Zusammenfassung
PURPOSE. Choroidal neovascularization in age-related macular degeneration is caused, to a large extent, by increased secretion of vascular endothelial growth factor (VEGF)-A by the retinal pigment epithelium (RPE). The purpose of the study was to identify pathways that lead to increased VEGF secretion by the RPE. METHODS. Ca2(+) signaling was studied in ARPE-19 and human RPE cells in primary ...
Zusammenfassung
PURPOSE. Choroidal neovascularization in age-related macular degeneration is caused, to a large extent, by increased secretion of vascular endothelial growth factor (VEGF)-A by the retinal pigment epithelium (RPE). The purpose of the study was to identify pathways that lead to increased VEGF secretion by the RPE. METHODS. Ca2(+) signaling was studied in ARPE-19 and human RPE cells in primary culture by means of Ca2(+) imaging. Membrane conductance was measured in the whole-cell configuration of the patch-clamp technique. VEGF-A secretion was measured by using ELISA. RESULTS. Freshly isolated RPE cells or ARPE-19 cells were shown to express TRPV1, -2, -3, and -4 channels. Increasing the temperature or stimulation by IGF-1 increased the VEGF-A secretion rate in both cell types. These effects were both sensitive to the TRPV channel blocker ruthenium red (20 mu M). The heat-inducible Ca2(+) signals were blocked by the TRPV channel blockers La3(+) and ruthenium red by 68% and 52%, respectively. In contrast, high concentrations of 2-APB (3 mM) increased [Ca2(+)](i), whereas the TRPV1 channel opener capsaicin and the TRPV3 channel opener camphor had no effect. Reduction of TRPV2 expression by siRNA attenuated the heat-evoked Ca2(+) response. In addition, a heat-activated inwardly rectifying current was measured that was completely blocked by ruthenium red. IGF-1 also increased whole-cell current with a corresponding increase in [Ca2(+)](i), which was blocked by the PI3-kinase blocker LY294002. CONCLUSIONS. The data strongly suggest that TRPV2 channels expressed by the RPE are involved in the Ca2(+) signaling that mediates both heat-dependent and IGF-1 (via PI3-kinase activation)-induced VEGF secretion. (Invest Ophthalmol Vis Sci.2010;51:6001-6008) DOI:10.1167/iovs.09-4720
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