Biomaterials for promoting neuron regeneration and directing axon growth following spinal cord injury (SCI) represent a highly promising field for treatment of this devastating condition. In particular, a material that mimics the highly aligned nature of the spinal cord will be particularly effective at directing axon regeneration through the site of injury. A number of studies have shown the effectiveness of gradients of growth factors NGF (for sensory neurons) and NT-3 or BDNF (for motor neurons), at both stimulating and directing axon regrowth following SCI. The proposed work outlines a strategy for functionalizing a highly aligned biomaterial hydrogel with gradients of these growth factors in order to direct and promote axon regeneration. The biomaterial will be composed of peptide amphiphile (PA) molecules that self-assemble into nanofibers and can be used to form a biodegradable hydrogel with the fibers aligned over macroscopic distances. Preliminary work by the applicant has demonstrated that aligned PAgels can efficiently entrap a number of proteins for several weeks, regardless of size and charge, including lysozyme (which has the same size and charge as most neurotrophins). In addition, a stable gradient of protein can be constructed along the aligned axis by using a simple diffusion-based method that allows for direct generation of the gradient immediately prior to biomaterial gelation. Building on this work, the ability of the gel t retain and create a stable gradient of growth factors will be investigated. The gel's ability to immobilize well-defined gradients of neurotrophins, in concentration ranges previously shown to promote bioactivity, will be investigated in detail. Following construction of the gel, neurons wil be incorporated into the material in order to determine its effectiveness at promoting and directing axon growth. Both dorsal root ganglia neurons (which respond to NGF), and corticospinal motor neurons (which respond to NT-3) will be incorporated into PA gels bearing the appropriate neurotrophin gradients. The length and number of axons, as well as their directionality and rate of growth will be determined, and the gradient parameters will be tuned to optimize these measures of axon growth. If successful, the work proposed will create the first biocompatible, degradable, and easily injectable biomaterial that possesses a long-lived gradient of growth factors on a highly aligned and cell-adhesive scaffold. In the long term, this material could be injected and gelled in the injury site following either acute or chronic SCI in order to stimulate axon growth and functional connectivity, reversing or at least mitigating the effects of the injury.
The proposed project, which seeks to synthesize a highly aligned and injectable scaffold bearing long-lived gradients of neuroactive growth factors, will create a novel biomaterial for neuron regeneration in spinal cord injury. The gradients of growth factors immobilized on the material will help stimulate axon growth, as well as direct it in the proper direction. The long term application of this material will be as a highly translatable medical treatment to help promote axon regeneration in order to reestablish functional connectivity following spinal cord injury (acute or chronic), restoring both sensory and motor control and mitigating the deleterious effects of the injury.
|Greene, Jacqueline J; McClendon, Mark T; Stephanopoulos, Nicholas et al. (2018) Electrophysiological assessment of a peptide amphiphile nanofiber nerve graft for facial nerve repair. J Tissue Eng Regen Med 12:1389-1401|
|Freeman, Ronit; Stephanopoulos, Nicholas; Álvarez, Zaida et al. (2017) Instructing cells with programmable peptide DNA hybrids. Nat Commun 8:15982|
|Rubert Pérez, Charles M; Stephanopoulos, Nicholas; Sur, Shantanu et al. (2015) The powerful functions of peptide-based bioactive matrices for regenerative medicine. Ann Biomed Eng 43:501-14|
|Stephanopoulos, Nicholas; Freeman, Ronit; North, Hilary A et al. (2015) Bioactive DNA-peptide nanotubes enhance the differentiation of neural stem cells into neurons. Nano Lett 15:603-9|
|Stephanopoulos, Nicholas; Ortony, Julia H; Stupp, Samuel I (2013) Self-Assembly for the Synthesis of Functional Biomaterials. Acta Mater 61:912-930|