Abstract
Purpose: :
VEGF-A is a key regulatory component in vascular development. The VEGF165 isoform of A is thought to be the predominant form for stimulating migration and proliferation of endothelial cells and angioblasts. An anti-angiogenic form of VEGF-A has been isolated, VEGF165b, which is generated by alternative splicing. This study investigates the localization of VEGF165 and VEGF165b during development of vasculatures in embryonic and fetal human eye.
Methods: :
Immunohistochemistry was performed on cryosections from 7 to 21 weeks gestation (WG) human eyes using a rabbit antibody against VEGF165 (Thermofisher) and a monoclonal antibody against VEGF165b (Abcam) and antibodies against CD31 and CXCR4 to label endothelial cells and progenitors respectively.
Results: :
The tunica vasculosa lentis (TVL) had moderate VEGF165 immunoreactivity at 7 WG, and very little VEGF165b. Both forms were elevated at 12 WG but VEGF165 was cytoplasmic and VEGF165b was nuclear in endothelial cells (EC). Both forms of VEGF-A were present diffusely throughout the retinal neuroblastic layer (NBL) at 7 WG. At 12 WG, VEGF165 was present diffusely through inner retina and inner NBL and VEGF165 and VEGF165b were present in most CXCR4+ progenitors in the inner retina, which includes angioblasts. By 21 WG, VEGF165 was present in nerve fibers and VEGF165b in inner Muller cell processes in central retina. In choroid, VEGF165 was present in forming choriocapillaris (CC) and RPE at 7 WG while VEGF165b was present in CC and mesenchymal progenitors in choroid and sclera. By 12 WG, both were present predominantly in developing CC but VEGF165 was present in the basal portion of RPE as well. At 21 WG, both forms were lower in choroidal blood vessels than previously but VEGF165b was also present in the nuclei of RPE.
Conclusions: :
In TVL at 12 WG, VEGF165 was cytoplasmic and VEGF165b was nuclear in endothelial cells. VEGF165 and VEGF165b had distinctly different localizations in retina at 12 and 21 WG. Most striking was the succinct nuclear association of VEGF165b within CXCR4+ progenitors in retina, some of which are angioblasts, and mesenchymal precursors in choroid. The localization of VEGF165 in nerve fibers and VEGF165b in Muller cells at 21 WG, suggests that both neurons and glia contribute to the control of retinal vasculogenesis.
Keywords: vascular endothelial growth factor • retinal development • choroid