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J. Yang, E. Duh, R. B. Caldwell, M. A. Behzadian; PEDF's Action Inhibiting VEGF-Induced Permeability in Retinal Endothelial Cells Involves Beta-Catenin Signaling Pathway and uPAR Gene Expression. Invest. Ophthalmol. Vis. Sci. 2007;48(13):1777. doi: https://doi.org/.
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To study the mechanism of pigment epithelium-derived factor (PEDF)’s inhibitory action on VEGF-induced vascular permeability. PEDF is found in many cell types including vascular endothelial cells. It has potent anti-permeability and anti-angiogenic activity, probably by blocking VEGF function. Decreased PEDF levels have been associated with microvascular hyper-permeability in retinopathy. Work in our laboratory indicates that, under diabetic conditions, PEDF levels remain higher and the blood-retinal barrier is preserved in uPAR knockout mice as compared to wild type mice. We have also shown previously that VEGF-induced endothelial-cell permeability is mediated by transcriptional activation of ß-catenin and activation of the uPA/uPAR system. Here we test the hypothesis that PEDF inhibits VEGF induced permeability by blocking ß-catenin signaling and inhibiting uPAR expression.
Bovine retinal endothelial (BRE) cells were used for permeability assay based on changes in trans-endothelial electrical resistance (TER) in the presence or absence of VEGF and PEDF. Confocal immunofluorescence microscopy, cell fractionation and Western blot analysis were used to evaluate PEDF effects on VEGF-induced redistribution of ß-catenin from the membrane into the cytosol and nucleus. Real-time PCR was used to quantify uPAR expression in VEGF- and PEDF-treated BRE cells.
TER assays indicated that pretreatment with PEDF blocks VEGF- induced paracellular permeability increases in BRE cells. This phenomenon seemed to be dose dependent in that high concentrations of PEDF enhanced the VEGF effect. Real-time PCR analysis showed that PEDF blocks the action of VEGF in increasing uPAR expression. Confocal imaging and cell fractionation studies showed that PEDF inhibits VEGF-induced cytosolic accumulation and nuclear translocation of ß-catenin.
PEDF blocks VEGF induced permeability by inhibiting ß-catenin nuclear translocation and preventing uPAR expression. Understanding the mechanism of PEDF's anti-permeability action at the intracellular signaling level is important for developing new PEDF-based strategies for treatment of retinopathy.
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