Prednisolone rescues Duchenne muscular dystrophy phenotypes in human pluripotent stem cell–derived skeletal muscle in vitro

Abstract

Significance Duchenne muscular dystrophy (DMD) is a devastating disease caused by mutation in the X-linked dystrophin gene, resulting in skeletal muscle loss and patient premature death. Here, we present an improved protocol for the differentiation of human pluripotent stem cells to the skeletal muscle lineage. Using this protocol for the differentiation of human isogenic-induced pluripotent stem cells lacking the DMD gene, we can reproduce several features of the pathology including mislocalization of proteins of the dystrophin-associated complex, increased fiber branching, contraction defects, and calcium signaling hyperactivity. Several of these defects can be rescued in part in vitro by treating cells with prednisolone, the standard of care for DMD patients, supporting direct action of this drug on the diseased fibers. Duchenne muscular dystrophy (DMD) is a devastating genetic disease leading to degeneration of skeletal muscles and premature death. How dystrophin absence leads to muscle wasting remains unclear. Here, we describe an optimized protocol to differentiate human induced pluripotent stem cells (iPSC) to a late myogenic stage. This allows us to recapitulate classical DMD phenotypes (mislocalization of proteins of the dystrophin-associated glycoprotein complex, increased fusion, myofiber branching, force contraction defects, and calcium hyperactivation) in isogenic DMD-mutant iPSC lines in vitro. Treatment of the myogenic cultures with prednisolone (the standard of care for DMD) can dramatically rescue force contraction, fusion, and branching defects in DMD iPSC lines. This argues that prednisolone acts directly on myofibers, challenging the largely prevalent view that its beneficial effects are caused by antiinflammatory properties. Our work introduces a human in vitro model to study the onset of DMD pathology and test novel therapeutic approaches.