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Michalska Z, Ostaszewska A, Fularczyk M, Dzierżyńska M, Bielak K, Morytz J, Sieradzan AK, Archacka K, Brzoska E, Rodziewicz-Motowidło S, Ciemerych MA. In Vitro Bioactivity Evaluation of IL-4 and SDF-1 Mimicking Peptides Engineered to Enhance Skeletal Muscle Reconstruction. J Biomed Mater Res A 2025; 113:e37898. [PMID: 40087853 DOI: 10.1002/jbm.a.37898] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/15/2025] [Revised: 02/25/2025] [Accepted: 03/04/2025] [Indexed: 03/17/2025]
Abstract
Skeletal muscle regeneration depends on satellite cells, which, in response to injury, activate, proliferate, and reconstruct damaged tissue. However, under certain conditions, such as large injuries or myopathies, this process may not be properly executed, and muscle function may be affected. Thus, pro-regenerative actions, such as the use of various factors or cells, are widely tested as a tool to improve muscle regeneration. In the current study, we designed peptides derived from the IL-4 and SDF-1 proteins, namely IL-4-X, IL-4-Y, SDF-1-X, and SDF-1-Y. We showed that these peptides can bind to appropriate receptors and can adopt proper structure in solution. Importantly, we documented, using in vitro culture, that they do not negatively affect the cells that are present and active in skeletal muscles, such as myoblasts and fibroblasts, bone marrow stromal cells, as well as induced pluripotent stem cells, which can serve as a source of myoblasts. The presence of peptides did not affect cell proliferation compared to untreated cells. In vitro culture and differentiation protocols documented that selected IL-4 and SDF-1 peptides increased cell migration and inhibited undesirable adipogenic differentiation. Thus, we proved that these peptides are safe to use in in vivo studies aimed at improving skeletal muscle regeneration.
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Affiliation(s)
- Zuzanna Michalska
- Department of Cytology, Institute of Developmental Biology and Biomedical Sciences, Faculty of Biology, University of Warsaw, Warsaw, Poland
| | - Anna Ostaszewska
- Department of Cytology, Institute of Developmental Biology and Biomedical Sciences, Faculty of Biology, University of Warsaw, Warsaw, Poland
| | - Martyna Fularczyk
- Department of Biomedical Chemistry, Faculty of Chemistry, University of Gdańsk, Gdańsk, Poland
| | - Maria Dzierżyńska
- Department of Biomedical Chemistry, Faculty of Chemistry, University of Gdańsk, Gdańsk, Poland
| | - Kacper Bielak
- Department of Cytology, Institute of Developmental Biology and Biomedical Sciences, Faculty of Biology, University of Warsaw, Warsaw, Poland
| | - Justyna Morytz
- Department of Cytology, Institute of Developmental Biology and Biomedical Sciences, Faculty of Biology, University of Warsaw, Warsaw, Poland
| | - Adam K Sieradzan
- Department of Theoretical Chemistry, Faculty of Chemistry, University of Gdańsk, Gdańsk, Poland
| | - Karolina Archacka
- Department of Cytology, Institute of Developmental Biology and Biomedical Sciences, Faculty of Biology, University of Warsaw, Warsaw, Poland
| | - Edyta Brzoska
- Department of Cytology, Institute of Developmental Biology and Biomedical Sciences, Faculty of Biology, University of Warsaw, Warsaw, Poland
| | | | - Maria A Ciemerych
- Department of Cytology, Institute of Developmental Biology and Biomedical Sciences, Faculty of Biology, University of Warsaw, Warsaw, Poland
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Rashid MI, Ito T, Miya F, Shimojo D, Arimoto K, Onodera K, Okada R, Nagashima T, Yamamoto K, Khatun Z, Shimul RI, Niwa JI, Katsuno M, Sobue G, Okano H, Sakurai H, Shimizu K, Doyu M, Okada Y. Simple and efficient differentiation of human iPSCs into contractible skeletal muscles for muscular disease modeling. Sci Rep 2023; 13:8146. [PMID: 37231024 DOI: 10.1038/s41598-023-34445-9] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2022] [Accepted: 04/30/2023] [Indexed: 05/27/2023] Open
Abstract
Pathophysiological analysis and drug discovery targeting human diseases require disease models that suitably recapitulate patient pathology. Disease-specific human induced pluripotent stem cells (hiPSCs) differentiated into affected cell types can potentially recapitulate disease pathology more accurately than existing disease models. Such successful modeling of muscular diseases requires efficient differentiation of hiPSCs into skeletal muscles. hiPSCs transduced with doxycycline-inducible MYOD1 (MYOD1-hiPSCs) have been widely used; however, they require time- and labor-consuming clonal selection, and clonal variations must be overcome. Moreover, their functionality should be carefully examined. Here, we demonstrated that bulk MYOD1-hiPSCs established with puromycin selection rather than G418 selection showed rapid and highly efficient differentiation. Interestingly, bulk MYOD1-hiPSCs exhibited average differentiation properties of clonally established MYOD1-hiPSCs, suggesting that it is possible to minimize clonal variations. Moreover, disease-specific hiPSCs of spinal bulbar muscular atrophy (SBMA) could be efficiently differentiated via this method into skeletal muscle that showed disease phenotypes, suggesting the applicability of this method for disease analysis. Finally, three-dimensional muscle tissues were fabricated from bulk MYOD1-hiPSCs, which exhibited contractile force upon electrical stimulation, indicating their functionality. Thus, our bulk differentiation requires less time and labor than existing methods, efficiently generates contractible skeletal muscles, and may facilitate the generation of muscular disease models.
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Affiliation(s)
- Muhammad Irfanur Rashid
- Department of Neural iPSC Research, Institute for Medical Science of Aging, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
- Department of Neurology, Aichi Medical University School of Medicine, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
| | - Takuji Ito
- Department of Neural iPSC Research, Institute for Medical Science of Aging, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
- Department of Neurology, Aichi Medical University School of Medicine, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
- Japan Society for the Promotion of Science, 5-3-1 Kojimachi, Chiyoda-ku, Tokyo, 102-0083, Japan
| | - Fuyuki Miya
- Center for Medical Genetics, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan
| | - Daisuke Shimojo
- Department of Neurology, Aichi Medical University School of Medicine, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
- Department of Physiology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan
| | - Kanae Arimoto
- Department of Biomolecular Engineering, Graduate School of Engineering, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi, 464-8603, Japan
| | - Kazunari Onodera
- Department of Neural iPSC Research, Institute for Medical Science of Aging, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
- Department of Neurology, Aichi Medical University School of Medicine, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
- Department of Neurology, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi, 466-8650, Japan
| | - Rina Okada
- Department of Neural iPSC Research, Institute for Medical Science of Aging, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
- Department of Neurology, Aichi Medical University School of Medicine, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
- Japan Society for the Promotion of Science, 5-3-1 Kojimachi, Chiyoda-ku, Tokyo, 102-0083, Japan
| | - Takunori Nagashima
- Department of Biomolecular Engineering, Graduate School of Engineering, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi, 464-8603, Japan
| | - Kazuki Yamamoto
- Department of Biomolecular Engineering, Graduate School of Engineering, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi, 464-8603, Japan
| | - Zohora Khatun
- Department of Neural iPSC Research, Institute for Medical Science of Aging, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
- Department of Neurology, Aichi Medical University School of Medicine, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
| | - Rayhanul Islam Shimul
- Department of Neural iPSC Research, Institute for Medical Science of Aging, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
| | - Jun-Ichi Niwa
- Department of Neurology, Aichi Medical University School of Medicine, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
| | - Masahisa Katsuno
- Department of Neurology, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi, 466-8650, Japan
- Department of Clinical Research Education, Nagoya University Graduate School of Medicine, Showa-ku, Nagoya, Aichi, 466-8650, Japan
| | - Gen Sobue
- Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
| | - Hideyuki Okano
- Department of Physiology, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo, 160-8582, Japan
| | - Hidetoshi Sakurai
- Department of Clinical Application, Center for iPS Cell Research and Application (CiRA), Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto, 606-8507, Japan
| | - Kazunori Shimizu
- Department of Biomolecular Engineering, Graduate School of Engineering, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi, 464-8603, Japan
| | - Manabu Doyu
- Department of Neurology, Aichi Medical University School of Medicine, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan
| | - Yohei Okada
- Department of Neural iPSC Research, Institute for Medical Science of Aging, Aichi Medical University, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan.
