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Daniyar L Dauletbekov, Alun R Barnard, Michelle McClements, Dominik M Fischer, Robert E MacLaren; Retention of cone function in a murine rhodopsin knock-out model of retinitis pigmentosa by rhodopsin gene delivery. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):244.
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© ARVO (1962-2015); The Authors (2016-present)
Subretinal delivery of wild type rhodopsin via adeno-associated virus (AAV) vectors provides structural and functional benefit to rod degeneration in different models of rhodopsin-linked retinitis pigmentosa (RP), however its impact on secondary cone degeneration has not yet been described. We hypothesized that the supplementation of wild type rhodopsin gene could slow down secondary cone loss in a murine model of RP lacking endogenous rhodopsin.
Homozygous rhodopsin knockout mice (rho-/-) were used in this study. Subretinal injection (2.0 ul containing 1.2E+9 genome copies) of a self complementary AAV vector encoding human rhodopsin driven by the human rhodopsin promoter - scAAV2/8.hRHOp.hRHO was administered at postnatal day 21 (P21). The contralateral eye in all mice received an equivalent volume and titer of sham injection of AAV2/8.control which didn’t encode for any protein to control for the effects of surgical intervention. Immunohistochemistry with rhodopsin staining was performed at P50 to confirm rhodopsin expression. Dark and light adapted electroretinography (ERG) at increasing flash intensities according to a standardized protocol were performed at P50 and P80 (2 month post-injection). In the rho-/- model light adapted ERG responses (reflecting cone function) are virtually extinct by P80, so retention of these responses at P80 was used as the primary outcome measure for this study.
Immunohistochemistry at P50 showed the presence of clearly defined rod outer segments expressing rhodopsin in the treated group and absence of rhodopsin/rod outer segments in the control group. At P80 average b-waves of light adapted ERG (only cones provide ERG responses at this type of ERG, as rod responses are supressed) at increasing flash intensities between 0.5 to 25 cd.s/m-2 were significantly higher in the treatment group (M = 69.74 uV; SE = 9.87; 95% CI = 47.41- 92.08) comparing to the control group (M = 25.15 uV; SE = 4.34 uV; 95% CI = 13.31 - 32.99; repeated measures 2 way ANOVA, F(2, 9) = 35.396, p = 0.0002; n = 10).
The preserved cone ERG responses confirmed our hypothesis of a functional benefit of delivering rhodopsin gene on secondary cone degeneration in a rhodopsin knock-out murine model of RP. The study represents potential progress for gene therapy addressing other forms of RP, where cone loss occurs secondary to rod degeneration.
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