Abstract
Purpose :
We discovered that many genes mutated in retinitis pigmentosa and related retinal dystrophies (RPRRD) were hypermethylated in human fetal retinas and mouse retinal progenitor cells (RPCs). The methylation of these genes was significantly reduced during RPC differentiation into photoreceptors (PH), leading to their increased expression. Since DNA methylation silences gene expression and DNA demethylation should occur to allow gene expression, we hypothesized that unsuccessful demethylation of the genes during the RPC-to-PH transition may reduce or even eliminate their activity, leading to PH abnormalities and retinal degeneration without any mutations in these genes.
Methods :
To test our hypothesis, we created a mouse model in which the DNA demethylation pathway was inactivated in RPCs. To this end, since the TET protein family has a vital role in the DNA demethylation pathway, we crossed Tet1-, Tet2-, Tet3-floxed (TET) animals and Chx10-Cre mice to generate triple conditional knockouts (Chx10-TET) with Cre recombinase expression directed to RPCs. TET mice were used as controls. To characterize these animals, we used various methods including immunohistochemistry, transmission electron microscopy (TEM), and electroretinogram (ERG) measures.
Results :
We found that the retinal and outer nuclear layer (ONL) thickness was significantly reduced in Chx10-TET mice compared to TET controls. However, we were able to detect many rows of nuclei in the ONL of the 3-month-old Chx10-TET animals, indicating slow PH death. Our data indicate that inactivation of the DNA demethylation pathway in RPCs prevents the development of the PH outer segments. The outer plexiform layer (OPL) is almost absent in Chx10-TET animals. This results in mixing of outer and inner nuclear layers in Chx10-TET mice. TEM examination of the retinas indicates the absence of synapses in TET-deficient PH. Light and dark-adapted ERG tests indicate that Chx10-TET animals are functionally blind.
Conclusions :
Our data suggest that inactivation of the DNA demethylation pathway prevents PH development. Slow death of undeveloped PH creates a window of opportunity to remove methylation and trigger the development of these neurons, restoring their function. Thus, the contribution of retina-specific epigenetic mechanisms to the pathogenesis of RPRRD may significantly change current approaches to diagnosing and treating these diseases.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.