| Summary: | Peripheral nerve injuries are one of the most prevelant trauma injuries, and the current golden standard for treatment is an autologous nerve graft. The use of tissue engineered nerve grafts has increased in recent years, a neural scaffold, cellular or acellular, is utilized to promote nerve repair. Prior research has shown that transcutaneous optogenetic stimulation on grafted engineered tissue can promote reinnervation and angiogensis in a rat volumetric muscle loss model. Our study queried the individual effects of biochemical and mechanical stimuli on neuronal growth to push the field of neuromuscular systems forward. We utilized optogenetic stimulation to emulate the biochemical effects and a magnetic fibrin platform to isolate the mechanical effect. To develop our magnetically actuatable substrate we optimized a fibrin hydrogel that would have a similar stiffness to skeletal muscle. Then, we added rectangular segments of 1:10 PDMS with 25% v/v 4 micron iron microparticles. These rectangular segments within the fibrin hydrogel were then cyclically actuated by a permanent neodymium magnet. Our results showed a substantial increase in neurite outgrowth in the experimental group which was supplemented with optogenetically exercised media from a muscle monolayer. The isolated biochemical effect was a substantial increase in the rate of neurite growth between the groups. In our preliminary neuromuscular system, we saw a degree of co-localized alignemnt between the neurites and differentiated muscle. This neuromuscular protocol seems to have physiological alignment similarities to in vivo tissue. We quantified alignment through a Fast Fourier Transform of the image data of the separate imaging channels, RFP for muscle and GFP for motor neuron. Finally, our magnetic fibrin platform found no significant increase in myofiber length and width when mechanical stimulation was applied after myoblast differentiation. In future research, we are exploring the biochemical and magnetic stimulation on our neuromuscular co-culture and actuate the myoblasts at an earlier cellular stage to impact alignment. In conclusion, our studies found that stimulated media aids in neurite outgrowth. In future research, we will perform RNAseq on our systems to verify the specific biological pathways and upregulated growth factors.
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