Learning, memory and blood-brain barrier pathology in Duchenne muscular dystrophy mice lacking Dp427, or Dp427 and Dp140.

Duchenne muscular dystrophy is a severe neuromuscular disorder that is caused by mutations in the DMD gene, resulting in a disruption of dystrophin production. Next to dystrophin expression in the muscle, different isoforms of the protein are also expressed in the brain and lack of these isoforms leads to cognitive and behavioral deficits in patients. It remains unclear how the loss of the shorter dystrophin isoform Dp140 affects these processes. Using a variety of behavioral tests, we found that mdx and mdx4cv mice (which lack Dp427 or Dp427 + Dp140, respectively) exhibit similar deficits in working memory, movement patterns and blood-brain barrier integrity. Neither model showed deficits in spatial learning and memory, learning flexibility, anxiety or spontaneous behavior, nor did we observe differences in aquaporin 4 and glial fibrillary acidic protein. These results indicate that in contrast to Dp427, Dp140 does not play a crucial role in processes of learning, memory and spontaneous behavior.

Low human dystrophin levels prevent cardiac electrophysiological and structural remodelling in a Duchenne mouse model.

Duchenne muscular dystrophy (DMD) is a progressive neuromuscular disorder caused by loss of dystrophin. This lack also affects cardiac structure and function, and cardiovascular complications are a major cause of death in DMD. Newly developed therapies partially restore dystrophin expression. It is unclear whether this will be sufficient to prevent or ameliorate cardiac involvement in DMD. We here establish the cardiac electrophysiological and structural phenotype in young (2-3 months) and aged (6-13 months) dystrophin-deficient mdx mice expressing 100% human dystrophin (hDMD), 0% human dystrophin (hDMDdel52-null) or low levels (~ 5%) of human dystrophin (hDMDdel52-low). Compared to hDMD, young and aged hDMDdel52-null mice displayed conduction slowing and repolarisation abnormalities, while only aged hDMDdel52-null mice displayed increased myocardial fibrosis. Moreover, ventricular cardiomyocytes from young hDMDdel52-null animals displayed decreased sodium current and action potential (AP) upstroke velocity, and prolonged AP duration at 20% and 50% of repolarisation. Hence, cardiac electrical remodelling in hDMDdel52-null mice preceded development of structural alterations. In contrast to hDMDdel52-null, hDMDdel52-low mice showed similar electrophysiological and structural characteristics as hDMD, indicating prevention of the cardiac DMD phenotype by low levels of human dystrophin. Our findings are potentially relevant for the development of therapeutic strategies aimed at restoring dystrophin expression in DMD.

Cross-sectional study into age-related pathology of mouse models for limb girdle muscular dystrophy types 2D and 2F.

Limb girdle muscular dystrophy (LGMD) types 2D and 2F are caused by mutations in the genes encoding for α- and δ-sarcoglycan, respectively, leading to progressive muscle weakness. Mouse models exist for LGMD2D (Sgca-/-) and 2F (Sgcd-/-). In a previous natural history study, we described the pathology in these mice at 34 weeks of age. However, the development of muscle pathology at younger ages has not been fully characterised yet. We therefore performed a study into age-related changes in muscle function and pathology by examining mice at different ages. From 4 weeks of age onwards, male mice were subjected to functional tests and sacrificed at respectively 8, 16 or 24 weeks of age. Muscle histopathology and expression of genes involved in muscle pathology were analysed for several skeletal muscles, while miRNA levels were assessed in serum. In addition, for Sgcd-/- mice heart pathology was assessed. Muscle function showed a gradual decline in both Sgca-/- and Sgcd-/- mice. Respiratory function was also impaired at all examined timepoints. Already at 8 weeks of age, muscle pathology was prominent, and fibrotic, inflammatory and regenerative markers were elevated, which remained relatively constant with age. In addition, Sgcd-/- mice showed signs of cardiomyopathy from 16 weeks of age onwards. These results indicate that Sgca-/- and Sgcd-/- are relevant disease models for LGMD2D and 2F.

