Development of a new miniaturized bioreactor for axon stretch growth.

Peripheral nerve injury requires a physical bridge across the lesion, which is limited by the insufficient supply of donor nerves. Here, we developed a new miniaturized bioreactor system for axon stretch growth. Dorsal root ganglia explants were first placed on two adjoining substrates and formed new synaptic connections. The axon bundles across the border between the top and bottom membranes were then stretched in a stepwise fashion by a microstepper motor system. After several days of stretch, the axon tracts could reach lengths that could develop into living nervous tissue constructs. In order to achieve appropriate neuronal culture to stimulate physiological conditions during axon stretch, we tested a variety of coating methods. Based on these results, the elongator substrates were coated with both poly-D-lysine and rat-tail collagen to maximize the number of axon bundles. Additionally, we found that increasing the axon stretch by 1[Formula: see text][Formula: see text]m at each step resulted in the highest stability. The bridging axons adapted to the stretch by increasing their length from 500[Formula: see text][Formula: see text]m to 5.94[Formula: see text]mm over 7 days of stretch growth. Immunocytochemical analysis confirmed that beta-III-tubulin, a major cytoskeletal constituent and neuronal marker, was present along axons. The findings demonstrate that bioreactor has the potential to generate transplant materials to address neural repair.