Cell Specific Guidance Of Nerve And Vascular Regeneration Through Sustained Growth Factor Release

Abstract

Regenerative peripheral nerve interfaces have been proposed as viable alternatives for the natural control of robotic prosthetic devices. However, sensory and motor axons at the neural interface are of mixed submodality types, which difficult the specific recording from motor axons and the eliciting of precise sensory modalities through selective stimulation. Here we evaluated the possibility of using type-specific neurotrophins to preferentially entice the regeneration of defined axonal populations from injured peripheral nerves. Segregation of mixed sensory fibers from dorsal root ganglion neurons was evaluated in vivo by compartmentalized diffusion delivery of nerve growth factor (NGF) and neurotrophin-3 (NT-3), to preferentially entice the growth of TrkA+ nociceptive and TrkC+ proprioceptive subsets of sensory neurons, respectively. A “Y”-shaped tubing was used to allow regeneration of the transected adult rat sciatic nerve into separate compartments filled with either NFG or NT-3. A significant increase in the number of CGRP+ pain fibers were attracted towards the sural nerve, while N-52+ large diameter axons were observed in the tibial and NT-3 compartments. Conversely, the preferential growth of pain fibers (CGRP+) and motor fibers (ChAT+) was evaluated using a polymeric sustained release method. Growth factor loaded PLGA microparticles created a gradient for a longer time compared to when they used directly. Two specific growth factors for pain fibers (NGF) and motor neurons (Pleiotrophin factor; PTN) were encapsulated in microparticles and tested in a femoral nerve double crush injury model. The common femoral nerve bifurcates to two branches: saphenous branch (SB) and motor branch (MB). After injury in the common femoral nerve, the sensory and motor axonal fibers are mixed and lose their ability to innervate the proper target. We used PLGA encapsulated NGF and PTN to guide the axons to their respective target. The results of this study confirmed that more nociceptive fibers and motor axons were attracted to NGF and PTN, respectively. The experimental group treated with BSA (negative control) did not show any preferential growth in either of branches. This study demonstrates the guided enrichment of sensory and motor axons and, and supports the notion that neurotrophic factors can be used to segregate sensory and perhaps motor axons in separate peripheral interfaces. In order to test if a growth factor gradient will enhance axonal guidance, we developed a method that consistent of coiling a polymeric fiber inside hydrogel microchannels. We used finite element analysis (FEA) to model drug delivery and to compare the diffusion dynamics between the non-gradients versus gradient model. This is needed both for the repair of sensory and motor branches and for the development of closed-loop peripheral neural interfaces. This strategy can be used to entice a specific sub-type of axons in a mixed population of nerves to the channels and eventually will guide them to the proper target. Together the data indicates that modality specific growth factors can be used to enrich the motor and sensory axonal fibers guidance to the correct target. However; the separation of the different subtype of the axonal fibers was not completely achieved indicating that either more than one growth factor or a combination of neurotrophic and pleiotrophic factors is needed to separate the axonal fibers more precisely. In addition, computer modeling and initial experiments indicate that gradient release of growth factor may help further in achieving enhanced selective nerve growth.