Tafluprost Promotes Axon Regeneration After Optic Nerve Crush via Zn <sup>2 </sup>-mTOR Pathway

Purpose: To investigate whether tafluprost could promote optic nerve regeneration in mice after optic nerve crush (ONC), and to find out the underlying molecular mechanism.Methods: Tafluprost was injected into the vitreous body immediately after ONC. The level of Zn2+ in the inner plexiform layer (IPL) of retina was stained by autometallography (AMG). The number of survival retinal ganglion cells (RGCs) were counted with co-stained by neuronal class βIII tubulin (TUJ1) and RBPMS. Individual axons that regenerated to 0.25mm, 0.5mm, 0.75mm and 1mm were manually counted in the whole-mount optic nerve labeled by cholera toxin B fragment (CTB). Immunofluorescence and western blot were performed to detect protein expression levels. Pattern electroretinogram was used to evaluate RGCs function.Results: 1μM tafluprost significantly decreased zinc transporter ZnT-3 expression and reduced Zn2+ accumulation in the IPL of retina. Tafluprost increased RGC survival, stimulated intense axonal regeneration, and maintained RGCs function as compared with control. Mechanistically, either tafluprost or Zn2+ elimination treatment (TPEN or ZnT-3 deletion) can activate the mTOR pathway with improved percentage of pS6+ RGC in retina. However, rapamycin, a specific inhibitor of the mTOR1, inhibited the activation of the mTOR pathway and abolished the regenerative effect mediated by tafluprost following ONC. Tafluprost also inhibited the upregulation of p62, LC3 and Bclin-1, attenuated the overactivation of microglia/macrophages and downregulated the expression of TNFα, IL-1β and IL-6.Conclusions: Our results suggest that tafluprost promoted axon regeneration via regulation of the Zn2+-mTOR pathway, and provide novel research directions for the mechanisms of glaucomatous optic nerve injury.