| PLGA microspheres were developed as an injectable depot for sustained, local delivery of vascular endothelial growth factor (VEGF) in infarcted myocardium. Cryogenic solid entrapment, solvent extraction microsphere fabrication incorporated 95+/-6% of the protein and maintained VEGF bioactivity. In vitro release of VEGF165 was hindered, relative to BSA or VEGF121, as a result of electrostatic interactions between the protein and polymer. All proteins released over a 5-8 week period, but a 4-5 week lag phase was observed for VEGF165 release. Heparin and protamine were evaluated as release-modifying excipients, but neither was overly efficacious.; The fate of particulate injected into ischemic myocardium was quantified using neutron-activated microspheres. On average, hearts retained 40-60% of the injected particulate, but retention was variable (5-100%). Leakage from the injection site and clearance through venous vessels were the primary modes of loss. Pluronic F127 and Matrigel reduced immediate leakage, relative to saline injections, but microsphere retention was unimproved. Data suggested that leakage occurred over a prolonged period with the gelling vehicles.; Injection of VEGF165-loaded PLGA microspheres (∼9mug VEGF) into rat myocardial infarcts induced a small, but statistically significant increase in vascularization, confirming VEGF bioactivity. A 93+/-3% reduction in ELISA-detectable VEGF was observed within 7 days of microsphere injection into infarcts. This clearance was markedly faster than predicted by in vitro studies. Microspheres elicited a mild foreign body response and disappeared within 2-3 weeks after injection, based on histology. However, the quantity of microspheres observed at 1 and 7 days was comparable, suggesting that protein degradation may be the primary cause of rapid VEGF clearance.; To our knowledge, this work represents the first investigation of PLGA microspheres for sustained protein delivery within infarcted myocardium. Incomplete retention, enhanced PLGA erosion, and accelerated VEGF degradation make this a challenging environment. Formulations delivering more VEGF over a longer duration would likely provide greater enhancement of infarct vascularization. This work highlights the critical importance of in vivo experiments and the challenges that limited protein availability impose on development of therapeutic microsphere formulations. Nevertheless, PLGA microspheres are a promising delivery system for angiogenic proteins. |
