Skeletal Ryanodine Receptors Are Involved in Impaired Myogenic Differentiation in Duchenne Muscular Dystrophy Patients

Abstract

Duchenne muscular dystrophy (DMD) is characterized by progressive muscle wasting following repeated muscle damage and inadequate regeneration. Impaired myogenesis and differentiation play a major role in DMD as well as intracellular calcium (Ca2+) mishandling. Ca2+ release from the sarcoplasmic reticulum is mostly mediated by the type 1 ryanodine receptor (RYR1) that is required for skeletal muscle differentiation in animals. The study objective was to determine whether altered RYR1-mediated Ca2+ release contributes to myogenic differentiation impairment in DMD patients. The comparison of primary cultured myoblasts from six boys with DMD and five healthy controls highlighted delayed myoblast differentiation in DMD. Silencing RYR1 expression using specific si-RNA in a healthy control induced a similar delayed differentiation. In DMD myotubes, resting intracellular Ca2+ concentration was increased, but RYR1-mediated Ca2+ release was not changed compared with control myotubes. Incubation with the RYR-calstabin interaction stabilizer S107 decreased resting Ca2+ concentration in DMD myotubes to control values and improved calstabin1 binding to the RYR1 complex. S107 also improved myogenic differentiation in DMD. Furthermore, intracellular Ca2+ concentration was correlated with endomysial fibrosis, which is the only myopathologic parameter associated with poor motor outcome in patients with DMD. This suggested a potential relationship between RYR1 dysfunction and motor impairment. Our study highlights RYR1-mediated Ca2+ leakage in human DMD myotubes and its key role in myogenic differentiation impairment. RYR1 stabilization may be an interesting adjunctive therapeutic strategy in DMD.

Calcium handling maturation and adaptation to increased substrate stiffness in human iPSC-derived cardiomyocytes: The impact of full-length dystrophin deficiency

“…This observation is also confirmed by the caffeineevoked Ca-Ts, which demonstrated a strong reduction of SR- Frontiers in Physiology frontiersin.org calcium content in DMD-hiPSC-CMs with respect to controls. In agreement, increased CaMKII activity and phosphorylation of RyR2 S2814 may be related to higher RyR open probability and to SR calcium leakage (Meyer et al, 2021) in DMD-hiPSC-CMs. Calcium leakage from the SR during diastole can partially account for the reduced Ca-T amplitude of DMD-hiPSC-CMs, as previously observed in the mdx mouse model (Ullrich et al, 2009;Ather et al, 2013;Kyrychenko et al, 2013).…”

Targeted transcript analysis in muscles from patients with genetically diverse congenital myopathies

“…On the other hand, the underlying cause leading to the expression of InsP3R may be linked to the lack of Ca 2+ signals originating from fewer or mutated RyR1 Ca 2+ channels which in turn may impact the muscle proteostatic machinery, leading to the aberrant expression of many proteins. Indeed, (i) in the absence of RyR1, mouse muscles aberrantly express >300 genes, including atypical expression of transcription factors, contractile proteins, muscle-specific structural proteins, and muscle regulatory factors 56 ; (ii) RYR1 silencing induces a decrease in myotube area and fusion index 57 ; (iii) pharmacologically blocking RyR1 activity inhibits in vitro differentiation of foetal myoblasts 58 ; (iv) muscles from XL-MTM patients express the lowest levels of RYR1 and show the greatest changes in overall transcript expression; and (v) expression of the MyHC-EO isoform during development is dependent on the function of the vestibular system and when reduced amounts of mutant RyR1 channels are present, extraocular muscles fail to express MyHC-EO. 59 , 60 Hence, it is reasonable to assume that calcium release mediated by RyR1 channels also regulates gene expression.…”

Proteomic profiling of impaired excitation–contraction coupling and abnormal calcium handling in muscular dystrophy

“…receptor dysregulation and ion channel leakiness disturb the proper linkage between motor neuron activity and skeletal muscle functioning [43][44][45]. In addition, stretch-induced fibre damage, Ca 2+ -entry through mechano-sensitive channels, altered membrane permeability and sarcosolic Ca 2+ -overload are also associated with mitochondrial dysregulation involving the mitochondrial transition pore in dystrophic muscle fibres.…”

“…This is reflected by significant alterations in the expression of a variety of proteins involved in excitation-contraction coupling [108][109][110] and store-operated Ca 2+ -entry [111][112][113] in X-linked muscular dystrophy. Of note, dystrophin-lacking skeletal muscles are characterised by significantly impaired Ca 2+ -release from the sarcoplasmic reticulum in muscular dystrophy [43][44][45]114] and the dystrophic phenotype of abnormal F I G U R E 4 Potential protein interactions between the sarcolemmal dystrophin-glycoprotein complex and proteins involved in the regulation of excitation-contraction coupling using bioinformatic STRING analysis [98] of proteins identified by the mass spectrometric survey of skeletal muscle membrane fractions [21,99]. The various evidence types that contribute to STRING are discernible in the visual networks by lines of different colours, including neighbourhood evidence, co-occurrence evidence, experimental evidence, database evidence and co-expression evidence [98].…”