PlGF silencing combined with PEDF overexpression: Modeling RPE secretion as potential therapy for retinal neovascularization

Ocular neovascularization is a defining feature of several blinding diseases. We have previously described the effectiveness of long-term pigment epithelium-derived factor (PEDF) expression in the retina of diabetic mice in ameliorating some diabetic retinopathy hallmarks. In this study, we aimed to investigate if the antiangiogenic potential of PEDF overexpression was enhanced in combination with placental growth factor (PlGF) silencing. Human RPE cells were transfected with a self-replicating episomal vector (pEPito) for PEDF overexpression and/or a siRNA targeting PlGF gene. Conditioned media from PEDF overexpression, from PlGF inhibition and from their combination thereof were used to culture human umbilical vein endothelial cells, and their proliferation rate, migration capacity, apoptosis and ability to form tube-like structures were analyzed in vitro. We here demonstrate that pEPito-driven PEDF overexpression in combination with PlGF silencing in RPE cells does not affect their viability and results in an enhanced antiangiogenic activity in vitro. We observed a significant decrease in the migration and proliferation of endothelial cells, and an increase in apoptosis induction as well as a significant inhibitory effect on tube formation. Our findings demonstrate that simultaneous PEDF overexpression and PlGF silencing strongly impairs angiogenesis compared with the single approaches, providing a rationale for combining these therapies as a new treatment for retinal neovascularization.