Using stem cells to replace degenerating muscle cells and restore lost skeletal muscle function is an attractive therapeutic strategy for treating neuromuscular diseases. Myogenic progenitors are a valuable cell type for cell-based therapy and also provide a platform for studying normal muscle development and disease mechanisms in vitro. Human pluripotent stem cells represent a valuable source of tissue for generating myogenic progenitors. Here, we present a novel protocol for deriving myogenic progenitors from human embryonic stem (hES) and induced pluripotent stem (iPS) cells using free-floating spherical culture (EZ spheres) in a defined culture medium. hES cell colonies and human iPS cell colonies were expanded in medium supplemented with high concentrations (100 ng/ml) of fibroblast growth factor-2 (FGF-2) and epidermal growth factor in which they formed EZ spheres and were passaged using a mechanical chopping method. We found myogenic progenitors in the spheres after 6 weeks of culture and multinucleated myotubes following sphere dissociation and 2 weeks of terminal differentiation. A high concentration of FGF-2 plays a critical role for myogenic differentiation and is necessary for generating myogenic progenitors from pluripotent cells cultured as EZ spheres. Importantly, EZ sphere culture produced myogenic progenitors from human iPS cells generated from both healthy donors and patients with neuromuscular disorders (including Becker's muscular dystrophy, spinal muscular atrophy, and familial amyotrophic lateral sclerosis). Taken together, this study demonstrates a simple method for generating myogenic cells from pluripotent sources under defined conditions for potential use in disease modeling or cell-based therapies targeting skeletal muscle.
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http://dx.doi.org/10.5966/sctm.2013-0143 | DOI Listing |
Int J Mol Sci
December 2024
Department of Biochemistry, Dongguk University College of Medicine, 123 Dongdae-ro, Gyeongju 38066, Republic of Korea.
Cortactin (CTTN) is an actin-binding protein regulating actin polymerization and stabilization, which are vital processes for maintaining skeletal muscle homeostasis. Despite the established function of CTTN in actin cytoskeletal dynamics, its role in the myogenic differentiation of progenitor cells remains largely unexplored. In this study, we investigated the role of CTTN in the myogenic differentiation of C2C12 myoblasts by analyzing its effects on actin cytoskeletal remodeling, myocardin-related transcription factor A (MRTFA) nuclear translocation, serum response factor (SRF) activation, expression of myogenic transcription factors, and myotube formation.
View Article and Find Full Text PDFBioengineering (Basel)
December 2024
Department of Orthopedic Surgery, Mayo Clinic, Rochester, MN 55905, USA.
Muscle stem cells (MuSCs) are essential for skeletal muscle regeneration, influenced by a complex interplay of mechanical, biochemical, and molecular cues. Properties of the extracellular matrix (ECM) such as stiffness and alignment guide stem cell fate through mechanosensitive pathways, where forces like shear stress translate into biochemical signals, affecting cell behavior. Aging introduces senescence which disrupts the MuSC niche, leading to reduced regenerative capacity via epigenetic alterations and metabolic shifts.
View Article and Find Full Text PDFJ Adv Res
December 2024
College of Animal Science, Shandong Provincial Key Laboratory for Livestock Germplasm Innovation & Utilization, Shandong Agricultural University, Taian, China; College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, China. Electronic address:
Brown adipose tissue (BAT) is responsible for non-shivering thermogenesis, but it is absent in some mammals, including pigs. During development, BAT progenitors are derived from paired box 7 (Pax7)-expressing somitic mesodermal stem cells, which also give rise to skeletal muscle. However, the intrinsic mechanisms underlying the fate decisions between brown fat and muscle progenitors remain elusive across species.
View Article and Find Full Text PDFJ Cachexia Sarcopenia Muscle
February 2025
Sprott Centre for Stem Cell Research, Regenerative Medicine Program, Ottawa Hospital Research Institute, Ottawa, Canada.
Background: Duchenne muscular dystrophy (DMD) is a devastating disease characterized by progressive muscle wasting that leads to diminished lifespan. In addition to the inherent weakness of dystrophin-deficient muscle, the dysfunction of resident muscle stem cells (MuSC) significantly contributes to disease progression.
Methods: Using the mdx mouse model of DMD, we performed an in-depth characterization of disease progression and MuSC function in dystrophin-deficient skeletal muscle using immunohistology, isometric force measurements, transcriptomic analysis and transplantation assays.
bioRxiv
December 2024
Sanford Burnham Prebys Medical Discovery Institute, Development, Aging and Regeneration Program, La Jolla, CA 92037, USA.
Generation of induced pluripotent cells (hiPSCs)-derived skeletal muscle progenitor cells (SMPCs) holds great promise for regenerative medicine for skeletal muscle wasting diseases, as for example Duchenne Muscular Dystrophy (DMD). Multiple approaches, involving ectopic expression of key regulatory myogenic genes or small molecules cocktails, have been described by different groups to obtain SMPC towards cell-transplantation as a therapeutic approach to skeletal muscle diseases. However, hiPSCs-derived SMPC generated using transgene-free protocols are usually obtained in a low amount and resemble a more embryonal/fetal stage of differentiation.
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