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Astra Dinculescu, Frank M Dyka, Seok-Hong Min, Rachel M Stupay, W. Clay Smith, William W Hauswirth; RPE targeted overexpression of mutant S163R C1QTNF5 in various mouse models of RPE/photoreceptor dysfunction. Invest. Ophthalmol. Vis. Sci. 2017;58(8):5229.
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© ARVO (1962-2015); The Authors (2016-present)
The S163R mutation in complement C1qTNF5 causes an autosomal dominant late-onset retinal degeneration (L-ORD) disorder, which shares several key features with AMD, including the presence of extracellular deposits between the RPE and choroid. We have previously shown that the S163R mutant exhibits a reversely polarized distribution, being routed towards the basal rather than apical RPE, where it forms thick deposits in-vivo. Here, we extend our previous study and characterize the behavior of the S163R mutant in mouse models of retinal degeneration following its AAV-mediated expression in the RPE.
We generated scAAV vectors (AAV2 quad YF) containing the mutant S163R C1QTNF5 tagged with HA epitope, driven by an RPE-specific BEST1 promoter (1012 vector genomes/mL). The viral vectors were injected subretinally into one eye of mice undergoing retinal degeneration, including rd10, rd12, ABCA4 KO and wild-type A/J. Retinal function was assessed periodically by full-field electroretinography (ERG) under scotopic and photopic conditions. The eyes were also examined by other non-invasive imaging methods, including digital fundus imaging and spectral-domain optical coherence tomography (SD-OCT). Transgene expression was detected by immunohistochemistry using an anti-HA antibody and by Western blotting. Hematoxylin and eosin (H & E) staining as well as various antibody markers for RPE and photoreceptor proteins were used to examine the morphology of photoreceptor/RPE/Bruch’s membrane/Choroid interface, and the associated extracellular S163R C1QTNF5 deposits.
The RPE-targeted overexpression of S163R C1QTNF5 mutant in all mice led to the accumulation of the S163R mutant as thick extracellular deposits between the RPE and choroid, independent of the mouse strain or the underlying pre-existing RPE or photoreceptor dysfunction analyzed. The presence of basal S163R deposits affected not only the RPE and photoreceptor cells, based on morphological and ERG analysis, but also the underlying choriocapillaris.
Every AAV-S163R injected mouse developed widespread RPE basal laminar deposits in a reliable, consistent manner. This system offers a unique opportunity to uncover pathological mechanisms associated with the extracellular basal deposit formation, and may represent a novel platform for developing therapeutic strategies for AMD and L-ORD.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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