(85d) Lentiviral Gene Delivery From Hydrogel Filled PLG Scaffolds

Strategies to promote regeneration of lost or damaged tissue frequently employ polymeric scaffolds to provide mechanical support and promote cell infiltration that also deliver tissue inductive factors. Gene delivery from scaffolds has the potential to target one or more processes that limit regeneration by delivering DNA encoding for inductive proteins. Releasing DNA from biomaterials often results in transgene expression adjacent to the implant, yet scaffolds aimed at regenerating lost tissue should promote substantial expression within the scaffold. To promote localized and sustained expression, we have investigated a strategy based on filling the pores of the scaffold with a hydrogel, which provides a barrier to release. The porous scaffold thus provides mechanical support to the injury, whereas the hydrogel design mediates cell interactions and regulates gene delivery. A lentiviral vector encoding for luciferase as a reporter gene was combined with the hydrogels collagen, fibrin, and alginate, and loaded into the pores of PLG scaffolds. These three hydrogels were selected as they have different rates of degradation and cell interactions, which was expected to influence the rate of cell infiltration and vector release. Upon gelation, these scaffolds were transplanted in the epididymal fat pad of mice and expression was followed in vivo using a bioluminescence system. Expression persisted for up to 4 weeks post implantation. In the first week after implantation, transgene expression was observed localized at the implant and also in the spleen and thymus. After 1 week, expression was observed only in the implant. The identity of transduced cells was determined by delivering a vector encoding for enhanced green fluorescence protein (EGFP), followed by dual immunostaining. Macrophages (F4/80+) and dendritic cells (CD11c+) were identified as GFP+ at the scaffold and the spleen. These results demonstrate the ability to localize gene expression within the scaffold, which has numerous applications in cell transplantation and gene therapy.