- Department of Neurology, Aichi Medical University School of Medicine, 1-1 Yazakokarimata, Nagakute, Aichi, 480-1195, Japan.
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3
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Liu M, Cheng L, Li X, Wang H, Wang M, Gan L. Resveratrol Reverses Myogenic Induction Suppression Caused by High Glucose Through Activating the SIRT1/AKT/FOXO1 Pathway. Nat Prod Commun 2023. [DOI: 10.1177/1934578x231159722] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/06/2023] Open
Abstract
Background Differentiated bone marrow mesenchymal stem cells (BMSCs) may be a therapeutic strategy to treat sarcopenia caused by high glucose. The effects of resveratrol in the myogenic induction of BMSCs under high glucose are unknown. We evaluated the effects and possible mechanisms of high glucose and resveratrol on myogenic induction of rat BMSCs. Methods Primary rat BMSCs were isolated and purified from Sprague-Dawley rats aged between 3 and 4 weeks. Rat BMSCs were differentiated into myogenic cells using conditioned medium and treated with glucose and/or resveratrol along with EX527 (a specific silent information regulator 1 [SIRT1] inhibitor). The expressions of MyoD1 and Myogenin were measured. The reactive oxygen species (ROS) level, superoxide dismutase (SOD) activity, and the expressions of FOXO1 and p-AKT/AKT during myogenic induction were also examined. Results High glucose decreased cell viability, cell proliferation, and SOD activity, increased intracellular ROS levels, and inhibited the AKT/FOXO1. Resveratrol reversed myogenic induction suppression caused by high glucose, partly through restoring cell proliferation and viability, reducing peroxidative damage, and activating the AKT/FOXO1 pathway; this effect was eliminated by EX527. Conclusion Our results indicate that resveratrol promoted myogenic induction and partially reversed the suppression of myogenic induction caused by high glucose through activating the SIRT1/AKT/FOXO1 pathway.
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Affiliation(s)
- Meiling Liu
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, Harbin, China
| | - Luyang Cheng
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, Harbin, China
| | - Xianglu Li
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, Harbin, China
| | - Hongzhi Wang
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, Harbin, China
| | - Manfeng Wang
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, Harbin, China
| | - Lu Gan
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, Harbin, China
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Generation of the First Human In Vitro Model for McArdle Disease Based on iPSC Technology. Int J Mol Sci 2022; 23:ijms232213964. [PMID: 36430443 PMCID: PMC9692531 DOI: 10.3390/ijms232213964] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2022] [Revised: 11/04/2022] [Accepted: 11/10/2022] [Indexed: 11/16/2022] Open
Abstract
McArdle disease is a rare autosomal recessive disorder caused by mutations in the PYGM gene. This gene encodes for the skeletal muscle isoform of glycogen phosphorylase (myophosphorylase), the first enzyme in glycogenolysis. Patients with this disorder are unable to obtain energy from their glycogen stored in skeletal muscle, prompting an exercise intolerance. Currently, there is no treatment for this disease, and the lack of suitable in vitro human models has prevented the search for therapies against it. In this article, we have established the first human iPSC-based model for McArdle disease. For the generation of this model, induced pluripotent stem cells (iPSCs) from a patient with McArdle disease (harbouring the homozygous mutation c.148C>T; p.R50* in the PYGM gene) were differentiated into myogenic cells able to contract spontaneously in the presence of motor neurons and generate calcium transients, a proof of their maturity and functionality. Additionally, an isogenic skeletal muscle model of McArdle disease was created. As a proof-of-concept, we have tested in this model the rescue of PYGM expression by two different read-through compounds (PTC124 and RTC13). The developed model will be very useful as a platform for testing drugs or compounds with potential pharmacological activity.
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Choi S, Ferrari G, Moyle LA, Mackinlay K, Naouar N, Jalal S, Benedetti S, Wells C, Muntoni F, Tedesco FS. Assessing and enhancing migration of human myogenic progenitors using directed iPS cell differentiation and advanced tissue modelling. EMBO Mol Med 2022; 14:e14526. [PMID: 36161772 PMCID: PMC9549733 DOI: 10.15252/emmm.202114526] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2021] [Revised: 08/19/2022] [Accepted: 08/19/2022] [Indexed: 02/05/2023] Open
Abstract
Muscle satellite stem cells (MuSCs) are responsible for skeletal muscle growth and regeneration. Despite their differentiation potential, human MuSCs have limited in vitro expansion and in vivo migration capacity, limiting their use in cell therapies for diseases affecting multiple skeletal muscles. Several protocols have been developed to derive MuSC-like progenitors from human induced pluripotent stem (iPS) cells (hiPSCs) to establish a source of myogenic cells with controllable proliferation and differentiation. However, current hiPSC myogenic derivatives also suffer from limitations of cell migration, ultimately delaying their clinical translation. Here we use a multi-disciplinary approach including bioinformatics and tissue engineering to show that DLL4 and PDGF-BB improve migration of hiPSC-derived myogenic progenitors. Transcriptomic analyses demonstrate that this property is conserved across species and multiple hiPSC lines, consistent with results from single cell motility profiling. Treated cells showed enhanced trans-endothelial migration in transwell assays. Finally, increased motility was detected in a novel humanised assay to study cell migration using 3D artificial muscles, harnessing advanced tissue modelling to move hiPSCs closer to future muscle gene and cell therapies.
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Affiliation(s)
- SungWoo Choi
- The Francis Crick InstituteLondonUK
- Department of Cell and Developmental BiologyUniversity College LondonLondonUK
| | - Giulia Ferrari
- Department of Cell and Developmental BiologyUniversity College LondonLondonUK
| | - Louise A Moyle
- Department of Cell and Developmental BiologyUniversity College LondonLondonUK
- Present address:
Institute of Biomedical EngineeringUniversity of TorontoTorontoONCanada
| | - Kirsty Mackinlay
- Department of Cell and Developmental BiologyUniversity College LondonLondonUK
- Present address:
Department of Physiology, Development and NeuroscienceUniversity of CambridgeCambridgeUK
| | - Naira Naouar
- Institut de Biologie Paris Seine FR3631, Plateforme de Bioinformatique ARTbioSorbonne UniversitéParisFrance
| | - Salma Jalal
- The Francis Crick InstituteLondonUK
- Department of Cell and Developmental BiologyUniversity College LondonLondonUK
| | - Sara Benedetti
- UCL Great Ormond Street Institute of Child HealthUniversity College LondonLondonUK
- National Institute for Health Research Great Ormond Street Hospital Biomedical Research CentreLondonUK
| | - Christine Wells
- Centre for Stem Cell SystemsThe University of MelbourneMelbourneVICAustralia
| | - Francesco Muntoni
- National Institute for Health Research Great Ormond Street Hospital Biomedical Research CentreLondonUK
- Dubowitz Neuromuscular CentreUCL Great Ormond Street Institute of Child Health & Great Ormond Street Hospital for ChildrenLondonUK
| | - Francesco Saverio Tedesco
- The Francis Crick InstituteLondonUK
- Department of Cell and Developmental BiologyUniversity College LondonLondonUK
- Dubowitz Neuromuscular CentreUCL Great Ormond Street Institute of Child Health & Great Ormond Street Hospital for ChildrenLondonUK
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6
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Messmer T, Klevernic I, Furquim C, Ovchinnikova E, Dogan A, Cruz H, Post MJ, Flack JE. A serum-free media formulation for cultured meat production supports bovine satellite cell differentiation in the absence of serum starvation. NATURE FOOD 2022; 3:74-85. [PMID: 37118488 DOI: 10.1038/s43016-021-00419-1] [Citation(s) in RCA: 79] [Impact Index Per Article: 26.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/07/2021] [Accepted: 11/01/2021] [Indexed: 01/08/2023]
Abstract
Cultured meat production requires the robust differentiation of satellite cells into mature muscle fibres without the use of animal-derived components. Current protocols induce myogenic differentiation in vitro through serum starvation, that is, an abrupt reduction in serum concentration. Here we used RNA sequencing to investigate the transcriptomic remodelling of bovine satellite cells during myogenic differentiation induced by serum starvation. We characterized canonical myogenic gene expression, and identified surface receptors upregulated during the early phase of differentiation, including IGF1R, TFRC and LPAR1. Supplementation of ligands to these receptors enabled the formulation of a chemically defined media that induced differentiation in the absence of serum starvation and/or transgene expression. Serum-free myogenic differentiation was of similar extent to that induced by serum starvation, as evaluated by transcriptome analysis, protein expression and the presence of a functional contractile apparatus. Moreover, the serum-free differentiation media supported the fabrication of three-dimensional bioartificial muscle constructs, demonstrating its suitability for cultured beef production.
