Lentiviral Gene Therapy from Multi-channel Bridges to Investigate Acute and Chronic Spinal Cord Repair

Spinal cord injury (SCI) causes paralysis below the level of injury which, at the cellular level, results from neuron and oligodendrocyte cell death, axonal loss, demyelination, and critically, the limited capacity of spinal cord neurons to regenerate1. Although central nervous system (CNS) tissue has the innate capacity to repair the local environment that develops after SCI lacks sufficient factors that promote regeneration and has an abundance of factors that inhibit regeneration2. Many strategies have been attempted to aid in the regeneration of CNS tissue, yet re-entry into intact spared tissue remains inadequately low. This is due to the complexity of the spinal cord microenvironment post injury and the barriers that must be addressed simultaneously to elicit adequate regeneration. The Shea lab has developed multi-channel poly (lactide-co-glycolide) (PLG) bridges to promote spinal cord regeneration and restore functional losses. These bridges are biodegradable and provide a temporary structure that promotes regeneration3. These bridges also serve as a platform for delivery of therapeutics including pharmacological agents, cells, and notably lentivirus. This dissertation investigated the use of lentiviral gene therapy from multi-channel bridges to barriers to regeneration during acute and chronic phases of SCI. We investigated myelination of regenerating axons by over-expression of platelet-derived growth factor-AA (PDGF) and noggin either alone or in combination in an acute mouse SCI model. The combination of noggin + PDGF enhanced total myelination of regenerating axons and notably oligodendrocyte myelination. Importantly, the increase in oligodendrocyte myelin enhanced functional recovery and was also associated with a greater density of cells of an oligodendroglial lineage. We investigated synergistic effects of anti-inflammatory and regenerative factors by bi-cistronic delivery of NT-3 and IL-10 using PLG bridges after acute SCI. This work was motivated by the need to delivered multiple factors simultaneously to address the multiple barriers to SCI regeneration. The combination of IL-10+NT-3 enhanced axonal growth and oligodendrocyte myelinated axon density significantly over control, although these factors do not act on oligodendrocyte cells directly. The increased oligodendrocyte myelin resulted in increased locomotor functional recovery compared to IL-10 or NT-3. Furthermore, we observed a strong positive correlation between oligodendrocyte myelinated axon density and functional recovery. These results show oligodendrocyte myelination as a limiting step in attaining enhanced functional recovery. Lastly, we investigated regeneration using the multi-channel bridge implanted into a chronic SCI following surgical resection of necrotic tissue. We noted that scar formation decreased at 4 and 8 weeks post injury (wpi), yet macrophage infiltration increased between 4 and 8 wpi. Subsequently, scar tissue was resected and bridges were implanted at 4 and 8 wpi. We observed robust axon growth into the bridge and remyelination at 6 months post initial injury. Axon densities were increased for 8 week bridge implantation relative to 4 week bridge implantation, whereas greater myelination, particularly by Schwann cells, was observed with 4 week bridge implantation. Taken together, the results of this dissertation show biomaterial bridges as a great tool to manipulate and investigate the spinal cord microenvironment to improve functional outcomes during acute and chronic stages of SCI. This work implies the focus of SCI treatments strategies should be reducing inflammation and enhancing myelination by oligodendrocytes for increased functional recovery. This work also implies resection of necrotic tissue in chronic SCI doesn’t negative impact functional recovery.PhDBiomedical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttps://deepblue.lib.umich.edu/bitstream/2027.42/153489/1/domsmi_1.pd