Abstract
Purpose :
Lafora disease (LD) is an autosomal recessive progressive neurologic disorder caused by mutations in EPM2A or EPM2B genes, both involved in glycogen structural integrity. Defective function of EPM2A or EPM2B results in accumulation of malformed insoluble glycogen termed Lafora bodies (LBs) in the central nervous system. This study aimed to characterise the retinal phenotype of Epm2a-/- mice to characterize retinal neuronal changes in LD.
Methods :
The study was approved by the animal care committee at the Toronto Center for Phenogenomics. Five to seven knockout (KO; Epm2a-/-) and control (WT) littermates were examined at two time points (10 and 14 months, respectively). Mice were injected 0.01 ml/g body weight of anesthetic solution (combination of 100mg/ml Ketamine and 20mg/ml Xylazine). Electroretinogram (ERG) testing was performed following overnight dark adaptation (DA) using Lab Cradle (Diagnosys LLC). A scotopic intensity series ERG was performed (9 steps; 0.0025 cd.s.m-2 – 10 cd.s.m-2) followed by a 10-minute light adaptation (LA; 30 cd.m-2). Subsequently, LA ERGs were performed using a 5 cd.s.m-2 stimulus flash to four stimulus frequencies (5 - 20 Hz). Retinal optical coherence tomography (OCT) and fundus imaging was performed using Phoenix MICRONTM. Periodic acid Schiff-Diastase (PASD) staining of the retina was performed to ascertain LB deposition. After staining, slides were imaged at 40x using 3DHistech Pannoramic Flash II Slide Scanner, analyzed using 3DHistech CaseViewer software and LBs were counted using 3DHistech QuantCenter. Regions of interest were selected on the retinal sections, and a scenario was built in QuantCentre that distinguishes LBs from background and other structures.
Results :
Retinal photographs were normal in both KO and WT mice. There was no significant difference in DA and LA ERG b-wave amplitudes between KO and WT mice groups at 10 and 14 months. The total retinal thickness was similar in both KO and WT groups. On PASD staining, LBs were observed only in KO mice in inner plexiform layer (IPL) and inner nuclear layers. Upon quantification, average number of LB accumulation in IPL in KO mice were 1743±533 and 2615±915 per mm2, at 10 and 14 months, respectively.
Conclusions :
This Epm2a-/- mice model demonstrates LB deposition in the bipolar cell nuclear layer and its synapse with retinal ganglion cells. Adaptive optics retinal imaging could visualize them in vivo and serve as a useful biomarker for LD.
This is a 2021 ARVO Annual Meeting abstract.