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7
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Zhang M, Niibe K, Kondo T, Limraksasin P, Okawa H, Miao X, Kamano Y, Yamada M, Jiang X, Egusa H. Rapid and efficient generation of cartilage pellets from mouse induced pluripotent stem cells by transcriptional activation of BMP-4 with shaking culture. J Tissue Eng 2022; 13:20417314221114616. [PMID: 35923173 PMCID: PMC9340412 DOI: 10.1177/20417314221114616] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/17/2022] [Accepted: 07/04/2022] [Indexed: 11/15/2022] Open
Abstract
Induced pluripotent stem cells (iPSCs) offer an unlimited source for cartilage
regeneration as they can generate a wide spectrum of cell types. Here, we
established a tetracycline (tet) controlled bone morphogenetic
protein-4 (BMP-4) expressing iPSC
(iPSC-Tet/BMP-4) line in which transcriptional activation
of BMP-4 was associated with enhanced chondrogenesis. Moreover,
we developed an efficient and simple approach for directly guiding
iPSC-Tet/BMP-4 differentiation into chondrocytes in
scaffold-free cartilaginous pellets using a combination of transcriptional
activation of BMP-4 and a 3D shaking suspension culture system.
In chondrogenic induction medium, shaking culture alone significantly
upregulated the chondrogenic markers Sox9, Col2a1, and
Aggrecan in iPSCs-Tet/BMP-4 by day 21. Of
note, transcriptional activation of BMP-4 by addition of tet
(doxycycline) greatly enhanced the expression of these genes. The cartilaginous
pellets derived from iPSCs-Tet/BMP-4 showed an oval morphology
and white smooth appearance by day 21. After day 21, the cells presented a
typical round morphology and the extracellular matrix was stained intensively
with Safranin O, alcian blue, and type II collagen. In addition, the homogenous
cartilaginous pellets derived from iPSCs-Tet/BMP-4 with 28 days
of induction repaired joint osteochondral defects in immunosuppressed rats and
integrated well with the adjacent host cartilage. The regenerated cartilage
expressed the neomycin resistance gene, indicating that the newly formed
cartilage was generated by the transplanted iPSCs-Tet/BMP-4.
Thus, our culture system could be a useful tool for further investigation of the
mechanism of BMP-4 in regulating iPSC differentiation toward the chondrogenic
lineage, and should facilitate research in cartilage development, repair, and
osteoarthritis.
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Affiliation(s)
- Maolin Zhang
- Division of Molecular and Regenerative Prosthodontics, Tohoku University Graduate School of Dentistry, Sendai, Miyagi, Japan
- Department of Prosthodontics, Ninth People’s Hospital affiliated to Shanghai Jiao Tong University, School of Medicine, Shanghai, China
| | - Kunimichi Niibe
- Division of Molecular and Regenerative Prosthodontics, Tohoku University Graduate School of Dentistry, Sendai, Miyagi, Japan
| | - Takeru Kondo
- Division of Molecular and Regenerative Prosthodontics, Tohoku University Graduate School of Dentistry, Sendai, Miyagi, Japan
| | - Phoonsuk Limraksasin
- Division of Molecular and Regenerative Prosthodontics, Tohoku University Graduate School of Dentistry, Sendai, Miyagi, Japan
| | - Hiroko Okawa
- Division of Molecular and Regenerative Prosthodontics, Tohoku University Graduate School of Dentistry, Sendai, Miyagi, Japan
| | - Xinchao Miao
- Division of Molecular and Regenerative Prosthodontics, Tohoku University Graduate School of Dentistry, Sendai, Miyagi, Japan
| | - Yuya Kamano
- Division of Molecular and Regenerative Prosthodontics, Tohoku University Graduate School of Dentistry, Sendai, Miyagi, Japan
| | - Masahiro Yamada
- Division of Molecular and Regenerative Prosthodontics, Tohoku University Graduate School of Dentistry, Sendai, Miyagi, Japan
| | - Xinquan Jiang
- Department of Prosthodontics, Ninth People’s Hospital affiliated to Shanghai Jiao Tong University, School of Medicine, Shanghai, China
| | - Hiroshi Egusa
- Division of Molecular and Regenerative Prosthodontics, Tohoku University Graduate School of Dentistry, Sendai, Miyagi, Japan
- Center for Advanced Stem Cell and Regenerative Research, Tohoku University Graduate School of Dentistry, Sendai, Miyagi, Japan
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Jalal S, Dastidar S, Tedesco FS. Advanced models of human skeletal muscle differentiation, development and disease: Three-dimensional cultures, organoids and beyond. Curr Opin Cell Biol 2021; 73:92-104. [PMID: 34384976 PMCID: PMC8692266 DOI: 10.1016/j.ceb.2021.06.004] [Citation(s) in RCA: 34] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/16/2021] [Accepted: 06/23/2021] [Indexed: 02/08/2023]
Abstract
Advanced in vitro models of human skeletal muscle tissue are increasingly needed to model complex developmental dynamics and disease mechanisms not recapitulated in animal models or in conventional monolayer cell cultures. There has been impressive progress towards creating such models by using tissue engineering approaches to recapitulate a range of physical and biochemical components of native human skeletal muscle tissue. In this review, we discuss recent studies focussed on developing complex in vitro models of human skeletal muscle beyond monolayer cell cultures, involving skeletal myogenic differentiation from human primary myoblasts or pluripotent stem cells, often in the presence of structural scaffolding support. We conclude with our outlook on the future of advanced skeletal muscle three-dimensional cultures (e.g. organoids and biofabrication) to produce physiologically and clinically relevant platforms for disease modelling and therapy development in musculoskeletal and neuromuscular disorders.
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Affiliation(s)
- Salma Jalal
- Department of Cell and Developmental Biology, University College London, WC1E 6DE London, United Kingdom
| | - Sumitava Dastidar
- Department of Cell and Developmental Biology, University College London, WC1E 6DE London, United Kingdom
| | - Francesco Saverio Tedesco
- Department of Cell and Developmental Biology, University College London, WC1E 6DE London, United Kingdom; The Francis Crick Institute, 1 Midland Road, London NW1 1AT, United Kingdom; Dubowitz Neuromuscular Centre, Great Ormond Street Institute of Child Health, University College London, London WC1N 1EH, United Kingdom; Department of Paediatric Neurology, Great Ormond Street Hospital for Children, WC1N 3JH London, United Kingdom.
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9
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Kim N, Yokobayashi Y. Novel RNA Viral Vectors for Chemically Regulated Gene Expression in Embryonic Stem Cells. ACS Synth Biol 2021; 10:2959-2967. [PMID: 34676762 DOI: 10.1021/acssynbio.1c00214] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
RNA viral vectors that replicate without DNA intermediates are attractive platforms for manipulation of cells for biomedical and veterinary applications because they have minimal risk of chromosomal integration. Vesicular stomatitis virus (VSV) vectors are among the most well-studied RNA viral vectors due to their low pathogenicity to humans and ability to express transgenes at high levels for weeks to months. However, their applications have been mostly limited to oncolytic and vaccine vectors due to their cytopathogenicity. We discovered two mutations in the VSV vector that synergistically confer improved stability in mouse embryonic stem cells (ESCs) with markedly lower cytopathic effects. We also demonstrated chemical regulation of transgene expression through embedded riboswitches. The ESCs infected with the mutant vector were shown to maintain pluripotency. This new vector sets the stage for precise regulation of gene expression in ESCs to produce a variety of differentiated cells without chromosomal alteration.
