HDAC1/2 are critical for RPE homeostasis and health

Abstract

Purpose : Histone deacetylases are a family of 18 known members, classified in four groups based on their homology to yeast proteins. In previous RNA-seq data, we found decreased expression of HDAC1/2 in RPE/choroid samples from human eyes with advanced dry age-related macular degeneration (AMD). Here, we report on the development of experimental models of HDAC1/2 deficiency and discover a critical role for these epigenetic regulators in the homeostasis and health of the RPE.

Methods : Hdac1^f/f, Hdac2^f/f, and Hdac1/2^f/f (gift of Eric Olson) mice were crossed with Vmd2-Cre (Jackson Labs). Conditional ablation of Hdac1/2 in RPE and localized Cre expression were confirmed by qPCR and immunofluorescence (IF). Fundus examination by multimodal imaging and ERG were performed (n=12-16). RPE morphology was examined by ZO-1 IF on RPE/choroidal flatmounts, and sections were analyzed by H&E staining (n=4-6). AAV2-iCre or control AAV2-Gfp sub-retinal injections were performed in WtC57BL/6J, Hdac1^f/f, Hdac2^f/f, and Hdac1/2^f/f (n=4-6) followed by fundus examination at 14 days. Intravitreous injections of small molecule class I HDAC inhibitors and siRNAs were studied in WtC57BL/6J mice (n=6-8).

Results : Hdac1/2^f/fx Vmd2Cre exhibited a robust phenotype characterized by pan-retinal degeneration with complete atrophy of the RPE by 1 month of age (Figure 1) and focal areas of depigmentation in their fur coat. Viable RPE was present at 2 weeks of age as shown in fundus imaging and ZO-1 IF. Loss of Hdac1/2 expression and Cre expression were confirmed in the RPE by qPCR and IF. At 1 month, SD-OCT revealed significant disruptions in the outer retina, and full-field ERGs had markedly diminished a- and b-wave amplitudes. Hdac1^f/f and Hdac2^f/f mice with Vmd2Cre demonstrated a less severe phenotype with focal RPE degeneration which developed at 2 months. AAV2-iCre (Figure 1) and siRNAs or small molecule inhibitors targeting HDAC1/2 induced focal areas of RPE degeneration as compared to control treatments.

Conclusions : Conditional ablation of Hdac1/2 in the mouse RPE resulted in severe degeneration and complete atrophy by 1 month. Similar results were achieved with postnatal ablation or inhibition of Hdac1/2 with siRNAs or small molecules. This pathway may be important in the pathogenesis of dry AMD, as the expression of Hdac1/2 is significantly decreased in the advanced stages of this disease.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.