| Spinal cord injury is a significant clinical challenge for which few treatment options are available. The complex biology of the central nervous system imparts many requirements to an effective therapy, such as reducing scar formation, providing growth factors to stimulate regeneration, and suppressing inhibitory signals. This research project has been focused at developing a combinatorial tissue engineering approach to facilitate nerve regeneration. In the proposed strategy, degradable polymer structures serve as cellular scaffolding and/or nerve guidance material while simultaneously functioning as a gene delivery vehicle to allow local expression of a therapeutic protein. Avoiding viral methods and the associated potential for immune response, this research has employed plasmid DNA that is complexed with a cationic lipid (referred to as lipoplexes) and is released from the FDA-approved degradable polymer poly(lactide-co-glycolide)(PLG). Methods have been established to incorporate lipoplexes into three-dimensional structures, from which they are released for over 50 days, thus constituting the first reported biomaterial-mediated sustained release of lipoplexes. Lipoplexes released from PLG disks retain their transfection activity, verified by the expression of both reporter proteins and biologically active nerve growth factor (NGF) in a variety of in vitro culture systems. Since non-viral methods are inefficient at transfecting neurons, a co-culture system was developed, in which PLG-mediated transfection was used to express NGF in accessory cells, which in turn produced sufficient levels of NGF to promote neurite outgrowth from primary neurons. Lipoplex-releasing PLG disks were shown to be effective for transfection in vivo by intramuscular implantation, producing a significant increase in luciferase expression relative to naked plasmid DNA released from PLG. Lastly, multilumen nerve guidance scaffolds were applied in preliminary studies of three different rat models of spinal cord injury. PLG scaffolds were shown to be capable of good apposition and integration with host spinal cord tissue. This research has set the stage for a novel and highly cross-disciplinary treatment strategy combining the physical benefits of a biocompatible bridging material with the local release of lipoplexes to express neurotrophic factors. This multifold approach offers a highly flexible and integrative method to potentiate regeneration following spinal cord injury. |