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Affiliation(s)
- Narae Kim
- Nucleic Acid Chemistry and Engineering Unit, Okinawa Institute of Science and Technology Graduate University, Onna, Okinawa 904 0495, Japan
| | - Yohei Yokobayashi
- Nucleic Acid Chemistry and Engineering Unit, Okinawa Institute of Science and Technology Graduate University, Onna, Okinawa 904 0495, Japan
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10
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Jin GZ. Enhanced growth and myogenic differentiation of spheroid-derived C2C12 cells. Biosci Biotechnol Biochem 2021; 85:1227-1234. [PMID: 33704409 DOI: 10.1093/bbb/zbab018] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/04/2020] [Accepted: 01/22/2021] [Indexed: 11/13/2022]
Abstract
Among many factors of controlling stem cell differentiation, the key transcription factor upregulation via physical force is a good strategy on the lineage-specific differentiation of stem cells. The study aimed to compare growth and myogenic potentials between the parental cells (PCs) and the 1-day-old C2C12 spheroid-derived cells (SDCs) in two-dimensional (2D) and three-dimensional (3D) culture conditions through examination of the cell proliferation and the expression of myogenic genes. The data showed that 1-day-old spheroids had more intense expression of MyoD gene with respect to the PCs. The proliferation of the SDCs is significantly higher than the PCs in a time-dependent manner. The SDCs had also significantly higher myogenic potential than the PCs in 2D and 3D culture conditions. The results suggest that MyoD gene upregulation through cell-cell contacts is the good approach for preparation of seed cells in muscle tissue engineering.
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Affiliation(s)
- Guang-Zhen Jin
- Institute of Tissue Regeneration Engineering (ITREN), Dankook University, Cheonan, Republic of Korea.,Department of Nanobiomedical Science and BK21 PLUS NBM Global Research Center for Regenerative Medicine, Dankook University, Cheonan, Republic of Korea.,Department of Biomaterials Science, College of Dentistry, Dankook University, Cheonan, Republic of Korea
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11
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Canonico F, Chirivi M, Maiullari F, Milan M, Rizzi R, Arcudi A, Galli M, Pane M, Gowran A, Pompilio G, Mercuri E, Crea F, Bearzi C, D'Amario D. Focus on the road to modelling cardiomyopathy in muscular dystrophy. Cardiovasc Res 2021; 118:1872-1884. [PMID: 34254111 DOI: 10.1093/cvr/cvab232] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/08/2021] [Accepted: 07/07/2021] [Indexed: 11/13/2022] Open
Abstract
Alterations in the DMD gene, which codes for the protein dystrophin, cause forms of dystrophinopathies such as Duchenne muscular dystrophy, an X-linked disease. Cardiomyopathy linked to DMD mutations is becoming the leading cause of death in patients with dystrophinopathy. Since phenotypic pathophysiological mechanisms are not fully understood, the improvement and development of new disease models, considering their relative advantages and disadvantages, is essential. The application of genetic engineering approaches on induced pluripotent stem cells, such as gene editing technology, enables the development of physiologically relevant human cell models for in vitro dystrophinopathy studies. The combination of induced pluripotent stem cells-derived cardiovascular cell types and 3 D bioprinting technologies hold great promise for the study of dystrophin-linked cardiomyopathy. This combined approach enables the assessment of responses to physical or chemical stimuli, and the influence of pharmaceutical approaches. The critical objective of in vitro microphysiological systems is to more accurately reproduce the microenvironment observed in vivo. Ground-breaking methodology involving the connection of multiple microphysiological systems comprised of different tissues would represent a move toward precision body-on-chip disease modelling could lead to a critical expansion in what is known about inter-organ responses to disease and novel therapies that have the potential to replace animal models. In this review, we will focus on the generation, development, and application of current cellular, animal and potential for bio-printed models, in the study of the pathophysiological mechanisms underlying dystrophin-linked cardiomyopathy in the direction of personalized medicine.
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Affiliation(s)
- Francesco Canonico
- Fondazione Policlinico Universitario A. Gemelli IRCCS, Department of Cardiovascular Sciences, Rome, Italy
| | - Maila Chirivi
- Institute of Biochemistry and Cell Biology, National Research Council of Italy (IBBC-CNR), Monterotondo, Rome, Italy.,Istituto Nazionale Genetica Molecolare (INGM) "Romeo ed Enrica Invernizzi", Milan, Italy
| | - Fabio Maiullari
- Istituto Nazionale Genetica Molecolare (INGM) "Romeo ed Enrica Invernizzi", Milan, Italy
| | - Marika Milan
- Institute of Biochemistry and Cell Biology, National Research Council of Italy (IBBC-CNR), Monterotondo, Rome, Italy.,Istituto Nazionale Genetica Molecolare (INGM) "Romeo ed Enrica Invernizzi", Milan, Italy
| | - Roberto Rizzi
- Istituto Nazionale Genetica Molecolare (INGM) "Romeo ed Enrica Invernizzi", Milan, Italy.,Institute of Biomedical Technologies, National Research Council of Italy (ITB-CNR), Segrate, Milan, Italy
| | - Alessandra Arcudi
- Fondazione Policlinico Universitario A. Gemelli IRCCS, Department of Cardiovascular Sciences, Rome, Italy
| | - Mattia Galli
- Fondazione Policlinico Universitario A. Gemelli IRCCS, Department of Cardiovascular Sciences, Rome, Italy
| | - Marika Pane
- Fondazione Policlinico Universitario A. Gemelli IRCCS, Department of Women, Children and Public Health Sciences, Rome, Italy
| | - Aoife Gowran
- Centro Cardiologico Monzino IRCCS, Unit of Vascular Biology and Regenerative Medicine, Milan, Italy
| | - Giulio Pompilio
- Centro Cardiologico Monzino IRCCS, Unit of Vascular Biology and Regenerative Medicine, Milan, Italy.,Department of Biomedical, Surgical and Dental Sciences, University of Milan, Italy
| | - Eugenio Mercuri
- Fondazione Policlinico Universitario A. Gemelli IRCCS, Department of Women, Children and Public Health Sciences, Rome, Italy
| | - Filippo Crea
- Fondazione Policlinico Universitario A. Gemelli IRCCS, Department of Cardiovascular Sciences, Rome, Italy
| | - Claudia Bearzi
- Istituto Nazionale Genetica Molecolare (INGM) "Romeo ed Enrica Invernizzi", Milan, Italy.,Institute of Genetic and Biomedical Research, National Research Council (IRGB-CNR), Milan, Italy
| | - Domenico D'Amario
- Fondazione Policlinico Universitario A. Gemelli IRCCS, Department of Cardiovascular Sciences, Rome, Italy
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12
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Abstract
Due to the ability to differentiate into variety of cell types, mesenchymal stem cells (MSCs) hold promise as source in cell-based therapy for treating injured tissue and degenerative diseases. The potential use of MSCs to replace or repair damaged tissues may depend on the efficient differentiation protocols to derive specialized cells without any negative side effects. Identification of appropriate cues that support the lineage-specific differentiation of stem cells is critical for tissue healing and cellular therapy. Recently, a number of stimuli have been utilized to direct the differentiation of stem cells. Biochemical stimuli such as small molecule, growth factor and miRNA have been traditionally used to regulate the fate of stem cells. In recent years, many studies have reported that biophysical stimuli including cyclic mechanical strain, fluid shear stress, microgravity, electrical stimulation, matrix stiffness and topography can also be sensed by stem cells through mechanical receptors, thus affecting the stem cell behaviors including their differentiation potential. In this paper, we review all the most recent literature on the application of biochemical and biophysical cues on regulating MSC differentiation. An extensive literature search was done using electronic database (Medline/Pubmed). Although there are still some challenges that need to be taken into consideration before translating these methods into clinics, biochemical and biophysical stimulation appears to be an attractive method to manipulate the lineage commitment of MSCs.
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13
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Alarcin E, Bal-Öztürk A, Avci H, Ghorbanpoor H, Dogan Guzel F, Akpek A, Yesiltas G, Canak-Ipek T, Avci-Adali M. Current Strategies for the Regeneration of Skeletal Muscle Tissue. Int J Mol Sci 2021; 22:5929. [PMID: 34072959 PMCID: PMC8198586 DOI: 10.3390/ijms22115929] [Citation(s) in RCA: 37] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/27/2021] [Revised: 05/21/2021] [Accepted: 05/26/2021] [Indexed: 12/11/2022] Open
Abstract
Traumatic injuries, tumor resections, and degenerative diseases can damage skeletal muscle and lead to functional impairment and severe disability. Skeletal muscle regeneration is a complex process that depends on various cell types, signaling molecules, architectural cues, and physicochemical properties to be successful. To promote muscle repair and regeneration, various strategies for skeletal muscle tissue engineering have been developed in the last decades. However, there is still a high demand for the development of new methods and materials that promote skeletal muscle repair and functional regeneration to bring approaches closer to therapies in the clinic that structurally and functionally repair muscle. The combination of stem cells, biomaterials, and biomolecules is used to induce skeletal muscle regeneration. In this review, we provide an overview of different cell types used to treat skeletal muscle injury, highlight current strategies in biomaterial-based approaches, the importance of topography for the successful creation of functional striated muscle fibers, and discuss novel methods for muscle regeneration and challenges for their future clinical implementation.
