Thesis (Ph. D.)--Biomedical Engineering, Georgia Institute of Technology, 2008. / Committee Chair: LaPlaca, Michelle; Committee Member: Bellamkonda, Ravi; Committee Member: Garcia, Andres; Committee Member: Hochman, Shawn; Committee Member: Wang, Yadong. Part of the SMARTech Electronic Thesis and Dissertation Collection.

[Truncated abstract] Microenvironmental changes associated with apoptotic neural degeneration may instruct a proportion of newly transplanted donor cells to differentiate towards the fate of the deteriorating host cellular phenotype. In the work described in this thesis, this hypothesis was tested by inducing apoptotic retinal ganglion cell (RGC) death in neonatal and adult rats and mice, and then examining whether intravitreally grafted cells from a range of sources of donor neural tissue became incorporated into these selectively depleted retinae. Donor tissues were: a postnatal murine cerebellar-derived immortalised neural precursor cell line (C17.2); an adult rat hippocampal-derived clonal stem-like line (HCN/GFP); mouse embryonic day 14 (E14) primary dissociated retinal cells (Gt[ROSA]26); and adult mouse ciliary pigmented margin-derived primary neurospheres (Gt[ROSA]26). In neonates, rapid RGC death was induced by removal of the contralateral superior colliculus (SC), and in adults, delayed RGC death was induced by unilateral optic nerve (ON) transection. Some adult hosts received ON transection coupled with an autologous peripheral nerve (PN) graft. Donor cells were injected intravitreally 6-48 h after SC ablation (neonates) or 0, 5, 7 or 14 days after ON injury (adults). Cells were also injected into non-RGC depleted neonatal and adult retinae. At 4 or 8 weeks, transplanted cells were identified, quantified and their differentiation fate within host retinae was assessed. Transplanted male C17.2 cells were identified in host retinae using a Y-chromosome marker and in situ hybridisation, or by their expression of the lacZ reporter gene product Escherichia coli beta-galactosidase (beta-gal) using Xgal histochemistry or a beta-gal antibody. No C17.2 cells were identified in axotomised adult-injected eyes undergoing delayed RGC apoptosis (n = 16). Donor cells were, however, stably integrated within the retina in 29% (15/55) of mice that received C17.2 cell injections 24 h after neonatal SC ablation; 6-31% of surviving cells were found in the RGC layer (GCL). These NSC-like cells were also present in intact retinae, but on average there were fewer cells in GCL. In SC-ablated mice, most grafted cells did not express retinal-specific markers, although occasional donor cells in the GCL were immunopositive for beta-III tubulin (TUJ1), a protein highly iii expressed by, but not specific to, developing RGCs. Targeted rapid RGC depletion thus increased C17.2 cell incorporation into the GCL, but grafted C17.2 cells did not appear to differentiate into an RGC phenotype.

Traumatic brain injury (TBI) results in over 50,000 deaths and 80,000 disabilities each year. Current treatment strategies aim to alleviate acute disturbances, but are not able to address the chronic disorders associated with TBI. Neural transplantation is one potential treatment that will provide multifaceted sustained therapy to degenerating injured tissue. Transplantation of multipotent neural stem cells (NSCs) has been shown to enhance functional recovery in TBI models; however, poor cell survival and integration with host tissue potentially restrict the efficacy of such transplants. This limitation may be due to the absence of inherent NSC pro-survival cues (e.g., cell-ECM interactions). Furthermore, the neural injury environment presents cell death factors to transplanted NSCs. It is hypothesized that a 3-D scaffold presenting specific CNS adhesive moieties will enhance donor cell survival and promote differentiation and migration. This project encompassed material development and in vitro characterization. Results highlighted the importance of ligand tethering chemistry and density and also the mechanical integrity of cell scaffold systems. Furthermore, the developed scaffold provides a controlled microenvironment to assess the influence of LN on NSC survival, migration, and differentiation. Lastly, co-delivering NSC with the MC-LN tissue engineered scaffold into a mechanically injured neural co-culture test-bed or in vivo TBI model confirmed the importance of ECM cues for NSC survival and migration, respectively.