(4br) Fundamentals of Gene Delivery From Tissue Engineering Scaffolds

My research focus has been the development of biomaterial scaffolds capable of localized gene delivery that can enhance the native ability of the body to regenerate or heal wounds. Porous scaffolds are widely used to mechanically support the injury, promote cell infiltration, and deliver tissue inductive factors. To locally deliver these inductive factors, an approach with the unique potential to target one or more processes is by delivering gene therapy vectors encoding for the factors. I have performed mechanistic studies of gene delivery from biomaterials, and applied these delivery systems to regenerate nerves following spinal cord injury. For the mechanistic studies, I investigated two approaches to improve the localization and duration of transgene expression: i) cationic polymers to modify the scaffolds to reversibly bind the vectors, and ii) hydrogels to fill the pores of the scaffold. For the first approach, I used scaffolds made from PLG microspheres modified with cationic polymers, such as PEI, PLL, and dopamine to reversibly associate with the DNA and delay release from the scaffold, which should increase the local concentration of the vector for enhanced delivery. The scaffolds modified with polydopamine enhanced plasmid incorporation and at the same time extended release. However, extended release did not impact the duration of transgene expression in vivo, suggesting that the initial response of the host to the implant may dictate the extent of transgene expression. In the second approach, collagen, fibrin, and alginate hydrogels were deposited within the pores of the scaffolds. These three hydrogels were selected as they have different rates of degradation and cell interactions, which were expected to influence the rate of cell infiltration and vector release. Both plasmid and lentiviral vectors were loaded into the hydrogels. The plasmid produced short-term expression, whereas expression following lentivirus delivery persisted for up to 4 weeks post implantation. These results indicate that expression can be localized and sustained for extended periods of time.

We subsequently investigated gene delivery from porous scaffolds as a means to improve regeneration following spinal cord injury. Lentivirus encoding for the neurotrophic factors NT3 and BDNF were complexed with nanoparticles, loaded into scaffolds, and implanted in a rat spinal cord hemisection spinal cord model. The spinal cords were retrieved at 1, 2 and 4 weeks post implantation and stained for neurofilament (NF200). Lentiviral delivered significantly increased neurofilament growth through the injury site. Gene delivery from scaffolds is a viable approach to enhance spinal cord regeneration, and does not require cell transplantation common to other approaches. Overall, these studies demonstrate that gene delivery from biomaterials can enhance regeneration and has potential for cell transplantation and gene therapy applications. I see opportunities to improve gene delivery applications with control release systems, targeting, and actuated release. The use of magnetic particles can be used to enhance delivery, as well as to study cell response to mechanical stimuli, which can alter regeneration. I have experience in magnetic targeting which will allow me to utilize magnetic systems applied to tissue engineering and gene delivery for future research.