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C. Q. Yu, M. Zhang, M. I. Rosenblatt; Vascular Endothelial Growth Factor Mediates Corneal Nerve Repair. Invest. Ophthalmol. Vis. Sci. 2008;49(13):2947.
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To examine the vascular endothelial growth factor (VEGF) dependence of corneal nerve regeneration after superficial injury.
Trigeminal neurons collected from mice were dissociated and cultured. Cells were grown in Neurobasal media and 2% FBS with either 250 ng/mL VEGF specific neutralizing antibody (bevacizumab) or 250 ng/mL nonspecific mouse immunoglobulin. After 48 hours samples were fixed, stained with nerve specific antibodies and neuronal outgrowth quantified. Groups of thy1-YFP neurofluorescent mice were anesthetized and received 5 uL intrastromal injections of either 25 mg/mL bevacizumab or 25 mg/mL nonspecific mouse immunoglobulin. 24 hours after treatment a 2 mm circular corneal epithelial defect was made in each anesthetized mouse which removed their corneal epithelium and subbasal nerve plexus. 72 hours after wounding mice were sacrificed, their corneas whole mounted, and nerve regeneration in the area of the wound was quantified by fluorescence microscopy.
After 48 hours growth cultures of dissociated trigeminal neurons showed extensive axon outgrowth which extended over the surface of each well. Those treated with anti-VEGF antibody showed a 17% reduction in nerve process density compared to control. In living mice three days after superficial keratectomy, extensive nerve regeneration had already occurred. Mice treated with bevacizumab displayed a 23% reduction in the neuronal density of their subbasal plexus in the area of the wound compared to those treated with non specific immunoglobulin.
This study provides evidence that VEGF signaling influences the repair of corneal nerves after injury by demonstrating that abrogation of VEGF signaling reduces nerve growth in vivo and in vitro.
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