Oscillation of angiogenesis with vascular dropout in diabetic retinopathy by VESsel GENeration analysis (VESGEN).

Diabetic retinopathy is the leading cause of visual loss among working-aged adults in the United States.1,2 The diagnosis and management of diabetic retinopathy are based on grading of features obtained from clinical examination, as was performed in the Early Treatment Diabetic Retinopathy Study (ETDRS).3–5 Although progression of diabetic retinopathy results from adverse vascular remodeling that includes vascular dropout, ischemia, and finally neovascularization, current diagnosis relies on the grading of secondary vascular effects, such as microaneurysms, leakage, and exudates. It is challenging to directly evaluate changes in retinal blood vessels because of the morphologic complexity of the overlapping, highly branching arterial and venous trees within the human retina. The goal of this study was to investigate changes in the branching patterns of the arterial and venous trees during the progression of diabetic retinopathy. To study vascular remodeling directly, we used VESsel GENeration Analysis (VESGEN) software6–11 to map and quantify arterial and venous trees extracted from clinical images obtained by fluorescein angiography (FA). The VESGEN computer program analyzes major vascular branching parameters in a binary (black/white) image of a vascular tree, vascular network, or tree-network composite. Mapping and quantification by VESGEN analysis automatically segments vessels within a tree into branching generations (G1, G2, … Gx) according to coordinate change in vessel diameter and branching. A recent review11 of VESGEN applications mapped and quantified vascular trees and networks in the human retina, transgenic mouse retina, and chorioallantoic membrane (CAM), an avian model of angiogenesis and lymphangiogenesis. Coronary vessel development was analyzed as an immature vasculogenic network, a transitional network-tree composite, and a mature tapering vascular tree. To develop VESGEN mapping capabilities, proangiogenesis and antiangiogenesis factors were first studied in the CAM. By VESGEN analysis, basic fibroblast growth factor (bFGF) stimulated specifically the robust growth of many small vessels.6 Vascular endothelial growth factor (VEGF)-A, however, had a more complex effect.9 At low concentrations, VEGF stimulated the growth of new small vessels in a manner resembling stimulation by bFGF. At high concentrations, regulation by VEGF resulted in a more pathologic regulatory phenotype, in which the diameter of larger vessels was significantly dilated (accompanied by vascular leakage). Inhibition of angiogenesis by transforming growth factor (TGF) β-1 retained a normal vascular morphology,7 whereas inhibition by angiostatin rendered the vascular tree highly abnormal and irregular.12 The steroid drug triamcinolone acetonide (TA) also inhibited the growth of small vessels and, furthermore, thinned the diameters of all vessels throughout the vascular tree except those of the smallest vessels.10 We have previously quantified, with the use of fractal analysis, vascular morphology in the retinas of patients with normal eyes and those with mild nonproliferative diabetic retinopathy (NPDR).13 That study showed that compared with the normal retina, the combined density of arteries and veins in the NPDR retina decreased in the macula but was unchanged in peripheral regions. In our present study of progressive vascular remodeling during diabetic retinopathy, VESGEN software mapped and quantified branching characteristics of separated arterial and venous trees to reveal generation-specific changes of oscillating vessel density.