Vasoinhibins prevent retinal vasopermeability associated with diabetic retinopathy in rats via protein phosphatase 2A–dependent eNOS inactivation
Celina García, … , Gonzalo Martínez de la Escalera, Carmen Clapp
Celina García, … , Gonzalo Martínez de la Escalera, Carmen Clapp
Published May 22, 2008
Citation Information: J Clin Invest. 2008;118(6):2291-2300. https://doi.org/10.1172/JCI34508.
View: Text | PDF
Research Article Ophthalmology

Vasoinhibins prevent retinal vasopermeability associated with diabetic retinopathy in rats via protein phosphatase 2A–dependent eNOS inactivation

Abstract

Increased retinal vasopermeability contributes to diabetic retinopathy, the leading cause of blindness in working-age adults. Despite clinical progress, effective therapy remains a major need. Vasoinhibins, a family of peptides derived from the protein hormone prolactin (and inclusive of the 16-kDa fragment of prolactin), antagonize the proangiogenic effects of VEGF, a primary mediator of retinal vasopermeability. Here, we demonstrate what we believe to be a novel function of vasoinhibins as inhibitors of the increased retinal vasopermeability associated with diabetic retinopathy. Vasoinhibins inhibited VEGF-induced vasopermeability in bovine aortic and rat retinal capillary endothelial cells in vitro. In vivo, vasoinhibins blocked retinal vasopermeability in diabetic rats and in response to intravitreous injection of VEGF or of vitreous from patients with diabetic retinopathy. Inhibition by vasoinhibins was similar to that achieved following immunodepletion of VEGF from human diabetic retinopathy vitreous or blockage of NO synthesis, suggesting that vasoinhibins inhibit VEGF-induced NOS activation. We further showed that vasoinhibins activate protein phosphatase 2A (PP2A), leading to eNOS dephosphorylation at Ser1179 and, thereby, eNOS inactivation. Moreover, intravitreous injection of okadaic acid, a PP2A inhibitor, blocked the vasoinhibin effect on endothelial cell permeability and retinal vasopermeability. These results suggest that vasoinhibins have the potential to be developed as new therapeutic agents to control the excessive retinal vasopermeability observed in diabetic retinopathy and other vasoproliferative retinopathies.

Authors

Celina García, Jorge Aranda, Edith Arnold, Stéphanie Thébault, Yazmín Macotela, Fernando López-Casillas, Valentín Mendoza, Hugo Quiroz-Mercado, Hebert Luis Hernández-Montiel, Sue-Hwa Lin, Gonzalo Martínez de la Escalera, Carmen Clapp

×

Figure 3

Vasoinhibins inhibit VEGF-induced eNOS phosphorylation and eNOS activity via PP2A activation.

Options: View larger image (or click on image) Download as PowerPoint
Vasoinhibins inhibit VEGF-induced eNOS phosphorylation and eNOS activity...
(A) Line diagrams depicting the different protocols used in experiments represented in B, C, and D. (B) Western blot analysis of eNOS-Ser1179 phosphorylation in BAECs preincubated or not with 50 nM OA for 10 minutes, then with or without 20 nM vasoinhibins for 1 hour, followed or not by a treatment with 5 nM VEGF for 5 minutes as indicated. Total eNOS served as loading control. (C) eNOS activity was determined by the [3H]l-citrulline assay in BAECs incubated or not with 5 nM VEGF, alone or combined with 20 nM vasoinhibins in the presence or absence of 50 nM OA for 1 hour. Values are mean ± SEM of 5 independent experiments. *P < 0.05 versus no treatment; **P < 0.05 versus VEGF; #P < 0.05 versus VEGF and vasoinhibins. (D) Vasopermeability was measured by HRP leakage in BAEC monolayers untreated (ctl) or treated for 48 hours with 50 nM OA, 5 nM VEGF, or the combination of VEGF and 20 nM vasoinhibins or VEGF, vasoinhibins, and OA. Values are mean ± SEM of 3 independent experiments. *P < 0.05 versus no treatment; **P < 0.05 versus VEGF; #P < 0.05 versus VEGF and vasoinhibins.