Longitudinal characterization of molecular and pathological changes in the retina of a large animal model of CLN3 disease.

Abstract

Purpose : CLN3 disease leads to retinal degeneration and vision loss in affected patients. However, because postmortem retina from CLN3 disease patients is severely degenerated, it is difficult to determine the primary versus secondary pathological changes in the human eye. CLN3^Δ7/8 minipig, a large animal model of CLN3 disease that shows vision loss provides an appropriate platform to study CLN3 retina (patho)physiology. Therefore, the purpose of this study was to perform longitudinal characterization of molecular and pathological changes in the CLN3^Δ7/8 minipig retina.

Methods : Methods: Age-matched Wild-type and CLN3^Δ7/8 minipig retina were compared longitudinally (1 month, 6 month, 14 month, 36 month and 48 month). Techniques used to evaluate retina function, structural integrity and molecular composition included i) electroretinogram (ERG), ii) optical coherence tomography (OCT), iii) histological analyses utilizing transmission electron microscopy (TEM) and confocal microscopy following immunohistochemistry and iv)Western blotting for cell-type specific retina markers (e.g., EZR: RPE, RHO, RCVRN: photoreceptors).

Results : CLN3^Δ7/8 minipig retina showed reduced level of RCVRN, RHO (photoreceptors), CHX10 (rod bipolar cells), and EZR (RPE) when compared to age-matched wild-type minipig retina at 6 month of age. ERG analysis showed reduced scotopic b-wave amplitude in the 6 month CLN3^Δ7/8 retina. CLN3^Δ7/8 minipig retina at the 6 month also showed reduced outer nuclear layer thickness and rod photoreceptor outer segment disorganization. In addition, EM evaluation showed presence of disease-associated fingerprint-like material and altered melanosome polarity in the 6 month CLN3^Δ7/8 minipig RPE when compared to wild-type minipig RPE. Furthermore, compared to wild-type minipig RPE, reduced autofluorescence/lipofuscin was seen in CLN3^Δ7/8 minipig RPE at the 36 month of age and this was followed by almost complete loss of outer nuclear layer in the CLN3^Δ7/8 minipig retina at 48 month of age.

Conclusions : CLN3^Δ7/8 minipig retina shows structural, functional and molecular alterations at 6 month of age. Furthermore, complete loss of outer nuclear layer in CLN3^Δ7/8 minipig model is preceded by rod photoreceptor outer segment disorganization and reduced RPE lipofuscin.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.