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Affiliation(s)
- Emine Alarcin
- Department of Pharmaceutical Technology, Faculty of Pharmacy, Marmara University, 34854 Istanbul, Turkey;
| | - Ayca Bal-Öztürk
- Department of Analytical Chemistry, Faculty of Pharmacy, Istinye University, 34010 Istanbul, Turkey;
- Department of Stem Cell and Tissue Engineering, Institute of Health Sciences, Istinye University, 34010 Istanbul, Turkey
| | - Hüseyin Avci
- Department of Metallurgical and Materials Engineering, Eskisehir Osmangazi University, 26040 Eskisehir, Turkey;
- Cellular Therapy and Stem Cell Research Center, Eskisehir Osmangazi University, 26040 Eskisehir, Turkey
- AvciBio Research Group, Eskisehir Osmangazi University, 26040 Eskisehir, Turkey;
- Translational Medicine Research and Clinical Center, Eskisehir Osmangazi University, 26040 Eskisehir, Turkey
| | - Hamed Ghorbanpoor
- AvciBio Research Group, Eskisehir Osmangazi University, 26040 Eskisehir, Turkey;
- Department of Biomedical Engineering, Ankara Yildirim Beyazit University, 06010 Ankara, Turkey;
- Department of Biomedical Engineering, Eskisehir Osmangazi University, 26040 Eskisehir, Turkey
| | - Fatma Dogan Guzel
- Department of Biomedical Engineering, Ankara Yildirim Beyazit University, 06010 Ankara, Turkey;
| | - Ali Akpek
- Department of Bioengineering, Gebze Technical University, 41400 Gebze, Turkey; (A.A.); (G.Y.)
| | - Gözde Yesiltas
- Department of Bioengineering, Gebze Technical University, 41400 Gebze, Turkey; (A.A.); (G.Y.)
| | - Tuba Canak-Ipek
- Department of Thoracic and Cardiovascular Surgery, University Hospital Tuebingen, Calwerstraße 7/1, 72076 Tuebingen, Germany;
| | - Meltem Avci-Adali
- Department of Thoracic and Cardiovascular Surgery, University Hospital Tuebingen, Calwerstraße 7/1, 72076 Tuebingen, Germany;
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14
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Kumar D, Anand T, Talluri TR, Kues WA. Potential of transposon-mediated cellular reprogramming towards cell-based therapies. World J Stem Cells 2020; 12:527-544. [PMID: 32843912 PMCID: PMC7415244 DOI: 10.4252/wjsc.v12.i7.527] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/26/2020] [Revised: 05/09/2020] [Accepted: 05/28/2020] [Indexed: 02/07/2023] Open
Abstract
Induced pluripotent stem (iPS) cells present a seminal discovery in cell biology and promise to support innovative treatments of so far incurable diseases. To translate iPS technology into clinical trials, the safety and stability of these reprogrammed cells needs to be shown. In recent years, different non-viral transposon systems have been developed for the induction of cellular pluripotency, and for the directed differentiation into desired cell types. In this review, we summarize the current state of the art of different transposon systems in iPS-based cell therapies.
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Affiliation(s)
- Dharmendra Kumar
- Animal Physiology and Reproduction Division, ICAR-Central Institute for Research on Buffaloes, Hisar 125001, India.
| | - Taruna Anand
- NCVTC, ICAR-National Research Centre on Equines, Hisar 125001, India
| | - Thirumala R Talluri
- Equine Production Campus, ICAR-National Research Centre on Equines, Bikaner 334001, India
| | - Wilfried A Kues
- Friedrich-Loeffler-Institut, Institute of Farm Animal Genetics, Department of Biotechnology, Mariensee 31535, Germany
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15
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Baci D, Chirivì M, Pace V, Maiullari F, Milan M, Rampin A, Somma P, Presutti D, Garavelli S, Bruno A, Cannata S, Lanzuolo C, Gargioli C, Rizzi R, Bearzi C. Extracellular Vesicles from Skeletal Muscle Cells Efficiently Promote Myogenesis in Induced Pluripotent Stem Cells. Cells 2020; 9:cells9061527. [PMID: 32585911 PMCID: PMC7349204 DOI: 10.3390/cells9061527] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/19/2020] [Revised: 06/13/2020] [Accepted: 06/15/2020] [Indexed: 12/11/2022] Open
Abstract
The recent advances, offered by cell therapy in the regenerative medicine field, offer a revolutionary potential for the development of innovative cures to restore compromised physiological functions or organs. Adult myogenic precursors, such as myoblasts or satellite cells, possess a marked regenerative capacity, but the exploitation of this potential still encounters significant challenges in clinical application, due to low rate of proliferation in vitro, as well as a reduced self-renewal capacity. In this scenario, induced pluripotent stem cells (iPSCs) can offer not only an inexhaustible source of cells for regenerative therapeutic approaches, but also a valuable alternative for in vitro modeling of patient-specific diseases. In this study we established a reliable protocol to induce the myogenic differentiation of iPSCs, generated from pericytes and fibroblasts, exploiting skeletal muscle-derived extracellular vesicles (EVs), in combination with chemically defined factors. This genetic integration-free approach generates functional skeletal myotubes maintaining the engraftment ability in vivo. Our results demonstrate evidence that EVs can act as biological "shuttles" to deliver specific bioactive molecules for a successful transgene-free differentiation offering new opportunities for disease modeling and regenerative approaches.
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Affiliation(s)
- Denisa Baci
- Institute of Biochemistry and Cell Biology, National Research Council, 00015 Rome, Italy; (D.B.); (M.C.); (V.P.); (M.M.); (A.R.); (D.P.)
- Department of Biotechnology and Life Sciences, University of Insubria, 21100 Varese, Italy
| | - Maila Chirivì
- Institute of Biochemistry and Cell Biology, National Research Council, 00015 Rome, Italy; (D.B.); (M.C.); (V.P.); (M.M.); (A.R.); (D.P.)
| | - Valentina Pace
- Institute of Biochemistry and Cell Biology, National Research Council, 00015 Rome, Italy; (D.B.); (M.C.); (V.P.); (M.M.); (A.R.); (D.P.)
| | | | - Marika Milan
- Institute of Biochemistry and Cell Biology, National Research Council, 00015 Rome, Italy; (D.B.); (M.C.); (V.P.); (M.M.); (A.R.); (D.P.)
| | - Andrea Rampin
- Institute of Biochemistry and Cell Biology, National Research Council, 00015 Rome, Italy; (D.B.); (M.C.); (V.P.); (M.M.); (A.R.); (D.P.)
| | - Paolo Somma
- Flow Cytometry Core, Humanitas Clinical and Research Center, 20089 Milan, Italy;
| | - Dario Presutti
- Institute of Biochemistry and Cell Biology, National Research Council, 00015 Rome, Italy; (D.B.); (M.C.); (V.P.); (M.M.); (A.R.); (D.P.)
| | - Silvia Garavelli
- Institute for Endocrinology and Oncology “Gaetano Salvatore”, National Research Council, 80131 Naples, Italy;
| | | | - Stefano Cannata
- Department of Biology, University of Rome Tor Vergata, 00133 Rome, Italy; (S.C.); (C.G.)
| | - Chiara Lanzuolo
- Institute of Biomedical Technologies, National Research Council, 20090 Milan, Italy;
- Fondazione Istituto Nazionale di Genetica Molecolare, 20122 Milan, Italy
| | - Cesare Gargioli
- Department of Biology, University of Rome Tor Vergata, 00133 Rome, Italy; (S.C.); (C.G.)
| | - Roberto Rizzi
- Institute of Biomedical Technologies, National Research Council, 20090 Milan, Italy;
- Fondazione Istituto Nazionale di Genetica Molecolare, 20122 Milan, Italy
- Correspondence: (R.R.); (C.B.); Tel.: +39-02-0066-0230 (R.R.); +39-02-0066-0230 (C.B.)
| | - Claudia Bearzi
- Institute of Biochemistry and Cell Biology, National Research Council, 00015 Rome, Italy; (D.B.); (M.C.); (V.P.); (M.M.); (A.R.); (D.P.)
