RESUMO
In this study. we investigate the potential for manipulating bone cell mechanotransducers in tissue engineering. Membrane ion channels such as voltage operated calcium channels (VOCC) have been shown to be a critical component of the bone cell transduction pathway with agonists and inhibitors of this pathway having profound effects on the load signal. By encapsulating a calcium channel agonist with slow release within a poly(L-lactide) (PLLA) scaffold, we can generate a 'mechano-active' scaffold for use in skeletal tissue engineering. PLLA scaffolds with and without a calcium channel agonist, BAY K8644, were seeded with primary human bone cells or the human MG63 bone cell line and cultured for 13 weeks followed by mechanical stimulation with a four-point bending model. Our results show that addition of the agonist for slow release is sufficient to enhance the load-related responses in bone cells within the scaffolds. Specifically, collagen type I expression and the ratio of alkaline phosphatase to protein are elevated in response to cyclical mechanical stimulation of approximately 1000 microstr which is then further enhanced in the mechano-active' scaffolds. As the agonists only act when the calcium channels are open by attenuating the calcium flux, the stimulation is specifically targeted to scaffolds subjected to load either in vitro or ultimately in vivo. Our results suggest that manipulating the VOCC and attenuating the opening of the calcium channels may be an effective technique to amplify matrix production via mechanical stimulation which may be applied to bone tissue engineering and potentially engineering of other load-bearing connective tissues.
