DMD-Q995* Mice

Catalog Number: C001518

Strain Name: C57BL/6JCya-Dmdem1(Q995X)/Cya

Genetic Background: C57BL/6JCya

Reproduction: Homozygous female × Heterozygous male

Strain Description

Duchenne muscular dystrophy (DMD) is a severe, progressive, and debilitating X-linked disorder characterized by muscle wasting. This condition precipitates difficulties with movement, eventually necessitating assisted ventilation, and often leads to premature death. The primary cause of DMD is mutations in the dystrophin muscular dystrophy (DMD) gene, which is responsible for encoding the dystrophin protein. These mutations effectively eliminate the production of dystrophin protein in muscle tissues, instigating muscle atrophy and a myriad of complications [1]. The absence of dystrophin protein culminates in the disintegration of the dystrophin-associated protein complex (DAPC) within the muscle membrane. This disintegration disrupts the interaction between actin and the extracellular matrix, rendering muscles devoid of dystrophin more susceptible to damage. This susceptibility results in the progressive loss of muscle tissue and function, as well as the development of cardiomyopathy [2].

DMD-Q995* mice carry a c.2983C>T (p.Q995) mutation in the Dmd gene, which results in the production of a premature termination codon (PTC). In eukaryotes, the nonsense-mediated mRNA decay (NMD) pathway degrades mRNAs containing PTCs to reduce errors in gene expression. This is because these abnormal mRNAs may encode harmful gain-of-function or dominant-negative proteins that can damage normal human physiological mechanisms. In DMD-Q995* mice, the mutation and the NMD pathway together result in the degradation of most Dmd transcripts. The remaining transcripts can only encode truncated dystrophin proteins that lack normal function, leading to the loss of dystrophin function [3-5]. This model, due to the lack of normal dystrophin expression, exhibits a series of muscle disease phenotypes similar to the clinical presentation of Duchenne muscular dystrophy (DMD), and can be used for research on DMD. Homozygous female mice and heterozygous males of this strain are viable and fertile.

Strain Strategy

The c.2983C>T mutation was introduced into the mouse Dmd gene using gene editing technology.


Application

This model is a valuable tool for studying the mechanisms of DMD and for screening potential therapeutic agents.

Validation Data

1. Detection of Dmd transcripts (mRNA)


Figure 1. Detection of Dmd transcripts in tissues of 9-week-old DMD-Q995* mice and wild-type (WT) mice. The qRT-PCR* analysis of Dmd gene expression indicates that the presence of premature termination codons (PTC) in the homozygous DMD-Q995* mice leads to the degradation of some Dmd transcripts. Compared to the wild-type (WT) mice, the expression of Dmd in various tissues has decreased to varying degrees, indicating the abnormal expression of the Dmd gene in this model.

*The forward and reverse primers used for detection were both located downstream of the mutation site.

2. Detection of dystrophin protein expression


Figure 2. Detection of dystrophin expression in the brain, heart, and skeletal muscle of 7-week-old DMD-Q995* mice and wild-type mice. Western blotting analysis of dystrophin expression in mice revealed that dystrophin was not expressed in the brain, heart, or skeletal muscle of DMD-Q995* mice.

*The antibody used for the assay was a recombinant anti-dystrophin antibody (Abcam, ab218198).

3. H&E staining of muscle tissue


Figure 3. Histological analysis of muscle tissue from DMD-Q995* mice and wild-type (WT) mice. H&E staining of muscle tissues from homozygous DMD-Q995* mice revealed variable muscle fiber size, nuclei aggregation, and inflammatory cell infiltration, which were not observed in wild-type mice. 

