Controlled release of vascular endothelial growth factor (VEGF) and pleiotrophin (PTN) stimulates nerve regeneration across long gap peripheral nerve defects

Peripheral nerve injury gaps resulting from tissue loss after trauma or tumor resection are routinely repaired by autogenous nerve grafts which provide partial sensory-motor recovery at the expense of normal donor nerves. To obviate the need of harvesting donor tissue, short nerve gaps (i.e., 1-2.5 cm) have been effectively bridged by synthetic nerve guides. However, nerve regeneration across gaps longer than 3 cm have failed when repaired using natural or synthetic nerve scaffolds. We hypothesize that neurotrophic support and early vascularization of the regenerated nerve are critical for successful nerve regeneration across long-gap nerve defects. Vascular endothelial growth factor (VEGF) and Pleiotrophin (PTN) are potent endothelial cell mitogens that promote angiogenesis. In addition, both PTN and VEGF have growth promoting effects on neurons in the central and peripheral nervous systems. The combination of neurotrophic and angiogenic activities of these molecules suggests them as candidates to promote simultaneous vascularization and nerve regeneration across long-gap injuries in peripheral nerves. To evaluate this hypothesis we developed a polymeric growth factor delivery system releasing VEGF or PTN and investigated its angiogenic and neurotrophic potential both in-vitro and in-vivo. PLGA nanospheres prepared by the double emulsion technique showed a sustained protein release over a period of four weeks. Robust angiogenesis was seen in adult aortic explant cultures supplemented with VEGF nanospheres. Similarly, increased neurite outgrowth was observed in ventral spinal motor neuron cultures treated with PTN nanospheres.;Using this system the regenerative response across a 3 cm nerve gap in the injured rabbit common peroneal nerve was evaluated in animals that received control treatments: (a) collagen filled simple tube and (b) multiluminal biosynthetic nerve implant (BNI) filled with collagen, and compared to those in which the BNI was supplemented with either (c) VEGF or (d) PTN nanospheres. Histological evaluation ten to twelve weeks post implantation revealed that nerve defects repaired with simple tubes failed to regenerate across the long gap, while those bridged with collagen-filled BNIs showed vascularized nerves within the hydrogel micro-channels. The number of neurofilament-positive axons within the regenerated microfascicles increased significantly in the BNIs supplemented with VEGF or PTN. Furthermore, functional recovery was demonstrated by recovery of target muscle mass, compound motor action potentials, and the behavioral toe-spreading assay. Our study indicates that nerve and vascular growth factors can stimulate nerve regeneration across long-gap peripheral nerve defects.