Making Skeletal Muscle from Human Pluripotent Stem Cells

Human pluripotent stem cells (hPSCs) proliferate in vitro for long periods without losing pluripotency and can be induced to differentiate into various cell types including skeletal muscle cells (SMCs). Human embryonic stem cells (hESCs) are generated from a preimplantation-stage embryo. Human-induced pluripotent stem cells (hiPSCs) are derived from somatic cells of both healthy donors and patients with muscle diseases of any age using reprogramming factors. Currently, there are two kinds of protocols to induce skeletal muscle from hPSCs. One type utilizes overexpression of a potent myogenic master regulator, MyoD, to directly induce skeletal muscle. Stepwise induction of skeletal muscle has also been reported by many research groups, but hiPSC-based cell therapy for muscular dystrophy is still experimental. On the other hand, hiPSCs derived from patients with muscle disease are widely used for disease modeling in vitro. Here, we review the recent literature on derivation of skeletal muscle from human pluripotent stem cells and discuss their application

α1-Syntrophin–deficient skeletal muscle exhibits hypertrophy and aberrant formation of neuromuscular junctions during regeneration

α1-Syntrophin is a member of the family of dystrophin-associated proteins; it has been shown to recruit neuronal nitric oxide synthase and the water channel aquaporin-4 to the sarcolemma by its PSD-95/SAP-90, Discs-large, ZO-1 homologous domain. To examine the role of α1-syntrophin in muscle regeneration, we injected cardiotoxin into the tibialis anterior muscles of α1-syntrophin–null (α1syn−/−) mice. After the treatment, α1syn−/− muscles displayed remarkable hypertrophy and extensive fiber splitting compared with wild-type regenerating muscles, although the untreated muscles of the mutant mice showed no gross histological change. In the hypertrophied muscles of the mutant mice, the level of insulin-like growth factor-1 transcripts was highly elevated. Interestingly, in an early stage of the regeneration process, α1syn−/− mice showed remarkably deranged neuromuscular junctions (NMJs), accompanied by impaired ability to exercise. The contractile forces were reduced in α1syn−/− regenerating muscles. Our results suggest that the lack of α1-syntrophin might be responsible in part for the muscle hypertrophy, abnormal synapse formation at NMJs, and reduced force generation during regeneration of dystrophin-deficient muscle, all of which are typically observed in the early stages of Duchenne muscular dystrophy patients

Mesenchymal Stem Cells for Regenerative Medicine for Duchenne Muscular Dystrophy

Mesenchymal stem cells (MSCs) are multipotent stem cells that can be isolated from both foetal and adult tissues. Several groups demonstrated that transplantation of MSCs promoted the regeneration of skeletal muscle and ameliorated muscular dystrophy in animal models. Mesenchymal stem cells in skeletal muscle, also known as fibro-adipogenic progenitors (FAPs), are essential for the maintenance of skeletal muscle. Importantly, they contribute to fibrosis and fat accumulation in dystrophic muscle. Therefore, MSCs in muscle are a pharmacological target for the treatment of muscular dystrophies. In this chapter, we briefly update the knowledge on mesenchymal stem/progenitor cells and discuss their therapeutic potential as a regenerative medicine treatment of Duchenne muscular dystrophy

Calcitonin Receptor Signaling Inhibits Muscle Stem Cells from Escaping the Quiescent State and the Niche

Masahiko Yamaguchi, Yoko Watanabe, Takuji Ohtani, Akiyoshi Uezumi, Norihisa Mikami, Miki Nakamura, Takahiko Sato, Masahito Ikawa, Mikio Hoshino, Kunihiro Tsuchida, Yuko Miyagoe-Suzuki, Kazutake Tsujikawa, Shin’ichi Takeda, Hiroshi Yamamoto, So-ichiro Fukada, Calcitonin Receptor Signaling Inhibits Muscle Stem Cells from Escaping the Quiescent State and the Niche, Cell Reports, Volume 13, Issue 2, 2015, Pages 302-314, ISSN 2211-1247, https://doi.org/10.1016/j.celrep.2015.08.083

Coordinate control of axon defasciculation and myelination by laminin-2 and -8

Schwann cells form basal laminae (BLs) containing laminin-2 (Ln-2; heterotrimer α2β1γ1) and Ln-8 (α4β1γ1). Loss of Ln-2 in humans and mice carrying α2-chain mutations prevents developing Schwann cells from fully defasciculating axons, resulting in partial amyelination. The principal pathogenic mechanism is thought to derive from structural defects in Schwann cell BLs, which Ln-2 scaffolds. However, we found loss of Ln-8 caused partial amyelination in mice without affecting BL structure or Ln-2 levels. Combined Ln-2/Ln-8 deficiency caused nearly complete amyelination, revealing Ln-2 and -8 together have a dominant role in defasciculation, and that Ln-8 promotes myelination without BLs. Transgenic Ln-10 (α5β1γ1) expression also promoted myelination without BL formation. Rather than BL structure, we found Ln-2 and -8 were specifically required for the increased perinatal Schwann cell proliferation that attends myelination. Purified Ln-2 and -8 directly enhanced in vitro Schwann cell proliferation in collaboration with autocrine factors, suggesting Lns control the onset of myelination by modulating responses to mitogens in vivo

