In vitro development of a tissue-engineered model of peripheral nerve regeneration to study neurite growth.

Abstract

A unique tissue-engineered model of peripheral nerve regeneration was developed in vitro to study neurite outgrowth. Mouse dorsal root ganglia neurons were seeded on a collagen sponge populated with human endothelial cells and/or human fibroblasts. Addition of nerve growth factor (NGF; 10 ng/ml) was not required for sensory neurons survival but was necessary to promote neurite outgrowth, as assessed by immunostaining of the 150 kDa neurofilament. A vigorous neurite elongation was detected inside the reconstructed tissue after 14 and 31 days of neurons culture, reaching up to 770 microm from day 14. Axons were often observed closely associated with the capillary-like tubes reconstructed in the model, in a similar pattern as in the human dermis. The presence of endothelial cells induced a significant increase of the neurite elongation after 14 days of culture. The addition of human keratinocytes totally avoided the twofold decrease in the amount of neurites observed between 14 and 31 days in controls. Besides the addition of NGF, axonal growth did not necessitate B27 supplement or glial cell coculture to be promoted and stabilized for long-term culture. Thus, this model might be a valuable tool to study the effect of various cells and/or attractive or repulsive molecules on neurite outgrowth in vitro.