Biomimetic Injectable 3D Hydrogels with Aligned Topography for Neural Tissue Engineering

Spinal cord trauma leads to destruction of the highly organized cytoarchitecture that carries information along the axis of the spinal column. Currently, there are no clinically accepted strategies that can help regenerate severed axons after spinal cord injury. Experimental neuroregenerative efforts include the creation of optimal biomaterials with aligned topography to support enhanced neuronal regeneration. Hydrogels are soft biomaterials with high water content that are widely used as scaffolds to interface with the central nervous system (CNS). Current available methods to create topography within the 3D amorphous hydrogels are typically complex. Here we examine a simple and reproducible method that results in consistently aligned fibrils within 3D matrices using thermally gelling biomimetic polymers that are compatible with neuronal cells. A collagen type I (Col)-based thermally gelling hydrogel system was used in combination with two other native extracellular matrix proteins: laminin I (LN), and hyaluronic acid (HA). Gelling kinetics for all gel types (Col, Col LN, Col HA) were examined, and we found that all three combinations of polymer formed consistent gels at 37°C. Col solution was faster to form gels (17 min), while Col LN and Col HA took longer (~22 minutes). A method of aspiration and ejection was used to produce Col-based hydrogels containing aligned fibrils. Gels were then examined using scanning electron microscopy (SEM). SEM images confirmed successful alignment in all gel types, and the size of fibers was consistent with reported values for collagen (~250 nm in diameter). We found that embryonic spinal cord neurons survive and produce processes aligned to collagen fibrils after 14 days in vitro. Next, we investigated the functionality of aligned and non-aligned Col hydrogels implanted acutely after a contusion type spinal cord injury to the thoracic spinal cord at T7/T8. Pan neuronal antibody-positive fibrils were found within all implants, aligned hydrogels supported neurite growth along the parallel direction of the implanted hydrogels. Our data indicate that thermally gelling biomimetic hydrogels can produce aligned matrices by a method of aspiration and ejection. The material composition and process of aligning hydrogels outlined here presents a novel platform for regenerative therapies for the CNS that is compatible with the survival and growth of neuronal cells in vitro and in vivo.