Differentiation of mammalian skeletal muscle cells cultured on microcarrier beads in a rotating cell culture system

The growth and repair of adult skeletal muscle are due in part to activatio n of muscle precursor cells, commonly known as satellite cells or myoblasts . These cells are responsive to a variety of environmental cues, including mechanical stimuli. The overall goal of the research is to examine the role of mechanical signalling mechanisms in muscle growth and plasticity throug h utilisation of cell culture systems where other potential signalling path ways (i.e. chemical and electrical stimuli) are controlled To explore the e ffects of decreased mechanical loading on muscle differentiation, mammalian myoblasts are cultured in a bioreactor (rotating cell culture system!, a m odel that has been utilised to simulate microgravity. C2C12 murine myoblast s are cultured on microcarrier beads in a bioreactor and followed throughou t differentiation as they form a network of multinucleated myotubes. In com parison with three-dimensional control cultures that consist of myoblasts c ultured on microcarrier beads in teflon bags, myoblasts cultured in the bio reactor exhibit an attenuation in differentiation. This is demonstrated by reduced immunohistochemical staining for myogenin and alpha -actinin. Weste rn analysis shows a decrease, in bioreactor cultures compared with control cultures, in levels of the contractile proteins myosin (47% decrease, p< 0. 01) and tropomyosin (63% decrease, p<0.01). Hydrodynamic measurements indic ate that the decrease in differentiation may be due, at least in part, to f luid stresses acting on the myotubes. In addition, constraints on aggregate size imposed by the action of fluid forces in the bioreactor affect differ entiation. These results may have implications for muscle growth and repair during spaceflight.