April 2014
Volume 55, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2014
PLVAP Modulates Angiogenesis By Tuning VEGF Signaling In Endothelial Cells
Author Affiliations & Notes
  • Joanna Wisniewska-Kruk
    Department of Ophthalmology, Academic Medical Center, Amsterdam, Netherlands
    Department of Cell Biology and Histology, Academic Medical Center, Amsterdam, Netherlands
  • Ingeborg Klaassen
    Department of Ophthalmology, Academic Medical Center, Amsterdam, Netherlands
    Department of Cell Biology and Histology, Academic Medical Center, Amsterdam, Netherlands
  • Ilse M Vogels
    Department of Cell Biology and Histology, Academic Medical Center, Amsterdam, Netherlands
  • Cornelis J Van Noorden
    Department of Cell Biology and Histology, Academic Medical Center, Amsterdam, Netherlands
  • Reinier O Schlingemann
    Department of Ophthalmology, Academic Medical Center, Amsterdam, Netherlands
    Department of Clinical and Molecular Ophthalmogenetics, Netherlands Institute for Neuroscience, Royal Netherlands Academy of Arts and Science (KNAW), Amsterdam, Netherlands
  • Footnotes
    Commercial Relationships Joanna Wisniewska-Kruk, None; Ingeborg Klaassen, None; Ilse Vogels, None; Cornelis Van Noorden, None; Reinier Schlingemann, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science April 2014, Vol.55, 2241. doi:
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      Joanna Wisniewska-Kruk, Ingeborg Klaassen, Ilse M Vogels, Cornelis J Van Noorden, Reinier O Schlingemann, ; PLVAP Modulates Angiogenesis By Tuning VEGF Signaling In Endothelial Cells. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2241.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: Plasmalemma vesicle associated protein (PLVAP) is an endothelial cell-specific protein, detected by the PAL-E antibody that has been used for decades as endothelial marker. PLVAP is expressed in a large part of the vascular system, but is absent from blood vessels with an intact blood-retinal barrier. However, in pathological conditions associated with neovascularization in the central nervous system, as occurs in diabetic retinopathy, PLVAP expression significantly increases. In this study, we investigated the role of PLVAP in angiogenesis and the potential molecular mechanisms that underlie its pro-angiogenic activity.

Methods: To elucidate the role of PLVAP in new blood vessel formation in vivo, we used the oxygen-induced retinopathy mouse model and injected siRNA intraocularly. Five days after injection, retinas were flat-mounted and the vasculature was analyzed. Furthermore, the possible pro-angiogenic function of PLVAP was investigated in in vitro models, including the aortic ring assay, the endothelial spheroid-based assay and migration assays. The silencing efficiency of siRNA and lentiviral delivered shRNA in all experiments was confirmed on mRNA and protein levels. The expression of VEGF receptors and its co-receptors after Plvap silencing in endothelial cells was investigated using western blot, immunohistochemistry and flow cytometry analysis.

Results: Silencing of Plvap in the oxygen-induced retinopathy model resulted in decreased vascular and neovascular areas, and increased avascular areas in the retinas. Plvap inhibition in cultured endothelial cells exhibited decreased sprout formation, reduced numbers of endothelial tip cells and diminished cell migration. However, no effect on cell viability or cell proliferation was observed after Plvap silencing in these cells. Reduced angiogenic capacity of cells that lack PLVAP expression was linked to reduced VEGFR2 protein levels, selective downregulation of VEGF co-receptors and VEGFR2 down-stream signaling kinases.

Conclusions: Our data highlight PLVAP as an important endothelium-specific cofactor for physiological and pathological angiogenesis through a VEGFR2 dependent mechanism. Therefore, PLVAP may serve as a potential therapeutic target in proliferative vascular eye disorders, such as diabetic retinopathy and retinopathy of prematurity.

Keywords: 700 retinal neovascularization • 499 diabetic retinopathy • 609 neovascularization  
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