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Peter Hao Tang, Mark Pierson, Neal D. Heuss, Katarina Wrzos, Dale S Gregerson; Elevated presence of dendritic cells in retina persist late into life of a mouse model for Type 2 Leber congenital amaurosis. Invest. Ophthalmol. Vis. Sci. 2016;57(12):4491.
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© ARVO (1962-2015); The Authors (2016-present)
Mutation of the gene RPE65 leads to disruption of retinoid metabolism and Type 2 Leber congenital amaurosis (LCA2), a form of retinal dystrophy characterized by early-onset cone death and progressive visual impairment. We previously shown that GFP+ dendritic cells (DC) were recruited at an early age to the outer retina of Rpe65-/- mice bred to CD11c-DTR/GFP background. The trigger for recruitment remains unclear. We investigate the timing, localization and persistence of DC recruitment to the outer retina in relation to the well-studied cone death cycle in this animal model to deduce its mechanism.
To develop the animal model, Rpe65-/- mice were bred to transgenic mice whose DC expressed a chimeric protein containing GFP and diphtheria toxin receptor driven by a CD11c promoter. Progeny were screened by PCR, flow cytometry, and immunofluorescence to confirm transgenic Rpe65-/- mice along with Rpe65+/+ mice as controls. Cone survival was evaluated by counts from retinal flatmounts stained for cone opsins, and morphology was evaluated by cryosections and retinal flatmounts. Eyecups were prepared and analyzed using flow cytometry to quantify GFP+ DC at various ages between different groups.
Morphology of cross sections and retinal flatmounts confirm that GFP+ DC are recruited to the outer retina, where majority residing in the OPL and extend processes to inner and outer segment layers. Furthermore, flow cytometry at 1 wk of age showed similarly elevated levels in the retinas of both Rpe65-/- and control groups. This elevated level decreased significantly by 2 weeks of age in the control but remained elevated in the Rpe65-/- group, a time when the cone densities between the two groups were similar. Analysis from 4 weeks all the way to 40 weeks of age confirmed that Rpe65-/- mice maintained a significantly elevated level of GFP+ DC in the retina compared to controls, and that this was observed far after the majority of cones had already died.
These results show that the GFP+ DC were initially recruited to the outer retina regardless of Rpe65-/- status; however, significantly elevated DC remained in Rpe65-/- retinas far beyond the first month of age where the majority of cones have already died. This suggests that other factors, possibly rod degeneration in Rpe65-/- mice, may play a key role in recruiting and maintaining DC to the outer retina.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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