Investigative ophthalmology & visual science · 2025
Novel Grm6 Variant in a no b-wave (nob) Mouse Model: Phenotype Characterization and Gene Therapy.
СтатьяПоддержка NIH: внешняя программаПоддержка исследования: вне правительства СШАНазвание и аннотация: Перевод в очереди
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Lin Pei-Hsuan, Kang Eugene Yu-Chuan, Makrides Neoklis, Lee Winston, Tseng Yun-Ju, Wu Pei-Liang, Peregrin John, Sherman Emmet, Wang Jason Hunghsuan, Swayne Theresa C, Li Tai-De, Agosto Melina A, Sparrow Janet R, Zhang Xin, Tsang Stephen H, Wang Nan-Kai
Аннотация
PURPOSE: To characterize a no b-wave (nob) mouse model of congenital stationary night blindness (CSNB) caused by a Grm6 variant that disrupts photoreceptor-to-bipolar cell signaling. Additionally, we aim to evaluate the efficacy of gene therapy in restoring visual function.
METHODS: The nob mouse was generated through selective breeding to regenerate the nob phenotype. Adeno-associated viruses encoding Grm6 and GFP, driven by two promoters (hGRM6 and CMV), were administered to nob mice at postnatal days 5 (P5) and 30 (P30), respectively. Electroretinography and spectral domain optical coherence tomography (SD-OCT) were conducted three months after gene therapy.
RESULTS: The nob phenotype was successfully regenerated, and a homozygous missense variant c.1037G>A (p.Arg346His) in Grm6 was identified as the causal variant. Scotopic b waves were absent, whereas a waves remained normal, indicating intact rod function but impaired bipolar cell function. SD-OCT revealed thinning of the retinal nerve fiber layer and outer plexiform layer (OPL) in affected mice. Immunofluorescence and immunoblotting revealed decreased mGluR6 levels and associated signaling proteins. Gene therapy restored mGluR6 expression and reestablished synaptic protein localization in the OPL, although improvements in b/a ratios and OPL thickness were modest. Notably, the hGRM6 promoter at P5 was more effective at restoring OPL.
CONCLUSIONS: We identified a new nob mouse model that mimics the CSNB phenotype in human patients. Whereas gene therapy successfully restored mGluR6 expression, functional improvements were limited. Early treatment using a specific promoter is critical, and increasing transduction efficiency may improve gene therapy strategies.
METHODS: The nob mouse was generated through selective breeding to regenerate the nob phenotype. Adeno-associated viruses encoding Grm6 and GFP, driven by two promoters (hGRM6 and CMV), were administered to nob mice at postnatal days 5 (P5) and 30 (P30), respectively. Electroretinography and spectral domain optical coherence tomography (SD-OCT) were conducted three months after gene therapy.
RESULTS: The nob phenotype was successfully regenerated, and a homozygous missense variant c.1037G>A (p.Arg346His) in Grm6 was identified as the causal variant. Scotopic b waves were absent, whereas a waves remained normal, indicating intact rod function but impaired bipolar cell function. SD-OCT revealed thinning of the retinal nerve fiber layer and outer plexiform layer (OPL) in affected mice. Immunofluorescence and immunoblotting revealed decreased mGluR6 levels and associated signaling proteins. Gene therapy restored mGluR6 expression and reestablished synaptic protein localization in the OPL, although improvements in b/a ratios and OPL thickness were modest. Notably, the hGRM6 promoter at P5 was more effective at restoring OPL.
CONCLUSIONS: We identified a new nob mouse model that mimics the CSNB phenotype in human patients. Whereas gene therapy successfully restored mGluR6 expression, functional improvements were limited. Early treatment using a specific promoter is critical, and increasing transduction efficiency may improve gene therapy strategies.