To characterize recombinant AAV2 （rAAV2）-mediated expression of L 132C/T 159C ChR2 mutant in retinal ganglion cells （RGCs） of young adult cynomolgus monkeys, rAAV2 vectors carrying a fusion construct of the ChR2 m...To characterize recombinant AAV2 （rAAV2）-mediated expression of L 132C/T 159C ChR2 mutant in retinal ganglion cells （RGCs） of young adult cynomolgus monkeys, rAAV2 vectors carrying a fusion construct of the ChR2 mutant and GFP （ChR2-GFP） were delivered to the vitreous chamber by intravitreal injection. Expression patterns of the ChR2 mutant in RGCs were examined by immunohistochemical methods three months after injection. The RNA-binding protein with multiple splicing （RBPMS） was used as an RGC specific marker to differentiate RGCs from other retinal neurons and non-neuronal cells. The numbers of RBPMS＋ and GFP＋ double-labeled RGCs in the central foveal varied with the eccentricity. The expression peaked within 100 p.m from the edge of the foveola and drastically decreased to a single superficial RGC layer approximately 300 ~tm from the edge. On average, the ratio of the double-labeled RGCs versus RBPMS＋ RGCs approached 0.324-0.15 （n=14 fields） at the central foveal region （0.1 to 0.53 mm）. We observed that the ratio reached 0.784-0.16 （n=21 fields） at peripheral retinal locations （eccentricity 〉7 mm）. This investigation demonstrates that RBPMS could serve as a valuable RGC specific marker for future investigations in this field.展开更多
基金supported by National Science Foundation of China(31571091 to Mingliang Pu),National Basic Research Program of China (2015CB351806 to Mingliang Pu)National Institutes of Health Grant (NIH) (EY17130 to Zhuo-Hua Pan)Dryer Foundation, the Ligon Research Center of Vision, and Research to Prevent Blindness to Department of Ophthalmology at Wayne State University
文摘To characterize recombinant AAV2 （rAAV2）-mediated expression of L 132C/T 159C ChR2 mutant in retinal ganglion cells （RGCs） of young adult cynomolgus monkeys, rAAV2 vectors carrying a fusion construct of the ChR2 mutant and GFP （ChR2-GFP） were delivered to the vitreous chamber by intravitreal injection. Expression patterns of the ChR2 mutant in RGCs were examined by immunohistochemical methods three months after injection. The RNA-binding protein with multiple splicing （RBPMS） was used as an RGC specific marker to differentiate RGCs from other retinal neurons and non-neuronal cells. The numbers of RBPMS＋ and GFP＋ double-labeled RGCs in the central foveal varied with the eccentricity. The expression peaked within 100 p.m from the edge of the foveola and drastically decreased to a single superficial RGC layer approximately 300 ~tm from the edge. On average, the ratio of the double-labeled RGCs versus RBPMS＋ RGCs approached 0.324-0.15 （n=14 fields） at the central foveal region （0.1 to 0.53 mm）. We observed that the ratio reached 0.784-0.16 （n=21 fields） at peripheral retinal locations （eccentricity 〉7 mm）. This investigation demonstrates that RBPMS could serve as a valuable RGC specific marker for future investigations in this field.
