Quantification of angiogenesis due to basic fibroblast growth factor in a modified rabbit corneal model

The rabbit corneal angiogenesis assay was modified to allow quantification of the neovascular response induced by growth factors. Basic Fibroblast Growth Factor (bFGF) was used for this experiment. bFGF was diluted in phosphate buffer solution (PBS) in such a way that 2 microliter of solution contained concentrations ranging from 17 to 1,200 ng. Each drop was absorbed by a 1 x 2 mm particle of a dried 70% hydratable hydrogel and implanted into a mid-stroma corneal pocket, 2 mm from the limbus. Iodinated bFGF was used to measure growth factor diffusion in the cornea, which was found to be isotropic. Autoradiography showed that bFGF was stored in the cornea at both epithelial and endothelial level. Corneal neovascularization occurred on the second day after implantation and was maximal on the 7th day. At this time the neovascular surface was measured by planimetry on corneal photographs and compared with controls. This method allows precise definition of the dose of any substance to be implanted into the cornea and induces a rapid neovascular response, thus allowing quantitative evaluation of neovascularization within one week. A neovascular response was detectable for doses as low as 35 ng of bFGF and increased proportionally to the dose of bFGF implanted.