Cell-Surface Protein Profiling Identifies Distinctive Markers of Progenitor Cells in Human Skeletal Muscle

SummarySkeletal muscle contains two distinct stem/progenitor populations. One is the satellite cell, which acts as a muscle stem cell, and the other is the mesenchymal progenitor, which contributes to muscle pathogeneses such as fat infiltration and fibrosis. Detailed and accurate characterization of these progenitors in humans remains elusive. Here, we performed comprehensive cell-surface protein profiling of the two progenitor populations residing in human skeletal muscle and identified three previously unrecognized markers: CD82 and CD318 for satellite cells and CD201 for mesenchymal progenitors. These markers distinguish myogenic and mesenchymal progenitors, and enable efficient isolation of the two types of progenitors. Functional study revealed that CD82 ensures expansion and preservation of myogenic progenitors by suppressing excessive differentiation, and CD201 signaling favors adipogenesis of mesenchymal progenitors. Thus, cell-surface proteins identified here are not only useful markers but also functionally important molecules, and provide valuable insight into human muscle biology and diseases

Residual laminin-binding activity and enhanced dystroglycan glycosylation by LARGE in novel model mice to dystroglycanopathy

Hypoglycosylation and reduced laminin-binding activity of α-dystroglycan are common characteristics of dystroglycanopathy, which is a group of congenital and limb-girdle muscular dystrophies. Fukuyama-type congenital muscular dystrophy (FCMD), caused by a mutation in the fukutin gene, is a severe form of dystroglycanopathy. A retrotransposal insertion in fukutin is seen in almost all cases of FCMD. To better understand the molecular pathogenesis of dystroglycanopathies and to explore therapeutic strategies, we generated knock-in mice carrying the retrotransposal insertion in the mouse fukutin ortholog. Knock-in mice exhibited hypoglycosylated α-dystroglycan; however, no signs of muscular dystrophy were observed. More sensitive methods detected minor levels of intact α-dystroglycan, and solid-phase assays determined laminin binding levels to be ∼50% of normal. In contrast, intact α-dystroglycan is undetectable in the dystrophic Largemyd mouse, and laminin-binding activity is markedly reduced. These data indicate that a small amount of intact α-dystroglycan is sufficient to maintain muscle cell integrity in knock-in mice, suggesting that the treatment of dystroglycanopathies might not require the full recovery of glycosylation. To examine whether glycosylation defects can be restored in vivo, we performed mouse gene transfer experiments. Transfer of fukutin into knock-in mice restored glycosylation of α-dystroglycan. In addition, transfer of LARGE produced laminin-binding forms of α-dystroglycan in both knock-in mice and the POMGnT1 mutant mouse, which is another model of dystroglycanopathy. Overall, these data suggest that even partial restoration of α-dystroglycan glycosylation and laminin-binding activity by replacing or augmenting glycosylation-related genes might effectively deter dystroglycanopathy progression and thus provide therapeutic benefits

Periostin Is Required for the Maintenance of Muscle Fibers during Muscle Regeneration

Skeletal muscle regeneration is a well-organized process that requires remodeling of the extracellular matrix (ECM). In this study, we revealed the protective role of periostin, a matricellular protein that binds to several ECM proteins during muscle regeneration. In intact muscle, periostin was localized at the neuromuscular junction, muscle spindle, and myotendinous junction, which are connection sites between muscle fibers and nerves or tendons. During muscle regeneration, periostin exhibited robustly increased expression and localization at the interstitial space. Periostin-null mice showed decreased muscle weight due to the loss of muscle fibers during repeated muscle regeneration. Cultured muscle progenitor cells from periostin-null mice showed no deficiencies in their proliferation, differentiation, and the expression of Pax7, MyoD, and myogenin, suggesting that the loss of muscle fibers in periostin-null mice was not due to the impaired function of muscle stem/progenitor cells. Periostin-null mice displayed a decreased number of CD31-positive blood vessels during muscle regeneration, suggesting that the decreased nutritional supply from blood vessels was the cause of muscle fiber loss in periostin-null mice. These results highlight the novel role of periostin in maintaining muscle mass during muscle regeneration