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Wei Du, Wen-tao Deng, Sanford L Boye, Jie Li, Wei Li, William W Hauswirth, Ji-Jing Pang; AAV-mediated Gene Therapy Restores M-Cone Function in S-opsin only Thrb Knockout Mouse. Invest. Ophthalmol. Vis. Sci. 2014;55(13):3318.
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Color vision is facilitated by cones expressing different opsin photopigments. M-cones are sensitive to middle (M, ‘green’) and S-cones short (S,‘blue’) wavelengths. Thyroid hormone receptor β2 (TRβ2) is a ligand-activated transcription factor that is expressed in the outer nuclear layer of the embryonic retina. Thrb (encoding Trβ2) knockout (KO) mice show selective loss of M-opsin and a concomitant increase in S-opsin. Therefore the Thrb KO mouse provides a model for the study of human Blue Cone Monochromacy. We therefore tested whether AAV-mediated M-opsin gene therapy targeting S-opsin only cones can restore M-cone system function/structure in this model.
At postnatal day 14 (P14), one μl of AAV8 (Y447, 733)-IRBP/GNAT2-M-opsin vector (1013 particles/ml) was injected subretinally into one eye of each TRb2 KO mouse. The other eye was uninjected as a control. M-cone mediated ERGs were recorded at 8 weeks after injection. Treated and control eyes were harvested for immunohistochemical studies.
Eight weeks after treatment, nearly normal dark-adapted ERG waveforms were recorded in either treated or untreated Thrb KO eyes with a mild reduction of ERG amplitudes in treated eyes, perhaps due to subretinal injection-related damage. Cone mediated light-adapted ERGs were recorded from either treated or untreated eyes. S-cone mediated ERGs were also recorded from either treated or untreated eyes. M-cone mediated ERGs were only seen in treated Thrb KO eyes, at about 20% of normal amplitudes. Cone-opsin staining showed that robust S-opsin was present in both treated and untreated eyes whereas no M-opsins were evident in untreated eyes. In contrast, in treated eyes M-opsin positive cells were seen, which were shown to be cones by cone-specific PNA staining.
Since Thrb KO mouse has only S-opsin, it can be considered a model for human Blue Cone Monochromacy. AAV8-IRBP/GNAT2-M-opsin mediated gene therapy restores M-cone function in Thrb KO mouse. These results serve as a baseline for studying long-term retinal rescue in Thrb KO mice.
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