| Spinal cord injury (SCI) is a devastating condition that affects more than 10,000 people in the US each year. Today, even though most SCI patients benefited from the rapid development in treatment and care and live much longer, currently there is no cure for this condition. In the past two decades, developments in neurotransplantation and gene transfer techniques have introduced a spectrum of promising strategies for treatment of SCI, including in vivo and ex vivo gene therapy. In the in vivo approach, vectors encoding therapeutic genes are used directly to transduce host cells to induce expression of the therapeutic products. In the ex vivo approach, cells are genetically modified in vitro and grafted into the host as a vehicle for delivery of the therapeutic agents. Gene therapy encompasses three dimensions: the therapeutic genes, the vectors and the donor cells. Examples of valuable therapeutic agents include the neurotrophic/neurotropic factors and their receptors; anti-apoptotic factors, growth associated proteins, cell adhesion molecules and extracellular matrix molecules. A variety of vectors are designed for in vivo and ex vivo gene delivery, such as herpes simplex virus, adenovius, retrovirus, adenoviral-associated virus, lentivirus and some non-viral vectors. Several types of donor cells have been studied, including primary cells, immortalized cell lines and stem cells/cell lines. The goal of this thesis is to develop strategies of in vivo and ex vivo intraspinal gene delivery and to apply them to experimental models of SCI. All dimensions of gene therapy were explored. A variety of viral vectors (adenoviral and retroviral vectors) and donor cells (primary fibroblasts and immortalized neural stem cells) were employed to deliver therapeutic genes (NT-3 and BDNF) and reporter genes (lacZ and GFP) into intact or injured spinal cord. In chapters 3 and 4, recombinant adenoviral vectors encoding reporter genes were used to demonstrate the feasibility of in vivo and ex vivo intraspinal gene delivery. In chapter 5, an immortalized neural stem cell line was genetically modified by a retroviral vector encoding the NT-3 gene and xenografted into adult rat spinal cord to demonstrate the feasibility of ex vivo intraspinal gene delivery with neural stem cells. Finally in chapters 6 and 7, primary fibroblasts engineered to produce BDNF were grafted into a cervical spinal cord hemisection cavity to demonstrate their effects on promoting rubrospinal axon regeneration and functional recovery and preventing axotomy induced retrograde neuron death and atrophy in the Red Nucleus. Results presented in this thesis provided direct evidence that gene therapy may eventually prove effective for treatment of human SCI. |