- Fondazione Istituto Nazionale di Genetica Molecolare, 20122 Milan, Italy
- Correspondence: (R.R.); (C.B.); Tel.: +39-02-0066-0230 (R.R.); +39-02-0066-0230 (C.B.)
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16
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Slanzi A, Iannoto G, Rossi B, Zenaro E, Constantin G. In vitro Models of Neurodegenerative Diseases. Front Cell Dev Biol 2020; 8:328. [PMID: 32528949 PMCID: PMC7247860 DOI: 10.3389/fcell.2020.00328] [Citation(s) in RCA: 149] [Impact Index Per Article: 29.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/11/2020] [Accepted: 04/16/2020] [Indexed: 12/12/2022] Open
Abstract
Neurodegenerative diseases are progressive degenerative conditions characterized by the functional deterioration and ultimate loss of neurons. These incurable and debilitating diseases affect millions of people worldwide, and therefore represent a major global health challenge with severe implications for individuals and society. Recently, several neuroprotective drugs have failed in human clinical trials despite promising pre-clinical data, suggesting that conventional cell cultures and animal models cannot precisely replicate human pathophysiology. To bridge the gap between animal and human studies, three-dimensional cell culture models have been developed from human or animal cells, allowing the effects of new therapies to be predicted more accurately by closely replicating some aspects of the brain environment, mimicking neuronal and glial cell interactions, and incorporating the effects of blood flow. In this review, we discuss the relative merits of different cerebral models, from traditional cell cultures to the latest high-throughput three-dimensional systems. We discuss their advantages and disadvantages as well as their potential to investigate the complex mechanisms of human neurodegenerative diseases. We focus on in vitro models of the most frequent age-related neurodegenerative disorders, such as Parkinson’s disease, Alzheimer’s disease and prion disease, and on multiple sclerosis, a chronic inflammatory neurodegenerative disease affecting young adults.
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Affiliation(s)
- Anna Slanzi
- Department of Medicine, University of Verona, Verona, Italy
| | - Giulia Iannoto
- Department of Medicine, University of Verona, Verona, Italy
| | - Barbara Rossi
- Department of Medicine, University of Verona, Verona, Italy
| | - Elena Zenaro
- Department of Medicine, University of Verona, Verona, Italy
| | - Gabriela Constantin
- Department of Medicine, University of Verona, Verona, Italy.,Center for Biomedical Computing (CBMC), University of Verona, Verona, Italy
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17
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Jin Y, Shen Y, Su X, Weintraub NL, Tang Y. Effective restoration of dystrophin expression in iPSC Mdx-derived muscle progenitor cells using the CRISPR/Cas9 system and homology-directed repair technology. Comput Struct Biotechnol J 2020; 18:765-773. [PMID: 32280431 PMCID: PMC7132053 DOI: 10.1016/j.csbj.2020.03.012] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2019] [Revised: 02/14/2020] [Accepted: 03/17/2020] [Indexed: 12/20/2022] Open
Abstract
Duchenne muscular dystrophy (DMD) is a progressive myopathic disease caused by mutations in the gene encoding dystrophin protein that eventually leads to the exhaustion of myogenic progenitor cells (MPC). Autologous induced pluripotent stem cells (iPSCs) provide an endless source of MPC, which can potentially replenish the progenitor cell pool, repair muscle damage, and prevent DMD progression. Deletion of mutant exon 23 (ΔEx23) with clustered regularly interspaced short palindromic repeats/CRISPR-associated 9 (CRISPR/Cas9) gene-editing technology can correct dystrophin gene expression in iPSCs. However, successful exon23 deletion and clonal isolation are very inefficient (~3%), and manual selection of each iPSC clone and genotyping to identify ΔEx23 is labor-intensive. To overcome these obstacles, we added a homology-directed repair (HDR) donor vector, which carries floxed fluorescent protein and antibiotic selection genes, thus allowing us to identify ΔEx23 iPSC with donor selective gene integration. Our results indicate that the HDR-mediated targeted integration enables ΔEx23 iPSC identification; the HDR donor vector increased the recognition efficiency of clonal isolation (>90% as confirmed by Sanger sequencing). After removal of the inserted genes by Cre-mediated recombination followed by doxycycline (Dox)-induced MyoD induction, ΔEx23 iPSC differentiated into MPC with restored dystrophin expression in vitro. Importantly, transplanted ΔEx23 iPSC-MPC express dystrophin in the muscles of a mouse model of DMD (Mdx mice). In conclusion, the use of HDR donor vector increased the efficiency of ΔEx23 gene correction by CRISPR/Cas9, and facilitate the identification of successfully edited iPSC clones for cell therapy of DMD.
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Affiliation(s)
| | | | | | | | - Yaoliang Tang
- Medical College of Georgia, Augusta University, Augusta, GA, USA
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18
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Liu M, Li X, Zhou C, Wang M, Wang H, Ding H, Cheng L, Gan L, Wu X, Du Z. Thioredoxin mitigates H 2 O 2 -induced inhibition of myogenic differentiation of rat bone marrow mesenchymal stem cells by enhancing AKT activation. FEBS Open Bio 2020; 10:835-846. [PMID: 32160414 PMCID: PMC7193161 DOI: 10.1002/2211-5463.12835] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/12/2019] [Revised: 02/06/2020] [Accepted: 03/06/2020] [Indexed: 11/20/2022] Open
Abstract
Thioredoxin (Trx) is a hydrogen acceptor of ribonucleotide reductase and a regulator of some enzymes and receptors. It has been previously shown that significantly elevated levels of Trx expression are associated with the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs), but it is not clear how Trx regulates the effects of hydrogen peroxide (H2O2) on myogenic differentiation of BMSCs. Here, we report that rat BMSCs treated with a high dose (150 µm) of H2O2 exhibited a significant reduction in viability, cell cycling, and superoxide dismutase and glutathione peroxidase levels, and an increase in reactive oxygen species and malondialdehyde levels, which was accompanied by reductions in protein kinase B activation and forkhead Box O1, myogenic differentiation 1 and myogenin expression during myogenic differentiation. Furthermore, treatment with recombinant human Trx significantly mitigated the effects of H2O2 on the myogenic differentiation of BMSCs, and this was abrogated by cotreatment with wortmannin [a specific phosphatidylinositol 3‐kinase inhibitor]. In summary, our results suggest that treatment with recombinant human Trx mitigates H2O2‐induced oxidative stress and may promote myogenic differentiation of rat BMSCs by enhancing phosphatidylinositol 3‐kinase/protein kinase B/forkhead Box O1 signaling.
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Affiliation(s)
- Meiling Liu
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, China
| | - Xianglu Li
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, China
| | - Changlin Zhou
- Department of Orthopedics, The Second Affiliated Hospital of Harbin Medical University, China
| | - Manfeng Wang
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, China
| | - Hongzhi Wang
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, China
| | - Haifeng Ding
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, China
| | - Luyang Cheng
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, China
| | - Lu Gan
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, China
| | - Xiaowei Wu
- Department of Geriatrics, The Second Affiliated Hospital of Harbin Medical University, China
| | - Zhimin Du
- Institute of Clinical Pharmacology, The Second Affiliated Hospital of Harbin Medical University, China.,State Key Laboratory of Quality Research in Chinese Medicines, Macau University of Science and Technology, Harbin, China
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19
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Chlebanowska P, Tejchman A, Sułkowski M, Skrzypek K, Majka M. Use of 3D Organoids as a Model to Study Idiopathic Form of Parkinson's Disease. Int J Mol Sci 2020; 21:ijms21030694. [PMID: 31973095 PMCID: PMC7037292 DOI: 10.3390/ijms21030694] [Citation(s) in RCA: 57] [Impact Index Per Article: 11.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2019] [Revised: 01/10/2020] [Accepted: 01/17/2020] [Indexed: 02/07/2023] Open
Abstract
Organoids are becoming particularly popular in modeling diseases that are difficult to reproduce in animals, due to anatomical differences in the structure of a given organ. Thus, they are a bridge between the in vitro and in vivo models. Human midbrain is one of the structures that is currently being intensively reproduced in organoids for modeling Parkinson’s disease (PD). Thanks to three-dimensional (3D) architecture and the use of induced pluripotent stem cells (iPSCs) differentiation into organoids, it has been possible to recapitulate a complicated network of dopaminergic neurons. In this work, we present the first organoid model for an idiopathic form of PD. iPSCs were generated from peripheral blood mononuclear cells of healthy volunteers and patients with the idiopathic form of PD by transduction with Sendai viral vector. iPSCs were differentiated into a large multicellular organoid-like structure. The mature organoids displayed expression of neuronal early and late markers. Interestingly, we observed statistical differences in the expression levels of LIM homeobox transcription factor alpha (early) and tyrosine hydroxylase (late) markers between organoids from PD patient and healthy volunteer. The obtained results show immense potential for the application of 3D human organoids in studying the neurodegenerative disease and modeling cellular interactions within the human brain.
