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Micah A Chrenek, Jana T Sellers, Kristie Ling Liao, Brooke Still, John M Nickerson, Jeffrey H Boatright; Rpe65 L450 allele increases sensitivity to light damage in C57BL/6J mice. Invest. Ophthalmol. Vis. Sci. 2019;60(9):2358. doi: https://doi.org/.
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To test whether C57BL/6J mice with the L450 allele of Rpe65 are more sensitive to LIRD than mice with the M450 allele. Danciger et al. (Mamm Genome 2000) reported a QTL on chromosome 3 that correlates with the location of RPE65 as influencing the severity of LIRD in mice. Based on this, we hypothesized that C57BL/6J mice with the wildtype L450 allele of Rpe65 would be more sensitive to light induced retinal degeneration (LIRD) compared to the typical C57BL/6J mouse with the M450 allele. We backcrossed the Rpe65-L450 allele into the C57BL/6J stain and assessed whether they are more sensitive to LIRD.
A line of C57BL/6J mice with the wildtype Rpe65-L450 allele was established by backcrossing 129/Sv mice to C57BL/6J mice for 10 generations. Mice heterozygous for Rpe65 were crossed to generate litters with homozygous M450, homozygous L450, and heterozygous pups. LIRD was induced using 40000 lux white LED light for 4 hours. ERGs were performed 7 days after exposure. Comparisons of gene expression, immunohistochemistry for stress markers, and counts of specific retinal cell types is ongoing. Sample sizes were 3 mice per group with statistical analysis using 2-way ANOVA with Tukey’s post-hoc analysis.
We observed a significant loss of ERG function in mice with either allele, with M450 homozygous mice having 47% a-wave and 58% b-wave amplitudes of dim controls (p<0.0001), and L450 homozygous mice having 26% a-wave and 25% b-wave amplitudes of dim controls (p<0.0001). The difference between the homozygous M450 and L450 mice was also significant (p=0.0013 for a-waves, p<0.0001 for b-waves). Counts from sections 200-300 mm from the optic nerve showed significant decreases in photoreceptor nuclei after light damage (p<0.0001). M450 homozygotes retained 40% and L450 homozygotes retained 15% of the photoreceptor nuclei.
C57BL/6J mice with the L450 allele of Rpe65 are more sensitive to LIRD than mice with the M450 allele. Since RPE65-M450 has 45% retinal isomerase activity as compared to RPE65-L450 (Redmond, 2007), we conclude that retinal recycling rate influences sensitivity to LIRD. Retinal recycling rate may also affect other photoreceptor degenerative processes and should be considered when studying models on the C57BL/6J background. We believe C57BL/6J mice with the L450 allele of Rpe65 will be useful for studies of retinal function in the context of a fully active vitamin A Cycle.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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