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Hongmin Yun, Robert L Hendricks; VEGF-A regulates sympathetic nerve innervation in murine herpes stromal keratitis (HSK). Invest. Ophthalmol. Vis. Sci. 2018;59(9):4934. doi: https://doi.org/.
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HSK is a potentially blinding corneal disease that manifests in both humans and mice as corneal edema, opacity, neovascularization, and hypoesthesia. Mice with primary HSK completely lose corneal sensory nerves leading to severe, diffuse inflammation that is maintained by subsequent hyperinnervation with sympathetic nerves. Eliminating sympathetic nerves from the cornea by excising the superior cervical ganglion regenerates corneal sensory nerves and reverses corneal opacity and neovascularization. Our current study explores the mechanism(s) of sympathetic innervation with the goal of blocking the ingrowth of sympathetic nerves and promoting the regeneration of sensory nerves as a novel therapy for HSK.
Mouse corneas were infected with HSV-1 and whole mounts were stained with fluorescent antibodies to VEGF-A, CD4, F4/80, CD31 and lyve-1, and were stained for β III tubulin(all nerves), tyrosine hydroxylase(sympathetic nerves), and substance P(sensory nerves) to define nerve changes. Mice received subconjunctival injections of soluble VEGF receptor 1(sVR1, 5ug, 3 times/week) to neutralize VEGF-A, clodronate liposomes(100 ul, once a week) to deplete macrophages, anti-CD4 antibody(150ug, once a week) to deplete CD4 T cells, or Avastin (500ug, 3 times/week) to block VEGF-A or appropriate controls. Mice were monitored for corneal opacity, neovascularization, and blink reflex. Z-stacks spanning entire cornea were acquired by confocal microscopy.
CD4 T cells, macrophages, and vascular endothelial cells expressed VEGF-A in HSK corneas. Depleting mouse corneas of macrophages or CD4 T cells dramatically decreased sympathetic innervation in the corneal stroma, and CD4 T cell depletion further initiated regeneration of corneal sensory nerves and recovery of corneal blink reflex. Subconjunctival injection of sVR1 caused partial recovery of cornea reflex, only sectorial innervation of sympathetic nerves, and less severe corneal opacity. Subconjunctival injection of Avastin which reportedly fails to completely neutralize mouse VEGF-A resulted in unstable recovery of cornea blink reflex and decreased sympathetic innervation. Optimization of VEGF blockade in the cornea is ongoing.
VEGF-A regulates HSK in part by promoting hyperinnervation of sympathetic nerves. We propose that CD4+ T cells maintain HSK at least in part through directly or indirectly regulating VEGF production in the cornea.
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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