Natural disease history of the D2-mdx mouse model for Duchenne muscular dystrophy.

The C57BL/10ScSn-Dmdmdx/J (BL10-mdx) mouse has been the most commonly used model for Duchenne muscular dystrophy (DMD) for decades. Their muscle dysfunction and pathology is, however, less severe than in patients with DMD, which complicates preclinical studies. Recent discoveries indicate that disease severity is exacerbated when muscular dystrophy mouse models are generated on a DBA2/J genetic background. Knowledge on the natural history of animal models is pivotal for high-quality preclinical testing. However, for BL10-mdx mice on a DBA2/J background (D2-mdx), limited data are available. We addressed this gap in the natural history knowledge. First, we compared histopathological aspects in skeletal muscles of young D2-mdx, BL10-mdx, and wild-type mice. Pathology was more pronounced in D2-mdx mice and differed in severity between muscles within individuals. Secondly, we subjected D2-mdx mice to a functional test regime for 34 weeks and identified that female D2-mdx mice outperform severely impaired males, making females less useful for functional preclinical studies. Direct comparisons between 10- and 34-wk-old D2-mdx mice revealed that disease pathology ameliorates with age. Heart pathology was progressive, with some features already evident at a young age. This natural history study of the D2-mdx mouse will be instrumental for experimental design of future preclinical studies.-Van Putten, M., Putker, K., Overzier, M., Adamzek, W. A., Pasteuning-Vuhman, S., Plomp, J. J., Aartsma-Rus, A. Natural disease history of the D2-mdx mouse model for Duchenne muscular dystrophy.

A modified diet does not ameliorate muscle pathology in a mouse model for Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is caused by a lack of dystrophin protein. Next to direct effects on the muscles, this has also metabolic consequences. The influence of nutrition on disease progression becomes more and more recognized. Protein intake by DMD patients may be insufficient to meet the increased demand of the constantly regenerating muscle fibers. This led to the hypothesis that improving protein uptake by the muscles could have therapeutic effects. The present study examined the effects of a modified diet, which composition might stimulate muscle growth, on disease pathology in the D2-mdx mouse model. D2-mdx males were fed with either a control diet or modified diet, containing high amounts of branched-chain amino acids, vitamin D3 and ursolic acid, for six weeks. Our study indicates that the modified diet could not ameliorate the muscle pathology. No effects on bodyweight or weight of individual muscles were observed. Neither did the diet affect severity of fibrosis or calcification of the muscles.

Voluntary exercise improves muscle function and does not exacerbate muscle and heart pathology in aged Duchenne muscular dystrophy mice.

Duchenne muscular dystrophy is a severe muscle wasting disease, characterized by a severely reduced lifespan in which cardiomyopathy is one of the leading causes of death. Multiple therapies aiming at dystrophin restoration have been approved. It is anticipated that these therapies will maintain muscle function for longer and extend the ambulatory period, which in turn will increase the cardiac workload which could be detrimental for cardiac function. We investigated the effects of voluntary running exercise in combination with low dystrophin levels on function and pathology of skeletal muscle and heart. We divided 15.5-month old female mdx (no dystrophin), mdx-XistΔhs (varying low dystrophin levels) and wild type mice (BL10-WT and XistΔhs-WT) to either a sedentary or voluntary wheel running regime and assessed muscle function at 17.5 months of age. Thereafter, a cardiac MRI was obtained, and muscle and heart histopathology were assessed. We show that voluntary exercise is beneficial to skeletal muscle and heart function in dystrophic mice while not affecting muscle pathology. Low amounts of dystrophin further improve skeletal muscle and cardiac function. These findings suggest that voluntary exercise may be beneficial for skeletal muscle and heart in DMD patients, especially in conjunction with low amounts of dystrophin.