Effects of stretching stress on the muscle contraction proteins of skeletal muscle myoblasts

Abstract

Several studies have reported that growth and differentiation of cultured myoblasts can be facilitated by applying appropriate mechanical stimulus. However, the effects of mechanical stimulus on the characteristics of muscle fi bers have not yet been fully elucidated. In this study, we gave mechanical stress to C2C12 cells, which were myoblasts derived from mice skeletal muscle. The following myosin heavy chain (MHC) isoforms were investigated in order to clarify muscle characteristics: MHC-2b, 2d and 2a, all of which are fast-twitch fi bers. After inoculating cells on a silicone chamber, the chamber was mechanically stretched, and a LightCycler TM was used to measure the mRNA expression of each MHC isoform at several times. The results showed that, with mechanical stretching, the expression of MHC-2b was initially high. On the other hand, without stretching, the expression of MHC-2d increased over time, but with stretching, it was hardly seen. Furthermore, the expression of MHC-2a was signifi cantly high in the stretching group. These results of this study suggest that, when intermittently stimulated, myoblasts express increased levels of MHC-2a isoform. Therefore, it is indicated that myocytes respond to environmental changes not only to facilitate growth and differentiation, but also to alter muscle function actively at the MHC isoform level.Myosin heavy chain (MHC), which is a muscle contraction protein, is known to be closely related to muscle function (14). Also, MHC consists of several isoforms, and changes in muscle function can be determined by the composition ratios of these isoforms (2, 14). In adult mice, there are four MHC isoforms; MHC-2b, 2d, 2a and 1, and in vivo studies have shown that these isoforms may undergo the following reversible changes: 2b → ← 2d → ← 2a → ← 1 (15). MHC-2b, 2d and 2a are all fast-twitch fi bers, and the contraction speed for MHC-2b is the fastest, followed by 2d and 2a, and MHC-1 is slow-twitch fiber (1).As to the relationship between the composition ratios of MHC isoforms and changes in muscle function, one study investigated the masseter muscle in fetal and newborn mice (19). The results showed that the composition ratios of MHC isoforms changed markedly at 16 days post conception when many oral tissues became ready for delivery and slight jaw movements began. Another study investigated the development process for the mouse masseter muscle after birth, and reported that the composition ratios of MHC isoforms changed markedly during weaning when the masseter muscle began to masticate instead of suckle (6, 17). These two investigations suggested a possibility that changes in muscle function leads to changes in MHC isoform composition. However, it is also possible that changes in MHC isoform are related to

Expression of Myosin Heavy-chain mRNA in Cultured Myoblasts Induced by Centrifugal Force

“…This result is consistent with the findings of a study by Sakiyama et al 13) demonstrating the enhancement of myoblast differentiation induced by extending stimulation as observed by electron microscopy.…”

“…Naumann and Pette administered electrical stimulation to cells obtained from rat hind leg and soleus muscles via culture medium and reported its effects on expression of myosin heavy-chain (MyHC) isoforms 10) . Sakiyama et al reported differences in myoblast proliferation and muscle fiber characteristics induced by extending stimulation 13) . Sakuraba et al examined the effects of magnetic stimulation of rat hind leg muscles on expression of mRNA by comparing stimulation and control groups 14) .…”

Effects of stretching stimulation with different rates on the expression of MyHC mRNA in mouse cultured myoblasts

“…Culture medium containing 2% FBS was used after allowing cells to adhere to the membrane for 1 day. The cells were then stretched according to the method of Sakiyama et al (21).…”

“…The stretch rate was established using the stretch rate setting method of a Flexercell ® strain unit. Since Sakiyama et al (21) performed an experiment at a stretch rate of 0.5/sec in a previous report, this condition was adopted as the standard. This rate represents the distance of membrane movement per second: 1/sec indicates that a membrane stretches from a stationary state then relaxes to return to the original state in 1 second.…”

“…Magnetic stimuli were applied to rat hindlimb muscle to compare mRNA expression with that of controls (22). Furthermore, in the third study (21), cells were stretched to determine changes in number of cell and nuclei and muscle fiber characteristics.…”