Semaphorin 7a actions on nerves and myeloid cells in the cornea promote nerve regeneration, thus linking neuronal and myeloid systems

Abstract

Purpose: We determined Sema7a localization and quantity in naive corneas and during nerve regeneration after lamellar flap surgery. We also performed structure-function analyses of Sema7a to determine whether peptides that span RGD integrin-binding or RTS disintegrin motifs selectively influence neuroregeneration.

Methods: Immunolocalization and Western blot analyses were performed to evaluate the abundance of Sema7a in naive corneas and corneas undergoing nerve regeneration after lamellar corneal surgery in thy1-YFP+ neurofluorescent mice. We used compartmental cultures of dissociated trigeminal ganglion cells to determine the effect of Sema7a exposure on neurite outgrowth in vitro. Finally, a Sema7a pellet was implanted under the corneal flap after lamellar transection surgery to determine the neuronal and inflammatory effects of Sema7a supplementation in vivo. We synthesized N- acetylated, C-amidated RGD- or RTS-containing peptides derived from the Sema7a sequence and determined the number of dissociated TG cells that adhered to collagen coated culture plates when incubated with the peptides.

Results: Sema7a is expressed in the cornea, mainly concentrated in the epithelium with less expression in the stroma. Corneal Sema7a expression increases after nerve transecting lamellar surgery and is localized near the regenerating nerve fronds. Sema7a induces neurite growth in vitro as potently as NGF. Exposure of trigeminal neurites to Sema7a (20 nM) in the side compartment significantly increased neurite length. The implanted Sema7a pellet significantly increased YFP+ cell (CD11b+GR1+ immature myeloid derived suppressor cells) influx into the cornea as well as increased corneal nerve length. In contrast to 7A-RGD peptide, 7A-RTS peptide does not stimulate neurite growth but it reduces adhesion of neurons to collagen surface.

Conclusions: Sema7a is constitutively expressed in the cornea and potently stimulates nerve regeneration and influx of immature myeloid cell. Sema7a peptides spanning integrin-binding motifs (RGD or RTS) selectively influence neuroregeneration. This immune semaphorin links nerve regeneration and myeloid systems in the cornea.

Keywords: 480 cornea: basic science • 687 regeneration