Analysis of Tissue Sparing and Circuit Regeneration in Spinal Cord Injuries Treated with Biomaterials and Gene Therapy
Chen, Jessica
2020
Abstract
Each year, the U.S. sees nearly 17,700 new cases of spinal cord injuries (SCIs). Despite intense rehabilitation, patients with SCIs most often suffer lifelong physical consequences and substantial increases in medical expenses per individual. While immobilization and surgery can be used for immediate stabilization of the injury, no clinical methods exist to address the subsequent inflammation and lack of tissue regeneration that further contribute to the motor and sensory deficits seen after an SCI. This dissertation aimed to understand how biomaterials and gene therapy treatment affect SCIs. In a mouse SCI model, where a left C5 hemisection results in loss of function of the left arm, a poly(lactide-co-glycolide) (PLG) scaffold or “bridge” can be implanted in place of the resected tissues. The bridge can be loaded with lentivirus for local delivery of gene therapy that can aid in control of the post-SCI microenvironment. Using lentiviral interleukin-10 (IL10), we found that IL10 animals significantly outperform animals that received a control lentivirus on a ladder beam test at 2- and 12- weeks post-injury (wpi). Closer examination of components of the forelimb motor circuitry suggest IL10 animals had increased sparing of lower motor neurons and neuromuscular junctions. Electrophysiological studies at 2 wpi showed that control injured animals had electromyogram recordings that were significantly dampened when compared to IL10 and control uninjured animals, thereby confirming that the motor circuitry remained more intact with IL10 treatment. These results, which were consistent in both male and female mice, are the first to show that IL10 spares motor circuitry directly responsible for enhanced muscle function. We then tested a combination therapy of lentiviral IL10 and brain-derived neurotrophic factor (BDNF), followed by examination of tissue sparing and regeneration. At 2 wpi, histological and electrophysiological analyses show that the tissue sparing effects of IL10 alone are only slightly enhanced by the addition of BDNF. By 12 wpi, most innervation differences among the treatment conditions disappeared, though electrophysiological examination suggests that IL10 may prevent some of the injury-associated shifts in muscle composition that result in increased fatigability in control injured animals. Within the spinal cord, we found that IL10 alone and IL10+BDNF cause a similar increase in axon growth across the injury site. 3D imaging using Clear Lipid-exchanged Acrylamide-hybridized Rigid Imaging / Immunostaining / in situ-hybridization-compatible Tissue Hydrogel (CLARITY) shows these axons do completely traverse the injury site, while electrophysiological studies suggest these axons are able to carry action potentials. These results are the first to show that regenerated axons can be electrophysiologically active. Taken together, these studies suggest early immunomodulation can have long-lasting benefits through tissue sparing, and that regenerated axons have the potential to transduce signals across an injury site. These findings provide novel insights into how the pathophysiology following an SCI can be altered using biomaterials and gene therapy. Future studies will involve identifying the synaptic targets of regenerated axons and determining how the formation of new circuits can influence motor function.Subjects
Tissue Sparing and Regeneration After A Spinal Cord Injury Biomaterial Implantation and Gene Therapy Threatment
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