2405-5808
• Publisher: Elsevier
• Peer Reviewed: Peer Reviewed
• Volume: 25
• Article Number: 100924
• DOI: https://doi.org/10.1016/j.bbrep.2021.100924
• Public URL: https://nottingham-repository.worktribe.com/output/5293791
• Publisher URL: https://www.sciencedirect.com/science/article/pii/S2405580821000182