4. Detection of serum creatine kinase (CK) level


Figure 4. Serum creatine kinase (CK) levels in 9-week-old DMD-Q995* mice and wild-type (WT) mice. Creatine kinase is a component of the cardiac enzyme panel, and an increase in serum creatine kinase is associated with muscle damage and myocardial damage. The results showed that the activity of creatine kinase in DMD-Q995* mice was significantly higher than that in wild-type mice, indicating that the muscle tissue of DMD-Q995* mice was damaged. In addition, the CK values of male DMD-Q995* mice were significantly higher than those of female DMD-Q995* mice, indicating that the muscle tissue damage in males was more severe than that in females, which is consistent with the conclusions of previous studies [6-8].

5. Rotarod 


Figure 5. Rotarod test of DMD-Q995* mice and wild-type mice. The test results show that compared to the wild-type mice, the latency to fall in the rotarod test of DMD-Q995* mice significantly shortened from the age of 6 weeks, indicating a defect in their motor coordination ability.

6. Grip Strength 


Figure 6. Grip strength test of DMD-Q995* mice and wild-type mice. The test results show that the grip strength of DMD-Q995* mice is significantly weaker compared to the wild-type mice, indicating that their limb strength is impaired due to muscle tissue damage.

7. Treadmill Study


Figure 7. Treadmill test of DMD-Q995* mice and wild-type mice. The test results show that compared to the wild-type mice, the total distance traveled (Distance Traveled) in the treadmill test of DMD-Q995* mice is significantly shortened, suggesting a decline in their exercise capacity and endurance. At the same time, the number of shocks (The Shock Times) has significantly increased, indicating defects in their motor coordination and learning ability. In addition, the latency (Latency) of DMD-Q995* mice is shorter, which may indicate a higher level of fatigue.

8. Gait Analysis


Figure 8. Gait analysis of DMD-Q995* mice and wild-type mice. The test results show that compared to the wild-type mice, the number of steps (Number of Steps) of DMD-Q995* mice increased, and the walking time (Walking Time) extended.

9. Conclusion

The DMD-Q995* mouse model demonstrates significant deficits in motor function and coordination. The decline in grip strength and reduced latency on the rotarod indicate impaired neuromuscular coordination and endurance. Additionally, reduced distance traveled, increased shock occurrences and shortened latency to the first shock highlight compromised locomotor activity and responsiveness. Alterations in gait patterns, including an increased number of steps and prolonged walking time, further support the presence of motor dysfunction. These findings collectively suggest that the DMD-Q995* mutation leads to progressive impairments in motor abilities, offering a robust model for studying neuromuscular disorders and potential therapeutic interventions.


 

References
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[3]Hoffman EP, Brown RH Jr, Kunkel LM. Dystrophin: the protein product of the Duchenne muscular dystrophy locus. Cell. 1987 Dec 24;51(6):919-28.
[4]Cox GA, Phelps SF, Chapman VM, Chamberlain JS. New mdx mutation disrupts expression of muscle and nonmuscle isoforms of dystrophin. Nat Genet. 1993 May;4(1):87-93.
[5]Sicinski P, Geng Y, Ryder-Cook AS, Barnard EA, Darlison MG, Barnard PJ. The molecular basis of muscular dystrophy in the mdx mouse: a point mutation. Science. 1989 Jun 30;244(4912):1578-80.
[6]Hermes TA, Kido LA, Macedo AB, Mizobuti DS, Moraes LHR, Somazz MC, Cagnon VHA, Minatel E. Sex influences diaphragm muscle response in exercised mdx mice. Cell Biol Int. 2018 Dec;42(12):1611-1621.
[7]Yoshida M, Yonetani A, Shirasaki T, Wada K. Dietary NaCl supplementation prevents muscle necrosis in a mouse model of Duchenne muscular dystrophy. Am J Physiol Regul Integr Comp Physiol. 2006 Feb;290(2):R449-55.
[8]Salimena MC, Lagrota-Candido J, Quírico-Santos T. Gender dimorphism influences extracellular matrix expression and regeneration of muscular tissue in mdx dystrophic mice. Histochem Cell Biol. 2004 Nov;122(5):435-44.