Access Type

Open Access Dissertation

Date of Award

January 2020

Degree Type

Dissertation

Degree Name

Ph.D.

Department

Biomedical Engineering

First Advisor

Harini Sundararaghavan

Abstract

Over 17,000 people per year in the United States sustain a spinal cord injury (SCI) for which there is no gold standard of care and life-long complications. SCI is a complex wound environment with various growth factors (GFs), cellular activity, and scar formation at various timepoints. For example, basic fibroblast growth factor (bFGF) is released immediately to protect local support cells, namely oligodendrocytes (OLs). Brain derived neurotrophic factor (BDNF) has peak expression at 2-3 days in mice and 5-7 days in humans, and aids axonal growth across the injury site. Incidentally this is around the same time as M1 macrophage invasion as part of the immune response after SCI.

Growth factor (GF) delivery is a common strategy in tissue engineering, however unbound GFs degrade quickly, and higher dosages are required to gain a therapeutic effect. We hypothesize that GFs delivered and maintained at times that match the native SCI response will increase the amount of neuronal reconnection and support cell presence as compared to one or none of the individual GFs released at the same times.

This thesis develops a spatiotemporal GF delivery system that provides bFGF immediately and BDNF delayed, with GF bioactivity maintained via heparin-hyaluronic acid nanofibers. Spatiotemporal release was controlled via hydrolytic degradation of poly(lactic-co-glycolic) acid microspheres (PLGA-MS) which release bFGF immediately, and enzymatic degradation of gelatin microspheres (GMS) which release BDNF in the presence of collagenase, which is secreted by M1 macrophages. Both PLGA-MS and GMS were electrospun with heparin-hyaluronic acid (HepHA) nanofibers, from which heparin actively sequesters GFs for increased bioactivity durations. This combined spatiotemporal system (STS) was then evaluated in a biomaterial-focused organotypic spinal cord (OSC) injury model for its effect on neurite reconnection as measured by percentage recovery of injury gap and OL number per gap and density (per mm^2).

Heparin-hyaluronic acid nanofibers were confirmed to sequester bioactive GFs in culture with L929 fibroblasts and dissociated chick neurons. GMS were confirmed to only release GFs when degraded and GMS electrospun into hyaluronic acid (HA) nanofibers were shown to have a positive effect on chick neurons, particularly with added M1 macrophage conditioned media. Within our OSC model, STS was compared with HA and HepHA, HepHA PLGA-MS+bFGF, and HepHA GMS+BDNF. STS had the highest percent gap recovery and OL number compared to all other groups, however HepHA had the highest OL density. Though there was no statistical difference between groups, the STS is a promising biomaterial for SCI, which warrants further study.

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