Abstract
Purpose :
Herpes stromal keratitis (HSK) is a potentially blinding corneal disease, and like many corneal dystrophies, is characterized by loss of sensation and persistent inflammation in both humans and mice. Corneal sensory nerve retraction in mouse primary HSK leads to severe, diffuse inflammation that is maintained by hyperinnervation with sympathetic nerves. Our current study illuminates a role for vascular endothelial growth factor (VEGF)-A in the promotion of sympathetic innervation in the cornea. Further, our data reveal that inhibition of VEGF signaling, independent of blood vessel formation, alleviates disease and re-establishes a more normal nerve network within the cornea.
Methods :
Mouse corneas were infected with HSV-1 and either: 1) corneal whole mounts were stained with antibodies against VEGF-A, immune cell populations, and sensory (substance P)/sympathetic nerves (tyrosine hydroxylase); or 2) corneas were analyzed by flow cytometry to detect immune cells producing VEGF-A. In vitro sensory or sympathetic neurons were cultured with varying concentrations of VEGF-A or with lysates from corneas that were experiencing HSK in the presence or absence of VEGF-A neutralizing antibodies. In a mouse model of HSK, we depleted immune cells or neutralized VEGF-A by subconjunctival injection of antibodies or isotype controls. Blinded observers measured pathology by assessing corneal opacity, neovascularization, and blink reflex.
Results :
During HSK, CD4+ T cells and macrophages are the primary producers of VEGF-A. Corneal VEGF-A during peak disease promotes sympathetic nerve growth while repressing sensory nerves. Locally depleting macrophages or CD4 T+ cells, or locally neutralizing VEGF-A with bevacizumab resulted in a diminishment of sympathetic innervation, a recovery of corneal blink reflex, and less corneal opacity.
Conclusions :
Our findings highlight a previously unrecognized role for VEGF-A in the maintenance of the corneal nerve landscape during health and disease. In addition to finding the sources of VEGF-A during HSK (CD4+ T cells and monocytes), we identify an alternative use for bevacizumab (Avastin), which targets sympathetic nerves in our model rather than blood vessels. It is likely that this mechanism influences other models of disease that rely on appropriate sensory/sympathetic innervation of tissues.
This is a 2020 ARVO Annual Meeting abstract.