Controlled release of nerve growth factor from a heparin-containing fibrin-based cell ingrowth matrix

The goal of this work was to develop a growth factor delivery system for use in nerve regeneration that would provide localized release of beta-nerve growth factor (beta-NGF) and other members of the neurotrophin family in a controlled manner. Although beta-NGF does not bind heparin with high affinity, we postulated that a basic domain found at the surface of native beta-NGF could interact with heparin and slow its diffusion from a heparin-containing delivery system. To test this hypothesis, we used a heparin-containing fibrin-based cell ingrowth matrix consisting of three components, namely an immobilized heparin-binding peptide, heparin and a neurotrophin with low heparin-binding affinity. The heparin-binding peptide contained a factor XIIIa substrate and was covalently cross-linked to fibrin matrices during polymerization. This cross-linked heparin-binding peptide served to immobilize heparin within the matrix, and this immobilized heparin interacted with the neurotrophin and slowed the passive release of the growth factor from the matrix. The ability of heparin-containing fibrin matrices, with a high excess of heparin-binding sites, to slow the diffusion-based release of beta-NGF from fibrin matrices was measured in the absence of cells. Conditions that provided for slow diffusion-based release of beta-NGF, brain-derived neurotrophic factor, and neurotrophin-3 were tested in an assay of neurite extension from dorsal root ganglia to determine the ability of the delivery system to release active growth factor. The results demonstrated that neurotrophins, interacting with fibrin matrices containing a large molar excess of heparin relative to growth factor, enhanced neurite extension by up to 100% relative to unmodified fibrin. In the absence of the delivery system, free neurotrophins within the fibrin matrix did not enhance neurite extension. The results suggest that these matrices could serve as therapeutic materials to enhance peripheral nerve regeneration through nerve guide tubes and may have more general usefulness in tissue engineering for the delivery of non-heparin-binding growth factors.