Histone deacetylase 2 inhibition promotes neurite outgrowth of retinal ganglion cells

Abstract

Purpose : Epigenetics mechanisms play an integral role in tissue development, damage, and regeneration and dictate the heterochromatin configuration of the genetic material to influence gene translational activities and mediate tissue and nerve regeneration. The acetylate states of histones have been used as an indicator of the developmental state of progenitor and differentiating cells. The goal of this study was to investigate if HDAC2, a histone deacetylase, holds a key to regulate the survival and neurite outgrowth of neurons, especially retinal ganglion cells (RGCs).

Methods : PC12 cells acquired from ATCC and primary RGCs isolated from mice aged postnatal day 9 (P9) were cultured and treated with suberoylanilide hydroxamic acid (SAHA; 1 µM), a pan HDAC inhibitor, and Santacruzamate A, an HDAC2 specific inhibitor (1 µM), respectively, for 3 – 4 days. The effects of both inhibitors on neurite outgrowth were analyzed following image acquisition. Additionally, PC12 cells were treated with increasing concentrations (0, 5 and 100 ng/mL) of nerve growth factor (NGF), and HDAC2 levels were evaluated using qPCR after 24 hours. Retinal explants from RGC specific HDAC2 knockout and littermate control mice were cultured for 5 days and stained for βIII-tubulin to reveal growing axons. Two-tailed Student's t-test was used for statistical analysis.

Results : Treatment with both SAHA and Santacruzamate A increased average neurite length and number in cultured PC12 cells and primary RGCs. qPCR data revealed a dose dependent decrease of HDAC2 levels in PC12 cells following treatment with increasing concentration of NGF. Additionally, retinal explants from RGC specific HDAC2 knockout mice exhibited increased neurite outgrowth relative to controls.

Conclusions : These data indicate a beneficial effect of blocking HDACs, especially HDAC2, at promoting neurite outgrowth. Our results point to an active role of histone acetylation/deacetylation in RGC axonal growth/regeneration. An in-depth understanding of HDACs and other associated underlying epigenetic mechanisms can help us design novel therapeutics to address glaucoma and other pathology that triggers tissue damage.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.