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Astra Dinculescu, Rachel M Stupay, Seok-Hong Min, Sanford L Boye, Vince Chiodo, William W Hauswirth; Clarin-1 expression in the inner retina: implications for gene therapy. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3632.
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© ARVO (1962-2015); The Authors (2016-present)
Mutations in clarin-1 (CLRN1) gene are the cause of Usher syndrome type III (USH3A), an autosomal recessive disorder, manifested as retinitis pigmentosa (RP) and progressive hearing loss. One previous study showed that Clrn1 is expressed in mouse photoreceptor cells. Here, we show that Clrn1 protein expression can also be detected in the inner retina. In addition, we describe the use rAAV vectors for studies aimed at developing therapeutic strategies for the treatment of RP caused by USH3A mutations.
Adult C57BL/6 mouse retinal sections were immunostained with a commercially available antibody directed against clarin-1, and visualized by immunofluorescence microscopy. Gene replacement therapy using intravitreal delivery of AAV-vectors expressing the mouse Clrn1 protein under the control of the ubiquitous CBA promoter were performed in 1 Month old Clrn1 knockout (KO) and wild-type control mice. Retinal function was assessed periodically by electroretinography (ERG) starting at 2 months post-injection. Transgene expression in AAV-treated eyes was detected by RT-PCR and Western Blotting.
Immunoreactivity for Clrn1 was detected in the inner retina, especially along the inner sublamina ending close to the ganglion cell layer in wild-type mice. The AAV-mediated gene-replacement strategy using an intravitreal approach resulted in a small, but significantly higher ERG amplitude in the Clrn1 KO group treated eyes relative to untreated partner eyes. Western blot analysis showed the presence of clarin protein tetramers similar to wild-type retinas in AAV-treated Clrn1 KO eyes but not in the contralateral controls.
Intravitreal delivery of clarin-1 using penetrating AAV vectors may represent a therapeutic strategy for the treatment of Ush3 retinal disorders, as it targets both the inner and outer retinal cells.
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