Stimulation of in vivo angiogenesis by cytokine-loaded hyaluronic acid hydrogel implants and potential gene expression mechanisms for new vessel growth

As a first step to developing cytokine-impregnated hydrogels capable of promoting desired physiologic responses, we have tested in vivo angiogenesis in a mouse model. Hyaluronic acid (HA) hydrogel samples pre-loaded with vascular endothelial growth factor (VEGF) or basic fibroblast growth factor (bFGF) were surgically implanted in an ear pocket in the mouse. The implants were then retrieved at 7 and 14 days post surgery, and the number of ingrowing microvessels evaluated by direct counting. As compared to contralateral controls, we show that released degrading hyaluronan alone is itself an effective stimulus for new vessel growth. Pre-loading of the gel with VEGF, however, dramatically augments the rate of new microvessel growth, resulting in a 2.3-fold increase in neovascularization over the sum of the rates produced by HA and VEGF individually. In contrast, although bFGF alone greatly augments vessel growth, no such potentiation effect was observed between bFGF and HA. We present physiologic data demonstrating vessel growth and correlate those data with gene expression changes measured by microarray analysis. The results show that HA-cytokine combinations can be designed to optimize angiogenesis, and suggest potential mechanisms by which angiogenic vessel proliferation may occur.