Optimizing alginate microbead delivery system for release of angiogenic protein for neovascularization.

摘要

The ability to stimulate the formation of new blood vessels (neovascularization) is essential for the treatment of ischemic tissues and key for the success of many tissue engineering applications. Using growth factors for the therapeutic stimulation of neovascularization has been investigated extensively, but the inability to control their temporal delivery may limit clinical success. To improve the administration of angiogenic factors, studies presented in this thesis focus on devising methods for the formulation of alginate microbeads for the encapsulation and release of protein to stimulate angiogenesis.;The first aim in addressing this objective concentrated on generating an understanding of the relationship between synthesis conditions and protein release kinetics in order to effectively design a system for controlled release. These studies indicate that alginate chemistry and synthesis conditions could be varied to control physical characteristics of alginate microbeads. In aim 2 and aim 3 the biological activity of FGF-1 released from these beads was determined in vitro and in vivo respectively. Cell assays confirmed the ability of released FGF-1 to stimulate proliferation of endothelial cells. A 3D co-culture model demonstrated protein loaded microbeads increased sprout formation significantly in cell aggregates compared to bolus of FGF-1 and empty beads. Protein loaded alginate beads implanted in a murine model of vascularized adipose tissue formation had a significant increase in vascular number density at 1 and 6 weeks than the group with bolus administration of FGF-1 and the empty bead group. Staining for smooth muscle actin showed that over 48% of vessels had associated mural cells. No differences in adipose formation were observed between any groups at any time points. Results in these studies suggest that sustained release of FGF-1 increases the duration of the vascular response in contrast to a bolus injection of FGF-1 however promoting angiogenesis alone was not sufficient for stimulating adipogenesis. In addition, lower total dose of FGF-1 can stimulate increased vascular density when delivered from alginate microbeads. These results presented in this dissertation provide a compelling impetus for experimental pursuit of FGF-1 loaded alginate microbeads for use in therapeutic stimulation of neovascularization in regenerative medicine applications.