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The cellular and molecular consequences of lack of dystrophin in humans are only partially known, which is crucial for the development of new therapies aiming to slow or stop the progression Duchenne and Becker muscular dystrophies. We analyzed muscle biopsies of DMD patients and controls using single nuclei RNA sequencing (snRNAseq) and correlated the results with clinical data. DMD samples displayed an increase in regenerative fibers, satellite cells and fibro-adipogenic progenitor cells (FAPs) and a decrease in slow fibers and smooth muscle cells. Samples from patients with stable mild weakness were characterized by an increase in regenerative fibers, while those from patients with progressive weakness had fewer muscle fibers and increased FAPs. DMD muscle fibers displayed a strong regenerative signature, while DMD FAPs upregulated genes producing extracellular matrix and molecules involved in several signaling pathways. An analysis of intercellular communication profile identified FAPs as a key regulator of cell signaling in DMD samples. We show significant differences in the gene expression profiled of the different cell populations present in DMD muscle samples compared to controls.

PurposeGreater muscle fragility is thought to cause the exhaustion of the muscle stem cells during successive degeneration/repair cycles, leading to muscle wasting and weakness in Duchenne muscular dystrophy. Chronic voluntary exercise can partially reduce the susceptibility to contraction induced-muscle injury, i.e., muscle fragility, as shown by a reduced immediate maximal force drop following lengthening contractions, in the dystrophic mdx mice. Here, we studied the effect of Prospero-related homeobox factor 1 gene (Prox1) transfer (overexpression) on fragility in chronically exercised mdx mice, because Prox1 promotes slower type fibres in healthy mice and slower fibres are less fragile in mdx muscle.Methodsmdx mice received or not Prox1 transfer into the tibialis anterior muscle and performed voluntary running into a wheel during 1 month. We also performed Prox1 transfer in sedentary mdx mice. In situ maximal force production of the muscle in response to nerve stimulation was assessed before, during and after 10 lengthening contractions. Molecular and cellular parameters were also evaluated.ResultsInterestingly, Prox1 transfer reduced the force drop following lengthening contractions in exercised mdx mice (p < 0.05 to 0.01), but not in sedentary mdx mice. It also increased the muscle expression of Myh7 (p < 0.001), MHC-2x (p < 0.01) and Trpc1 (p < 0.01), whereas it reduced that one of Myh4 (p < 0.001) and MHC-2b (p < 0.01) in exercised mdx mice. Moreover, Prox1 transfer decreased the maximal force (p < 0.01) before lengthening contraction in exercised mdx mice (p < 0.01), and reduced muscle weight (p < 0.0001) despite increased Mstn expression (p < 0.001).ConclusionOur results indicate that the beneficial effect of Prox1 transfer on muscle fragility is only observed in chronically exercised mdx mice. Thus, Prox1 transfer combined to chronic exercise have the potential to substantially slow the progression of the dystrophic disease in the long term.

Background Voluntary exercise can improve skeletal muscle fragility, i.e. higher susceptibility to contraction induced-injury, as shown by a greater force drop following lengthening contractions, in the dystrophic Mdx mice as compared to healthy mice with dystrophin. This beneficial effect is related to the activation of the calcineurin activation. Unfortunately, voluntary running only partly rescued fragility, so it would be interesting to combined the effects of exercise, for example, with those of others treatments activating the calcineurin pathway and promoting slow and more oxidative fibres. This is of particular interest because slow muscle fibres are apparently less affected and genetic or pharmacological treatments promoting slow and more oxidative fibres are been shown to be beneficial in the Mdx mice. Methods Here, we tested whether voluntary exercise (1 month of running in a wheel) combined with Prospero-related homeobox factor 1 gene ( Prox1) transfer would better improve functional dystrophic features in Mdx mice as compared to the voluntary exercise single approach. Prox1 is known to promote the promotion of slow contractile gene program in healthy muscle. Results We found that Prox1 transfer promoted slower molecular and functional contractile features in both voluntary exercised and sedentary Mdx mice. However, it improved fragility only in exercised Mdx mice. Moreover, Prox1 transfer reduced absolute maximal force production by causing reduction in muscle weight in both exercised and sedentary Mdx mice. Conclusion In conclusion, our results indicate that the beneficial effects of voluntary exercise and Prox1 transfer on fragility are additive in Mdx mice.