Effects of differentiation and mechanical stimulation on the morphology, protein expression, and mechanical properties of mammalian myocytes

The adaptive responses of skeletal muscle to exercise and mechanical loading are often beneficial. However, muscle can also atrophy, a serious consequence of human spaceflight. The research goal was to determine the effects of mechanical stimulation on mammalian skeletal muscle cell morphology and mechanical properties. Additional objectives were to characterize mechanical properties and protein expression of myocytes throughout differentiation. This work will advance the understanding of how muscle responds to loading and aid in the optimization of preventative countermeasures for spaceflight.; The objectives were accomplished by using an in vitro model to mechanically stimulate murine C2C12 myoctes and isolate the effects of stretch on cellular responses. Morphology changes were determined by measuring orientation and length of fibers. Changes in mechanical properties were identified using atomic force microscopy to measure the apparent elastic modulus and hysteresis of the cells. Levels of protein expression were semi-quantified using Western blots.; A continuous unidirectional stretch, simulating long bone growth, caused myocytes to lengthen and orient along the direction of stretch. Cellular responses were independent of strain rate for the range tested. The apparent elastic modulus was a function of differentiation, increasing after eight days of differentiation into myofibers from myoblasts. Hysteresis was independent of differentiation. Neither expression of myosin or tubulin proteins correlated to changes in mechanical properties. Microtubules were also not found to be major contributors to the mechanical properties.; Exercise and hypertrophy models were evaluated at 12 and 24 hours to determine the effects of oscillatory and constant strain on mechanical properties. Both mechanical stretch and time significantly affected mechanical properties. Each type of stretch caused different cellular responses, resulting in independent changes in elastic and viscous properties.; This work reveals the effects of stretch on myocyte mechanical and morphological properties, indicating the importance of loading in the maintenance and function of skeletal muscle. These results suggest that stretching protocols for spaceflight and in 1 g may benefit from cyclic and constantly held stretching. The characterization of cellular mechanical properties as a function of differentiation has application in tissue engineering where the optimization of functional compatibilities for tissue replacement is critical.