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. 2023 Mar 31;18(11):2526–2534. doi: 10.4103/1673-5374.371372

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Copyright: © Neural Regeneration Research

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The survival of RGCs after ONC requires the activity of S6K1 and 4E-BP1.

(A) AAV-EGFP, AAV-caS6K1, and AAV-DN-S6K1 were administrated intravitreally to mice 2 weeks before ONC. Eyes were harvested on day 14 after ONC. (B) Overexpression of caS6K1 but not DN-S6K1 increased RGC survival. Fluorescent photomicrographs of retinal whole-mounts showing surviving Tuj1⁺ RGCs 14 dpc from eyes injected with AAV-EGFP, AAV-caS6K1, and AAV-DN-S6K1. (C) Quantification of sizes of surviving RGCs from AAV-EGFP, AAV-caS6K1, and AAV-DN-S6K1 groups (mean ± SD, n = 4). (D) Quantification of surviving Tuj1⁺ RGCs from eyes injected with AAV-EGFP, AAV-caS6K1, and AAV-DN-S6K1 (mean ± SD, n = 6–8). € AAV-EGFP, AAV-sh4E-BP1, and AAV-4E-BP1-4A were administrated intravitreally to mice 2 weeks before ONC. Eyes were harvested on day 14 after ONC. (F) Overexpression of 4E-BP1-4A increased RGC survival while knockdown of 4E-BP1 decreased RGC survival. Immunofluorescence of retinal whole-mounts showing surviving RGCs 14 dpc from AAV-EGFP, AAV-sh4E-BP1, and AAV-4E-BP1-4A groups. Scale bars: 60 µm (B, F). (G) Quantification of surviving RGCs from eyes injected with AAV-EGFP, sh4E-BP1, and 4E-BP1-4A (mean ± SD, n = 6–8). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 (one-way analysis of variance with Bonferroni’s post hoc test). AAV: Adeno-associated virus; caS6K1: constitutively active S6K1; DN-S6K1: dominate negative S6K1; EGFP: enhanced green fluorescent protein; ONC: optic nerve crush; RGC: retinal ganglion cell; S6K1: mammalian target of rapamycin complex 1 downstream effector; Tuj1: β3-tubulin.