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Takeshi Yoshida, Shikun He, Mizuki Kitamura, Christine Spee, Stephen Ryan, David Hinton; The role of VEGFR-3 in the pathogenesis of choroidal neovascularization in age-related macular degeneration. Invest. Ophthalmol. Vis. Sci. 2013;54(15):334.
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Vascular endothelial growth factor (VEGF) plays a key role in the pathogenesis of choroidal neovascularization (CNV) in age-related macular degeneration (AMD). Recently, it’s reported that VEGFR-3 has a potential role in the pathogenesis of angiogenesis. The aim of the present study is to investigate the role of VEGFR-3 induced by interferon gamma IFNγ in the development of CNV in vitro.
Cryostat retinal sections from laser-induced CNV eyes of C57/BL6 mice were used for immunohistochemistry of VEGFR-3. IFNγ-induced VEGF-C in cultured human fetal retinal pigment epithelium cells (RPE) was analyzed by western blotting, and IFNγ-induced VEGFR-3 in human umbilical vein endothelial cells (HUVEC) was analyzed as well. HUVEC were transfected with scrambled or VEGFR-3 siRNA and then treated with or without IFNγ. The cells were stimulated with supernatant of RPE with or without IFNγ, and the angiogenic activity was measured using tube-formation assay. Furthermore, induction of VEGFR-2/VEGFR-3 heterodimer in HUVEC by the RPE supernatant was examined by immunoprecipitation and western blotting.
Markedly increased expression of VEGFR-3 was observed in RPE and endothelial cells in mouse CNV lesions induced by laser. In vitro, RPE stimulated with IFNγ significantly increased VEGF-C expression in supernatant of the RPE. In scrambled siRNA transfected RPE, pretreatment of HUVEC with IFNγ and treatment with the supernatant of IFNγ-treated RPE induced marked angiogenesis in tube formation assay; however, the angiogenic effect was diminished by inhibition of VEGFR-3 using siRNA in HUVECs. Furthermore, the supernatant treatment mentioned above also upregulated expression of VEGFR-2/VEGFR-3 heterodimer and phospho-ERK1/2 in HUVEC. VEGFR3 knockdown by siRNA also diminished the expression of the heterodimer and phospho-ERK1/2 in HUVEC.
Our data shows that VEGFR-3 and VEGF-C induced by IFNγ play an important role in angiogenesis. These results suggest that they might be a new therapeutic target for the treatment of wet AMD.
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