Abstract
Purpose :
Under physiological conditions, calcium and phosphate concentrations are tightly regulated, ensuring that calcification is restricted to bones and teeth. However, ectopic calcification in the extracellular space of soft tissues has been associated with ageing disorders including age-related macular degeneration (AMD), pseudoxanthoma elasticum (PXE), and generalized calcification in infancy (GACI). Ectopic calcification is associated with mutations on ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1). Here, we characterised the effects of exon 9 deletion in ENPP1 on ocular calcification and retinal structure and function.
Methods :
ENPP1 transgenic mice were generated by breeding mice with loxP sites flanking ENPP1 exon 9 with mice expressing Cre recombinase behind a CAG synthetic germline promoter.. Wild type, heterozygous and knockout animals were maintained for up to 6 months. Animals were regularly assessed by colour fundus photography (CFP), optical coherence tomography (OCT) and electroretinography (ERG). In addition, cadaveric eyes were assessed for calcification using OsteoSense680EX and confocal fluorescent microscopy. ENPP1-/- animals were compared to ENPP1+/- and ENPP1+/+ controls.
Results :
On CFP we identified visible lesions in the ENPP1-/- animals. No measurable difference could be found in OCT segmentation. Analysis of the ERG traces showed no appreciable changes to scotopic A-wave. However, scotopic B-wave amplitudes were significantly increased with increased implicit time in ENPP1-/- animals(2.5 cd.s/m2: 567.7 µV vs. 442.8 µV, P value: 0.0193; 8.0 cd.s/m2: 685.2 µV vs. 511.2 µV, P value: 0.0037; 25.0 cd.s/m2: 779.8 µV vs. 561.2 µV, P value: 0.0015) . Staining of cadaveric eye tissues with Osteosense showed ectopic calcification in the Bruch’s membrane but not in the retina. BrM calcification was more extensive in ENPP1-/- animals compared to ENPP1+/- and ENPP1+/+ animals.
Conclusions :
Our preliminary studies indicate that deletion of exon 9 in the ENPP1 results in changes to retinal structure and function. The functional changes in the retina and the associated Bruch’s membrane calcification could become a model for studying the effects of calcification and for the development of intervention strategies for diseases like AMD, PXE and GACI.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.