Histone Deacetylase 3 (HDAC3) plays an important role in retinal ganglion cell death after acute optic nerve injury

Abstract

Purpose: HDAC activity mediates gene silencing and apoptosis in dying retinal ganglion cells. An important role in these processes may be played by HDAC3, which exhibits nuclear translocation in ganglion cells shortly after optic nerve injury. The purpose of this study was to test the hypothesis that knock out of Hdac3 transcription will lead to protection of retinal ganglion cells from histone deacetylation-related gene silencing and death following acute injury.

Methods: Hdac3^fl/fl mice were given 1μL intravitreal injections of AAV2-CRE virus (10¹² genome copies/mL) into the OS eye to excise Hdac3. Rosa26-LacZ^fl/fl or Rosa26-LacZ^fl/fl mice were treated identically to serve as controls. After eight weeks, injected eyes underwent optic nerve crush. Five days post-optic nerve crush, retinas were harvested and analyzed using fluorescent microscopy or qPCR quantification of ganglion cell mRNAs. Analyses included HDAC2, HDAC3, activated caspase 3, and acetylated histone 4 (AcH4) labeling. Eyes were also harvested 14 days post-optic nerve crush and retinas analyzed using fluorescent microscopy for total retinal ganglion cell counts.

Results: : Rosa26-LacZ^fl/fl mouse eyes expressed HDAC2 and HDAC3 at 5 days following optic nerve crush, but exhibited wide-spread histone deacetylation in the ganglion cell layer characteristic of optic nerve crush. Deletion of Hdac3 in ganglion cells of Hdac3^fl/fl mice led to loss of HDAC3 expression and retained expression of HDAC2 and AcH4. Ganglion cells of Rosa26-LacZ^fl/fl mice also labeled positive for active caspase 3, an indicator of cell apoptosis, more frequently than those of Hdac3^fl/fl mice (P<0.0001). Cell counts at 14 days post optic nerve crush indicated significant preservation of retinal ganglion cell populations in injected Hdac3^fl/fl eyes in comparison to injected Rosa26-LacZ^fl/fl and non-injected Hdac3^fl/fl mice (P<0.001). Quantitative analysis of ganglion cell gene expression is on-going and will be reported.

Conclusions: These results support the hypothesis that Hdac3 plays a critical role in the pathology of retinal ganglion cells following acute optic nerve injury, making HDAC3 a possible target for glaucoma drug therapy.

Keywords: 615 neuroprotection • 531 ganglion cells • 536 gene modifiers