Advisory Committee Chair
Candace L Floyd
Advisory Committee Members
Date of Award
Degree Name by School
Master of Science in Biomedical Engineering (MSBME) School of Engineering
In chronic spinal cord injury (SCI), a glial encapsulated cyst develops at the injury site and acts as a physical and biochemical barrier to regeneration. Discussion has suggested that filling the cyst with a growth promoting scaffold is a promising approach to cure chronic SCI, though the research to date lacks a systematic approach. Dorsal root ganglia (DRG) explants are an extremely common tissue culture model of neural regeneration employed in this field, however there is no uniform method for objectively quantifying substrate biocompatibility and neurite outgrowth. Therefore, we aimed to develop a novel DRG growth assessment methodology and to demonstrate its efficacy for judging neurite outgrowth on electrospun polymer scaffolds. This study developed an assessment system for judging axonal outgrowth from DRG explants. The methodology involved assessment by three separate descriptive tests: 1) basic biocompatibility demonstrated by presence of neurite growth, 2) assessment of growth pattern characteristics using a three category Likert scale, and 3) pixel quantification of neurites from fluorescent micrographs. Having sufficiently refined these tests to give consistent scores under control conditions on laminin, we demonstrated the utility of the methodology in an experimental setting on various substrates. We electrospun and mechanically characterized three polymer scaffolds of polydioxanone (PDO), a novel poly(ester urethane urea) (PHH), and a 1:1 PDO/PHH blend, with random and aligned fiber configurations of each to compare the effects of polymer composition and fiber orientation on neurite outgrowth. We found that the size scale of our polymers was comparable to sprouting neurites and the modulus of all materials exceeded that of native tissue, and thus we posit that these substrates are physically suitable for supporting and directing nerve regrowth. We saw decreased growth scores between the laminin controls and all experimental substrates except the aligned PDO which also demonstrated excellent biocompatibility scores. The findings from all three growth tests indicate that PDO is highly biocompatible, and increasing PDO fiber alignment directs nerve growth along the direction of fiber alignment. We concluded that our in vitro assessment methods were inter-reliable and dependable measures of scaffold biocompatibility, making them an extremely effective and user-friendly tool for assessing substrate effects on neurite culture outgrowth. We also concluded that electrospun PDO is a promising material for development into a three dimensional SCI lesion filling scaffold, and we are prepared to perform direct in vitro comparisons between PDO and other substrates in preparation for in vivo studies.
Kezar, Sarah Margaret, "Development of an Axonal Outgrowth Assessment Methodology for Dorsal Root Ganglia Explants and Demonstration of Method Efficacy on Electrospun Polymer Scaffolds" (2014). All ETDs from UAB. 2132.