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Z. Ablonczy; The Effects of Vegf on the Tight Junction Proteome of Arpe–19 Cells . Invest. Ophthalmol. Vis. Sci. 2006;47(13):2891.
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© ARVO (1962-2015); The Authors (2016-present)
Macular edema is a major cause of reduced visual acuity in several eye diseases, and following ocular surgery, or injury. Subretinal fluid can accumulate following an increase in retinal pigment epithelium (RPE) paracellular permeability. These changes in the RPE permeability can be ascribed to reversible and irreversible modifications to the tight junction proteome; however, the cellular and molecular events that mediate these changes have not been established.
The human RPE cell line, ARPE–19, was used as a model to investigate VEGF–induced modifications on the barrier properties and on the tight–junction proteome of cultured RPE cells. The VEGF–induced changes in the RPE barrier function were characterized by trans–epithelial resistance (TER) measurements, and changes in the ARPE–19 cell proteome were analyzed in terms of abundance and post–translational modifications using the iTRAQ quantitative proteomics approach. The effects of VEGF inhibitors and competitors were also investigated.
Addition of VEGF (5 ng/mL) produced a prolonged robust (∼50%) drop in the TER, paralleled by a quantitatively similar rearrangement in proteins associated with both cytoskeleton and cell adhesion molecules. In the VEGF–treated samples, collagen 1A1, α–actinin, phospholipase A2, ubiquitin protease, pyruvate dehydrogenase, MMP–9 were present in higher quantities than in vehicle–treated controls. At the same time the levels of annexin 5, filamin C, prolyl–4–hydroxylase, ubiquitin/actin fusion protein, thymosin–α, keratin 1, serine protease 18 were significantly lower when compared to controls.
These data demonstrate that RPE permeability is regulated by VEGF and that the proteins, structurally connecting the tight junctions to the cytoskeleton, play a crucial role in regulating the barrier function of the RPE cell layer.
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