Abstract
Cataracts are the leading cause of vision loss in the world, although surgical treatment can restore vision in cataract patients. Until now, there have been no adequate animal models for in vivo studies of artificial lens safety and drug interactions. Genetic studies have demonstrated that GJA8 is involved in maintaining lens opacity and proper lens development. In this study, a cataract model with GJA8 gene knockout was developed via co-injection of Cas9/sgRNA mRNA into rabbit zygotes. Our results showed that gene mutation efficiency in the GJA8 locus reached 98.7% in embryos and 100% in pups, demonstrating that the Cas9/sgRNA system is a highly efficient tool for gene editing in rabbits. In agreement with other studies, our genetic and histology results showed that impaired GJA8 function caused microphthalmia, small lens size and cataracts. In summary, our novel rabbit model of cataracts will be an important drug-screening tool for cataract prevention and treatment.
Highlights
Cataracts are the leading cause of vision loss in the world, surgical treatment can restore vision in cataract patients
GJA8 encodes connexin[50] (Cx50) and has abundant expression in the lens, which is necessary for lens growth and maturation of lens fiber cells[10,11]
CRISPR/Cas9-mediated gene targeting of GJA8 in zygotes
Summary
Cataracts are the leading cause of vision loss in the world, surgical treatment can restore vision in cataract patients. A cataract model with GJA8 gene knockout was developed via co-injection of Cas9/sgRNA mRNA into rabbit zygotes. The lens specific gap junction proteins GJA1, GJA3 and GJA8 play critical roles in transmitting information from highly metabolically active cells to less metabolically active cells These are critical genes in development and function of vertebrate lenses[5]. Because rabbits and humans are similar, the objective of this study was to create a novel animal model that recapitulated human congenital cataracts by using CRISPR/Cas9-mediated GJA8 gene knockout in rabbits. To evaluate this cataract model, we examined the gene editing efficiency of CRISPR/Cas[9], the phenotypes and mutant gene heritability
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