Two central problems exist in nerve repair after traumatic injury. The first is finding a nerve guide suitable for large gaps in large diameter nerves. The second is that regeneration from proximal injuries occurs too slowly, which allows target muscle to atrophy, preventing reinnervation and functional motor recovery after regenerating neurites arrive. Nerve guides that accelerate regeneration could improve motor recovery after nerve repair. Most nerve guides produce inferior results, even when filled or coated with extracellular matrix proteins (ECM) or growth factors that promote neurite outgrowth, including laminin, regarded as the most potent substrate-bound promoter of neurite outgrowth. Recently, aligned nanofibers have been introduced into nerve guidance channels to provide topographical cues that direct regenerating neurites. These fibers have been found to orient, direct, and promote faster growth of regenerating neurites in vitro and in vivo. However, their performance still lags behind autografts. Since the primary problem in achieving functional regeneration slow speed, we hypothesize that taking peptide sequences from laminin that promote neurite outgrowth and covalently-binding them to aligned nanofibers could speed nerve regeneration and make it more effective. A proposal to fabricate and test such fibers is presented.
Specific aim 1 begins with electrospinning poly(lactide-co-glycolide) (PLGA) nanofibers that can be covalently bound to IKVAV, a peptide in laminin that promotes neurite growth. Fibers will be characterized for alignment, diameter, and rate of degradation. Dissociated motor neurons and dorsal root ganglia explants will be grown on these surfaces and their neurite outgrowth assayed to compare to controls without IKVAV and to laminin-coated controls.
In specific aim 2, gradients of IKVAV and YIGSR, another laminin peptide, will be covalently bound to PLGA nanofibers to see if gradients cause faster neurite growth. These fibers will be tested similarly as fibers in aim 1.
In specific Aim 3, longitudinally-oriented aligned nanofibers will be placed in semipermeable membranes as nerve guides and implanted in rat sciatic nerve gaps. In these experiments, PLGA with covalently bound fibers as developed in the first two aims will be compared against plain fibers and autologous nerve grafts. It is hoped that this project will further knowledge of both the production of functionalized nanofiber nerve guides and their potential to be harnessed as a tool for nerve regeneration.
Over 21,000 combat wounded American servicemen and women have sustained injuries to the peripheral nerves of the face and limbs since March 2003. This produces dramatic disability impacting activities of daily life. Nerve injury is also a common cause of disability in the civilian population. Unfortunately, the present technology of surgically implanted nerve guidance channels used to promote regeneration after injury produces poor outcomes, leaving patients disabled. We have developed aligned polymer nanofibers that direct the growth of regenerating axons in tissue culture. We also have preliminary evidence that implanting these nanofibers in surgically created nerve injuries in rats helps nerves to regenerate. Here we propose binding the surfaces of these nanofibers with segments of protein known to make nerve cells grow faster. The proposed experiments may lead to improved nerve guidance channels that produce better nerve regeneration for injured soldiers and civilians alike.
