Date of Award

5-2015

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Legacy Department

Bioengineering

Advisor

Ken Webb

Committee Member

Zhi Gao

Committee Member

Alexey Vertegel

Committee Member

Frank Alexis

Abstract

Over 50 million Americans are affected by ailments to the central nervous system (CNS) and it impacts the American economy over $400 billion a year. The number of people in the United States who have spinal cord injury (SCI) has been estimated to be approximately 276,000 persons as of 2014 with a range from 240,000 to 337,000 persons. The annual incidence of SCI, not including those who die at the scene of the accident is approximately 12,500 new cases each year. Due to the limited regenerative capacity of the adult CNS and lack of clinically effective therapies, these conditions commonly result in permanent functional deficits. SCI damages both ascending sensory and descending motor axonal pathways interrupting the transmission of synaptic signals between the brain and peripheral tissues. Although damaged axons attempt an initial regenerative response, this is rapidly aborted due to the presence of growth inhibitory molecules in CNS myelin and the glial scar and intrinsic limitations of adult CNS neuronal biochemistry such as the ability to maintain cAMP levels and upregulate the expression of `regeneration-associated genes'. On the other hand, TBI, stroke, and Parkinson's disease result in neuronal cell death. The CNS has limited capacity to replace lost neurons because the neurons themselves are terminally differentiated and post-mitotic. Although neural stem cells (NSCs) have been identified in specialized regions of the adult brain such as the sub-ventricular zone (SVZ) and the sub-granular zones (SGZ), their number is insufficient and the pathological environment inadequate to support an effective regenerative response. The end goal of this project is to develop a biomimetic scaffold using grooved fibers for neural regeneration. This goal was met with a two-pronged approach. In the first approach, grooved fibers immobilized with bioactive adhesive molecule were developed to topographically guide regenerating axons. In the second approach, grooved fiber staples were used as cell-laden microcarriers and integrated into a composite hydrogel which demonstrated its ability to serve as a platform for cell proliferation. This latter approach can be translated into an injectable in situ crosslinkable scaffold that can be used for neural stem cell (NSC) delivery with the prospect of stem cell differentiation into neurons to replenish cell loss. The first part of this research focused on immobilizing a bioactive 140 kDa fragment of L1 neural cell adhesion molecule on uniquely designed groovy capillary channel polymer (CCP) fibers. L1-CAM is an attractive candidate for growth of spared axonal growth cones upon injury. It mediates CNS maturation, by means of neurite outgrowth, adhesion, fasciculation, migration, survival, myelination, axon guidance, synaptic plasticity and regeneration after trauma. High levels of L1 are expressed by growing axons during development and after SCI and there is a positive correlation between their expression and axonal growth. CCP fibers with surface immobilized L1-CAM were demonstrated to guide growth of primary neurons in vitro. In the latter part of this research, a methodology to fabricate CCP fiber staples was developed and these were employed as cell-laden microcarriers. These microcarriers were then integrated into a composite hydrogel blend and demonstrated high cell proliferation in vitro compared to control gels. This composite system can be a promising platform for NSC delivery and differentiation into neurons.

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