A novel rabbit model of Duchenne muscular dystrophy generated by CRISPR/Cas9

Abstract

Duchenne muscular dystrophy (DMD) is an X-linked muscle-wasting disorder caused by mutations in the dystrophin gene, with an incidence of 1 in 3500 in new male births. Mdx mice are widely used as an animal model for DMD. However, these mice do not faithfully recapitulate DMD patients in many aspects, rendering the preclinical findings in this model questionable. Although larger animal models of DMD, such as dogs and pigs, have been generated, usage of these animals is expensive and only limited to several facilities in the world. Here, we report the generation of a rabbit model of DMD by co-injection of Cas9 mRNA and sgRNA targeting exon 51 into rabbit zygotes. The DMD knockout (KO) rabbits exhibit the typical phenotypes of DMD, including severely impaired physical activity, elevated serum creatine kinase levels, and progressive muscle necrosis and fibrosis. Moreover, clear pathology was also observed in the diaphragm and heart at 5 months of age, similar to DMD patients. Echocardiography recording showed that the DMD KO rabbits had chamber dilation with decreased ejection fraction and fraction shortening. In conclusion, this novel rabbit DMD model generated with the CRISPR/Cas9 system mimics the histopathological and functional defects in DMD patients, and could be valuable for preclinical studies.This article has an associated First Person interview with the first author of the paper.

CRISPR-Generated Animal Models of Duchenne Muscular Dystrophy

“…Compared to pigs, rabbits are more suitable in terms of productivity and cost whilst maintaining greater similarity at the genetic, physiological, and anatomical levels to humans than mice [67,74,75]. Rabbits have a gestational duration of 31 days, the capacity to breed throughout the year, and have reduced space requirements than dogs or pigs [67].…”

“…Their intermediate size also provides the advantages that come with increased body size without much of the financial burden [76]. Sui et al (2018) recently developed a DMD rabbit model by co-injecting Cas9 mRNA and two gRNAs targeting DMD exon 51 into rabbit zygotes [75]. A total of 33 live rabbits were obtained out of 128 injected zygotes, 26 of which had at least one mutated DMD allele.…”

“…Four times higher serum CK levels were consistently observed in the mutants between 8-20 weeks of age. More importantly, unlike mdx mice that do not experience obvious cardiac dysfunction until 1 year of age [75,77], DMD rabbits show significantly reduced ejection fraction (EF) and fractional shortening (FS) compared to wild-type as early as 4 months old. Histological analysis of cardiac muscle revealed dystrophic symptoms, despite the lack of obvious differences in gross heart anatomy between mutant and healthy control rabbits.…”

Rabbit Biomodels of Human Diseases Developed Using New Genomic Technologies. Crispr/Cas9 (Review)

“…признаки, очень похожие на таковые у человека с DMD. Эффективность получения Dmd KO-кроликов была очень высокой: 78% F0 крольчат, из которых 85% были Dmd -/- [77].…”

Genome Editing for the Understanding and Treatment of Cardiomyopathies

“…In line with this, a related thrust in the field is to use CRISPR to develop other, more phenotypically representative DMD animal models. Rat [117], rabbit [118], and pig [119] models have already been developed, and all of these present with cardiac features reminiscent of DMD. These can be used for testing future DMD therapies, not just for genome editing, but also for those aiming to make improvements in the dystrophic heart.…”