Figure 4.

Long-term silencing of HTT with tetra-siRNA^Htt

(A) Fluorescent fundus images and retinal cross-section of mice injected with 15 μg of the tetra-siRNA^Htt and the tetra-siRNA^NTC imaged at 3 months postinjection. (B) Retinal cross-sections at 3 months postinjection stained for expression of the HTT protein (purple signal). Shown is an entire cross-section (left) and a magnified view (right). Reduction of HTT protein in PR cell ISs is indicated by black arrowheads. (C and D) Retinal cross-sections stained for GFAP (red signal) and Iba1 (red signal) at 3, 6, and 9 months postintravitreal injection of 15 μg siRNAs to visualize the activation of gliosis and migration of microglia, respectively. (E) Quantification of remaining retinal HTT protein over time after 1 intravitreal injection of 15 μg siRNAs. Top shows example of protein bands on western blot and bottom shows quantification. Shown is the percentage of remaining HTT protein compared to the NTC for each time point. N = 6–7 retinas per time point and siRNA type; error bars: SD. ∗∗∗p < 0.001. (A, C, and D) Blue: nuclei marked with DAPI; green: cone segments marked with peanut agglutinin lectin (PNA); red: siRNAs in (A) marked by Cy3 label and GFAP, and by Iba1 in (C) and (D). In (A), half of the blue and green was removed to better visualize the red signal. Scale bars: 50 μm, except in (B), as indicated. Histology in (A)–(D) was repeated with at least N = 3 retinas.