Ιούλιος 2020, Characterization of Optimal Strain, Frequency and Duration of Mechanical Loading on Skeletal Myotubes' Biological Responses - In Vivo
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Abstract
Background/aim: Mechanical loading of differentiated myoblasts in vitro may mimic loading patterns of skeletal muscle in vivo. However, it is still uncharacterized the loading conditions that can produce the most effective muscle cells' biological responses, in vitro. This study investigated the effects of different loading protocols on the expression of myogenic regulatory factors, anabolic, atrophy and pro-apoptotic factors in skeletal myotubes.
Materials and methods: C2C12 myoblasts were differentiated and underwent various stretching protocols by altering their elongation, frequency and duration, utilizing an in vitro cell tension system. The loading-induced expression changes of MyoD, Myogenin, MRF4, IGF-1 isoforms, Murf1, Atrogin, Myostatin, Foxo and Fuca were measured by Real Time-PCR.
Results: Stretching by 2% elongation at 0.25 Hz for 12 h was overall the most effective in inducing beneficial responses.
Conclusion: A low strain, low frequency intermediate duration stretching can most effectively up-regulate myogenic/anabolic factors and down-regulate pro-apoptotic and atrophy genes in myotubes.
Keywords: Mechanical loading; mechanotransduction; myogenesis; myotubes.
Figure 1.
Effects of cyclic mechanical stretch on the expression of MRFs. Quantitative analysis of (A) MyoD, (B) Myogenin and (C) MRF4 mRNA expression in myotubes subjected to mechanical stretch compared to control (non-stretched myotubes); The mRNA values of MRFs in stretched myotubes were normalized to the corresponding GAPDH mRNA and are expressed as fold changes compared to control. Percentages on the columns represent the degree of elongation of the cell culture surface. Note that in the selected stretching protocols as the elongation and frequency (Hz) decrease the duration (Time) increases. *significantly different compared to control; ¤significantly different compared to 15% elongation; &significantly different compared to 10% elongation/1 Hz; #significantly different compared to 10% elongation/0.25 Hz; ¥significantly different compared to 2% elongation/12 h; Mean+SE of 3 independent experiments performed in triplicate; p<0.05.