Geographic characterization of RPE structure and lipid changes in the PEX1-p.Gly844Asp mouse model for Zellweger spectrum disorder.

Peroxisome Biogenesis Disorders-Zellweger Spectrum (PBD-ZSD) are a heterogenous group of autosomal recessive disorders caused by defects in PEX genes whose proteins are required for peroxisome assembly and function. Peroxisomes are ubiquitous organelles that play a critical role in complex lipid metabolism. Dysfunctional peroxisomes in ZSD cause multisystem effects, with progressive retinal degeneration (RD) leading to childhood blindness being one of the most frequent clinical findings. Despite progress in understanding the role of peroxisomes in normal cellular functions, much remains unknown about how their deficiency causes RD, and there is no treatment. To study RD pathophysiology in this disease, we used the knock-in PEX1-p.GlyG844Asp (G844D) mouse model of milder ZSD, which represents the common human PEX1-p.Gly843Asp allele. We previously reported diminished retinal function, functional vision, and neural retina structural defects in this model. Beyond the neural retina, structural defects in retinal pigment epithelium (RPE) have been reported in ZSD patients and murine models with single peroxisome enzyme deficiency, suggesting that RPE degeneration may contribute to overall RD progression in this disease. Here, we investigate the RPE phenotype in our PEX1-G844D mouse model, observing morphological, inflammatory, and lipid changes at 1, 3, and 6 months of age. We report that RPE cell degeneration appears at 3 months of age and worsens with time, starts in the dorsal pole, and is accompanied by subretinal inflammatory cell infiltration. We match these events with lipid remodelling using imaging mass spectrometry which allowed regional analysis specific to the RPE cell layer. We identified 47 lipid alterations that precede structural changes, 10 of which are localized to the dorsal pole. 32 of these lipid alterations persist to 3 months, with remodelling of the lipid signature at the dorsal pole. 14 new alterations occur concurrent with histological changes. Changes in peroxisome-dependent lipids detected by liquid chromatography tandem mass spectrometry (reduced docosahexanoic acid and increased very long chain lysophosphatidylcholines) are exacerbated over time. This study represents the first characterization of RPE in any animal model of ZSD, and the first in situ lipid analysis in any peroxisome-deficient tissue. Our findings reveal candidate lipid drivers that could be targeted to alleviate RD progression in ZSD, as well as candidate biomarkers that could be used to evaluate retinopathy progression and response to therapy.