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A. Gvritishvili, Y. Liu, J. Tombran-Tink; PEDF78-121 Limits Oxidative Stress in RPE Cells Through Akt and Erk Signaling and Reduces Stress Promoted Expression of VEGFR1. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1442.
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© ARVO (1962-2015); The Authors (2016-present)
PEDF treatment can reduce both retinal cell death and neovascularization. The neurotrophic activity of the protein is localized to an N terminal fragment corresponding to amino acid residues 78-121. In this study we examined the effects of this peptide on oxidative stress in RPE cells.
PEDF78-121 peptide was cloned into pET32a expressed in E.coli, and peptide purified using Ni-NTA chromatography. The effects of purified PEDF78-121 peptide were examined in ARPE19 cultures challenged with 640µM H2O2. Cells were treated for 2h with H2O2 followed by treatment with various concentration of PEDF78-121 for 24h. The protective effects of PEDF78-121 on H2O2 toxicity were assessed using LDH assay and PI staining. Modulations in VEGF, VEGF receptors, and various signaling pathways were also determined.
PEDF78-121 protects RPE cells from hydrogen peroxide toxicity at concentrations as low as 25 ng/ml. Approximately 30% decrease in cell death is seen in cultures after treatment with the peptide. This protective effect is mediated through the Akt and Erk signaling pathways. H202 decreased the phosphorylation of both signal molecules after 2h, a condition immediately reversed by PEDF78-121. Specificity of PEDF78-121 actions was shown with the pharmacological inhibitors Akt and MEK1/2 which blocked Akt and Erk1/2 activation, respectively by PEDF78-121. Inhibition of Akt and Erk signaling abrogated the protective effects of PEDF78-121 on RPE cells. We also show that oxidative stress increases the expression of VEGFR1 in RPE cells and that PEDF78-121 reduced this expression by 3.4 fold, an effect that was abolished when Akt and Erk signaling pathways were blocked.
Oxidative stress can modulate Akt and Erk signaling and induce expression of VEGR1 in RPE cells. PEDF78-121 is an effective pharmacological tool that reduces oxidative stress induced death in RPE cells and may limit the actions of these cells in angiogenic events in the retina.
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