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20
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Breuls N, Giacomazzi G, Sampaolesi M. (Epi)genetic Modifications in Myogenic Stem Cells: From Novel Insights to Therapeutic Perspectives. Cells 2019; 8:cells8050429. [PMID: 31075875 PMCID: PMC6562881 DOI: 10.3390/cells8050429] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/09/2019] [Revised: 05/06/2019] [Accepted: 05/07/2019] [Indexed: 12/17/2022] Open
Abstract
The skeletal muscle is considered to be an ideal target for stem cell therapy as it has an inherent regenerative capacity. Upon injury, the satellite cells, muscle stem cells that reside under the basal lamina of the myofibres, start to differentiate in order to reconstitute the myofibres while maintaining the initial stem cell pool. In recent years, it has become more and more evident that epigenetic mechanisms such as histon modifications, DNA methylations and microRNA modulations play a pivatol role in this differentiation process. By understanding the mechanisms behind myogenesis, researchers are able to use this knowledge to enhance the differentiation and engraftment potential of different muscle stem cells. Besides manipulation on an epigenetic level, recent advances in the field of genome-engineering allow site-specific modifications in the genome of these stem cells. Combining epigenetic control of the stem cell fate with the ability to site-specifically correct mutations or add genes for further cell control, can increase the use of stem cells as treatment of muscular dystrophies drastically. In this review, we will discuss the advances that have been made in genome-engineering and the epigenetic regulation of muscle stem cells and how this knowledge can help to get stem cell therapy to its full potential.
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Affiliation(s)
- Natacha Breuls
- Translational Cardiomyology Lab, Department of Development and Regeneration, Stem Cell Institute Leuven, 3000 KU Leuven, Belgium.
| | - Giorgia Giacomazzi
- Translational Cardiomyology Lab, Department of Development and Regeneration, Stem Cell Institute Leuven, 3000 KU Leuven, Belgium.
| | - Maurilio Sampaolesi
- Translational Cardiomyology Lab, Department of Development and Regeneration, Stem Cell Institute Leuven, 3000 KU Leuven, Belgium.
- Human Anatomy Unit, Department of Public Health, Experimental and Forensic Medicine, and Interuniversity Institute of Myology, University of Pavia, 27100 Pavia, Italy.
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21
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Piga D, Salani S, Magri F, Brusa R, Mauri E, Comi GP, Bresolin N, Corti S. Human induced pluripotent stem cell models for the study and treatment of Duchenne and Becker muscular dystrophies. Ther Adv Neurol Disord 2019; 12:1756286419833478. [PMID: 31105767 PMCID: PMC6501480 DOI: 10.1177/1756286419833478] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/21/2018] [Accepted: 11/27/2018] [Indexed: 12/31/2022] Open
Abstract
Duchenne and Becker muscular dystrophies are the most common muscle diseases and are both currently incurable. They are caused by mutations in the dystrophin gene, which lead to the absence or reduction/truncation of the encoded protein, with progressive muscle degeneration that clinically manifests in muscle weakness, cardiac and respiratory involvement and early death. The limits of animal models to exactly reproduce human muscle disease and to predict clinically relevant treatment effects has prompted the development of more accurate in vitro skeletal muscle models. However, the challenge of effectively obtaining mature skeletal muscle cells or satellite stem cells as primary cultures has hampered the development of in vitro models. Here, we discuss the recently developed technologies that enable the differentiation of skeletal muscle from human induced pluripotent stem cells (iPSCs) of Duchenne and Becker patients. These systems recapitulate key disease features including inflammation and scarce regenerative myogenic capacity that are partially rescued by genetic and pharmacological therapies and can provide a useful platform to study and realize future therapeutic treatments. Implementation of this model also takes advantage of the developing genome editing field, which is a promising approach not only for correcting dystrophin, but also for modulating the underlying mechanisms of skeletal muscle development, regeneration and disease. These data prove the possibility of creating an accurate Duchenne and Becker in vitro model starting from iPSCs, to be used for pathogenetic studies and for drug screening to identify strategies capable of stopping or reversing muscular dystrophinopathies and other muscle diseases.
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Affiliation(s)
- Daniela Piga
- Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), Neurology Unit, IRCCS Foundation Ca’ Granda Ospedale Maggiore Policlinico, University of Milan, Milan, Italy
| | - Sabrina Salani
- Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), Neurology Unit, IRCCS Foundation Ca’ Granda Ospedale Maggiore Policlinico, University of Milan, Milan, Italy
| | - Francesca Magri
- Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), Neurology Unit, IRCCS Foundation Ca’ Granda Ospedale Maggiore Policlinico, University of Milan, Milan, Italy
| | - Roberta Brusa
- Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), Neurology Unit, IRCCS Foundation Ca’ Granda Ospedale Maggiore Policlinico, University of Milan, Milan, Italy
| | - Eleonora Mauri
- Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), Neurology Unit, IRCCS Foundation Ca’ Granda Ospedale Maggiore Policlinico, University of Milan, Milan, Italy
| | - Giacomo P. Comi
- Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), Neurology Unit, IRCCS Foundation Ca’ Granda Ospedale Maggiore Policlinico, University of Milan, Milan, Italy
| | - Nereo Bresolin
- Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), Neurology Unit, IRCCS Foundation Ca’ Granda Ospedale Maggiore Policlinico, University of Milan, Milan, Italy
| | - Stefania Corti
- Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), Neurology Unit, IRCCS Foundation Ca’ Granda Ospedale Maggiore Policlinico, University of Milan, Via Francesco Sforza 35, 20122, Milan, Italy
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In Vitro Evaluation of Exon Skipping in Disease-Specific iPSC-Derived Myocytes. Methods Mol Biol 2019; 1828:173-189. [PMID: 30171542 DOI: 10.1007/978-1-4939-8651-4_11] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/17/2023]
Abstract
Patient-derived disease-specific induced pluripotent stem cells (iPSCs) have opened the door to recreating pathological conditions in vitro using differentiation into diseased cells corresponding to each target tissue. To investigate muscular disease, we have established a myogenic differentiation protocol mediated by inducible MYOD1 expression that drives human iPSCs into myocytes. This highly reproducible differentiation protocol yields a homogenous skeletal muscle cell population, reaching efficiencies as high as 70-90%. Such high efficiency enables us to evaluate the efficacy of exon skipping in disease-specific myocytes. These disease-specific iPSC-derived myocytes can be applied not only for the validation of therapeutic efficacy of specific antisense oligonucleotide but also for the screening of exon skipping chemicals combined with the multiwell differentiation system.
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Chua MWJ, Yildirim ED, Tan JHE, Chua YJB, Low SMC, Ding SLS, Li CW, Jiang Z, Teh BT, Yu K, Shyh-Chang N. Assessment of different strategies for scalable production and proliferation of human myoblasts. Cell Prolif 2019; 52:e12602. [PMID: 30891802 PMCID: PMC6536385 DOI: 10.1111/cpr.12602] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/19/2018] [Revised: 01/11/2019] [Accepted: 01/16/2019] [Indexed: 12/13/2022] Open
Abstract
OBJECTIVES Myoblast transfer therapy (MTT) is a technique to replace muscle satellite cells with genetically repaired or healthy myoblasts, to treat muscular dystrophies. However, clinical trials with human myoblasts were ineffective, showing almost no benefit with MTT. One important obstacle is the rapid senescence of human myoblasts. The main purpose of our study was to compare the various methods for scalable generation of proliferative human myoblasts. METHODS We compared the immortalization of primary myoblasts with hTERT, cyclin D1 and CDK4R24C , two chemically defined methods for deriving myoblasts from pluripotent human embryonic stem cells (hESCs), and introduction of viral MyoD into hESC-myoblasts. RESULTS Our results show that, while all the strategies above are suboptimal at generating bona fide human myoblasts that can both proliferate and differentiate robustly, chemically defined hESC-monolayer-myoblasts show the most promise in differentiation potential. CONCLUSIONS Further efforts to optimize the chemically defined differentiation of hESC-monolayer-myoblasts would be the most promising strategy for the scalable generation of human myoblasts, for applications in MTT and high-throughput drug screening.
