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Jin-Hong Chang, Kyu Yeon Han, Dimitri T Azar; Characterization of Injury-induced Corneal Angiogenesis and Lymphangiogenesis in Prox1-GFP/Flt1-dsRed Transgenic Mice. Invest. Ophthalmol. Vis. Sci. 2018;59(9):2578.
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The molecular mechanisms responsible for the innate avascularity of the cornea as well as vascular and lymphatic growth and regression in response to insult remain poorly understood. To characterize the kinetics of injury- and growth factor-induced blood and lymphatic vessel growth in the cornea, we bred Prox1-GFP/Flt1-dsRed (PGFD) transgenic mice with conditional knockout mice deficient in vascular endothelial growth factor receptors (VEGFRs).
We bred Prox1-GFP mice and Flt1-dsRed mice to generate PGFD mice and then bred PGFD mice with Prox1CreERT2, CDh5CreERT2, and VEGFR2lox mice to generate CDh5CreERT2/VEGFR2lox/PGFD and Prox1creERT2/VEGFR2lox, Prox1cre/CDh5CreERT2VEGFR2lox mice. VEGF-A and -C pellets, alkali burn, and five combinations of hemi-limbal injury were applied to PGFD and conditional KO mouse corneas. Injured corneas were imaged using Axiozoom V16 stereomicroscopy and analyzed with Zeiss Zen imaging programs, and the areas of induced blood and lymphatic vessels were calculated using Adobe Photoshop. RNA-seq was used to assay the mRNA levels in VEGF-C pellet-induced untreated and tamoxifen-treated Prox1cre/CDh5CreERT2VEGFR2lox mouse corneas.
Injury of half the limbus, the whole limbus, or the whole cornea caused little to no growth of blood or lymphatic vessels in the PGFD mouse cornea compared to that induced by VEGF-A or VEGF-C pellet implantation from day 1 to 14 after treatment. Simultaneous injury via scraping of half the limbus and the whole cornea (hemilimbal deficiency model) resulted in blood and lymphatic vessel growth. Conditional deletion of vascular endothelial VEGFR2 resulted in diminished corneal lymphangiogenesis (LA) in the hemilimbal deficiency injury model and abolished VEGF-A–induced corneal LA. In addition to this, conditional deletion of both vascular and lymphatic VEGFR2 in Prox1CreERT2VEGFR2lox/PGFD mice resulted in abolished VEGF-C–induced LA compared to that in the control. RNA-seq data showed diminished PDGF receptor pathway signaling after VEGF-C pellet implantation in conditional lymphatic endothelial VEGFR2 KO mice.
The Prox1-GFP/Flt1-dsRed transgenic mouse provides an excellent model for characterizing injury-induced corneal neovascularization. Also, vascular and lymphatic endothelial VEGFR2 plays a vital role in regulating injury-induced corneal LA.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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