Nerve conduits based on immobilization of nerve growth factor onto modified chitosan by using genipin as a crosslinking agent

Abstract

Incorporation of nerve growth factor (NGF) into a nerve conduit can improve peripheral nerve regeneration. Here, genipin, a natural and low toxic agent, was used to crosslink chitosan, a natural polysaccharide, and concurrently to immobilize NGF onto modified chitosan, followed by fabrication of chitosan (CS)–genipin (GP)–NGF nerve conduits. MTT test showed that the cell viability of Schwann cells cultured in the conduit extract was not significantly different from that in plain medium. The neurite outgrowth measurement and immunocytochemistry with anti-growth-associated protein-43 and anti-neurofilament indicated that NGF released from CS–GP–NGF nerve conduits retained the bioactivity of stimulating neuronal differentiation of PC12 cells. Fracture strength measurements and vitamin B12 release analysis confirmed that CS–GP–NGF nerve conduits possessed good mechanical properties and adequate permeability. We also investigated the in vitro release kinetics of NGF from CS–GP–NGF nerve conduits by ELISA. The continuous release profile of NGF, within a 60-day time span, consisted of an initial burst that was controlled by a concentration gradient-driven diffusion, followed by a zero-order release that was controlled by a degradation of chitosan matrix. Collectively, CS–GP–NGF nerve conduits had an integrated system for continuous release of NGF, thus holding promise for peripheral nerve repair applications.