Fig. 4: Porf-2 affects axon regeneration and growth cone formation by altering the activity of Rac1. | Cell Death & Disease

Fig. 4: Porf-2 affects axon regeneration and growth cone formation by altering the activity of Rac1.

From: Knockdown of Porf-2 restores visual function after optic nerve crush injury

Fig. 4

A Experimental timeline. Vehicle (Veh) or NSC23766 (NSC) was administered intraperitoneally once every two days after AAV2-shPorf-2 injection. B Confocal images of optic nerves from mice injected with AAV2-shPorf-2 14 days before ONC injury and treated with either Veh or NSC intraperitoneally. Asterisks indicate the optic nerve crush site. Scale bar, 200 μm. C Quantification of optic nerve regeneration in B (two-way ANOVA followed by Bonferroni’s multiple-comparisons test, p < 0.0001 at 0.20, 0.50, and 1.00 mm from the crush site; p < 0.05 at 1.50 mm from the crush site; n = 8 mice in each group). D Representative confocal images of retinal sections showing surviving RBPMS+ RGCs in the NSC-injected and control mouse groups. Scale bar, 20 μm. E Quantification of the RGC survival rate in D (Mann–Whitney test, p < 0.01; n = 6 mice in each group, at least eight non-adjacent retinal sections per mouse). F Experimental timeline. For in vivo studies, Veh or NSC was administered intraperitoneally once every two days after the AAV2-shPorf-2 injection. For in vitro experiments, Veh or NSC was added to the retinal explant medium, with the medium being changed once every two days. G Representative microphotographs of retinal explants stained with anti-beta III tubulin (Tuj1) showing neurite growth in Veh- or NSC-treated explants. Scale bar, 100 μm. H Quantification of neurites at different distances from the edges of the explants in (G) (two-way ANOVA followed by Bonferroni’s multiple-comparisons test, p < 0.0001 at 50, 100, 200, and 300 μm from the edges of the explants; p < 0.05 at 400 μm from the edges of the explants; n = 5 mice in each group). I Representative confocal images of neurites at the end of an axon in retinal explants from Veh- or NSC-treated groups stained with AAV2-EGFP (green), phalloidin (red), and Tuj1 (gray). Scale bar, 5 μm. J Percentage of growth cone-forming neurites in retinal explants in I (Mann–Whitney test, p < 0.01; n = 6 mice in each group, six to eight growth cones were analyzed for each retinal explant). Data are presented as the mean ± SEM. *p < 0.05, **p < 0.01, ****p < 0.0001. ns, not significant.

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