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Affiliation(s)
- Min-Wen Jason Chua
- NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, Singapore City, Singapore.,Stem Cell & Regenerative Biology, Genome Institute of Singapore, Agency for Science Technology and Research, Singapore City, Singapore.,Laboratory of Cancer Therapeutics, Program in Cancer and Stem Cell Biology, Duke-NUS Medical School, Singapore City, Singapore.,Institute of Molecular and Cell Biology, Agency for Science Technology and Research, Singapore City, Singapore.,Division of Medical Science, Laboratory of Cancer Epigenome, National Cancer Centre Singapore, Singapore City, Singapore
| | - Ege Deniz Yildirim
- Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore City, Singapore
| | - Jun-Hao Elwin Tan
- NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, Singapore City, Singapore.,Stem Cell & Regenerative Biology, Genome Institute of Singapore, Agency for Science Technology and Research, Singapore City, Singapore.,Institute of Molecular and Cell Biology, Agency for Science Technology and Research, Singapore City, Singapore.,Division of Medical Science, Laboratory of Cancer Epigenome, National Cancer Centre Singapore, Singapore City, Singapore
| | - Yan-Jiang Benjamin Chua
- NUS Graduate School for Integrative Sciences and Engineering, National University of Singapore, Singapore City, Singapore.,Stem Cell & Regenerative Biology, Genome Institute of Singapore, Agency for Science Technology and Research, Singapore City, Singapore.,Institute of Molecular and Cell Biology, Agency for Science Technology and Research, Singapore City, Singapore.,Division of Medical Science, Laboratory of Cancer Epigenome, National Cancer Centre Singapore, Singapore City, Singapore
| | - Suet-Mei Crystal Low
- Stem Cell & Regenerative Biology, Genome Institute of Singapore, Agency for Science Technology and Research, Singapore City, Singapore
| | - Suet Lee Shirley Ding
- Stem Cell & Regenerative Biology, Genome Institute of Singapore, Agency for Science Technology and Research, Singapore City, Singapore
| | - Chun-Wei Li
- Department of Clinical Nutrition, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China
| | - Zongmin Jiang
- State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China.,University of Chinese Academy of Sciences, Beijing, China.,Institute of Stem cell and Regeneration, Chinese Academy of Sciences, Beijing, China
| | - Bin Tean Teh
- Laboratory of Cancer Therapeutics, Program in Cancer and Stem Cell Biology, Duke-NUS Medical School, Singapore City, Singapore.,Institute of Molecular and Cell Biology, Agency for Science Technology and Research, Singapore City, Singapore.,Division of Medical Science, Laboratory of Cancer Epigenome, National Cancer Centre Singapore, Singapore City, Singapore
| | - Kang Yu
- Department of Clinical Nutrition, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China
| | - Ng Shyh-Chang
- State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China.,University of Chinese Academy of Sciences, Beijing, China.,Institute of Stem cell and Regeneration, Chinese Academy of Sciences, Beijing, China
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24
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The Regenerative Capability of the Urodele Amphibians and Its Potential for Plastic Surgery. Ann Plast Surg 2018; 81:511-515. [DOI: 10.1097/sap.0000000000001619] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/28/2022]
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25
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Abstract
Purpose of Review Muscular dystrophies (MDs) are a spectrum of muscle disorders, which are caused by a number of gene mutations. The studies of MDs are limited due to lack of appropriate models, except for Duchenne muscular dystrophy (DMD), myotonic dystrophy type 1 (DM1), facioscapulohumeral muscular dystrophy (FSHD), and certain type of limb-girdle muscular dystrophy (LGMD). Human induced pluripotent stem cell (iPSC) technologies are emerging to offer a useful model for mechanistic studies, drug discovery, and cell-based therapy to supplement in vivo animal models. This review will focus on current applications of iPSC as disease models of MDs for studies of pathogenic mechanisms and therapeutic development. Recent Findings Many and more human disease-specific iPSCs have been or being established, which carry the natural mutation of MDs with human genomic background. These iPSCs can be differentiated into specific cell types affected in a particular MDs such as skeletal muscle progenitor cells, skeletal muscle fibers, and cardiomyocytes. Human iPSCs are particularly useful for studies of the pathogenicity at the early stage or developmental phase of MDs. High-throughput screening using disease-specific human iPSCs has become a powerful technology in drug discovery. While MD iPSCs have been generated for cell-based replacement therapy, recent advances in genome editing technologies enabled correction of genetic mutations in these cells in culture, raising hope for in vivo genome therapy, which offers a fundamental cure for these daunting inherited MDs. Summary Human disease-specific iPSC models for MDs are emerging as an additional tool to current disease models for elucidating disease mechanisms and developing therapeutic intervention.
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Affiliation(s)
- Guangbin Xia
- Department of Neurology, College of Medicine, University of New Mexico, Albuquerque, NM USA
| | - Naohiro Terada
- Department of Pathology, Immunology & Laboratory Medicine, College of Medicine, Gainesville, FL USA
| | - Tetsuo Ashizawa
- Houston Methodist Neurological Institute and Research Institute, 6670 Bertner Ave R11-117, Houston, TX USA
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26
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Pantelic MN, Larkin LM. Stem Cells for Skeletal Muscle Tissue Engineering. TISSUE ENGINEERING PART B-REVIEWS 2018; 24:373-391. [PMID: 29652595 DOI: 10.1089/ten.teb.2017.0451] [Citation(s) in RCA: 54] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Volumetric muscle loss (VML) is a debilitating condition wherein muscle loss overwhelms the body's normal physiological repair mechanism. VML is particularly common among military service members who have sustained war injuries. Because of the high social and medical cost associated with VML and suboptimal current surgical treatments, there is great interest in developing better VML therapies. Skeletal muscle tissue engineering (SMTE) is a promising alternative to traditional VML surgical treatments that use autogenic tissue grafts, and rather uses isolated stem cells with myogenic potential to generate de novo skeletal muscle tissues to treat VML. Satellite cells are the native precursors to skeletal muscle tissue, and are thus the most commonly studied starting source for SMTE. However, satellite cells are difficult to isolate and purify, and it is presently unknown whether they would be a practical source in clinical SMTE applications. Alternative myogenic stem cells, including adipose-derived stem cells, bone marrow-derived mesenchymal stem cells, perivascular stem cells, umbilical cord mesenchymal stem cells, induced pluripotent stem cells, and embryonic stem cells, each have myogenic potential and have been identified as possible starting sources for SMTE, although they have yet to be studied in detail for this purpose. These alternative stem cell varieties offer unique advantages and disadvantages that are worth exploring further to advance the SMTE field toward highly functional, safe, and practical VML treatments. The following review summarizes the current state of satellite cell-based SMTE, details the properties and practical advantages of alternative myogenic stem cells, and offers guidance to tissue engineers on how alternative myogenic stem cells can be incorporated into SMTE research.
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Affiliation(s)
- Molly N Pantelic
- 1 Department of Molecular and Integrative Physiology, University of Michigan , Ann Arbor, Michigan
| | - Lisa M Larkin
- 1 Department of Molecular and Integrative Physiology, University of Michigan , Ann Arbor, Michigan.,2 Department of Biomedical Engineering, University of Michigan , Ann Arbor, Michigan
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27
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Jevons LA, Houghton FD, Tare RS. Augmentation of musculoskeletal regeneration: role for pluripotent stem cells. Regen Med 2018; 13:189-206. [PMID: 29557248 DOI: 10.2217/rme-2017-0113] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
The rise in the incidence of musculoskeletal diseases is attributed to an increasing ageing population. The debilitating effects of musculoskeletal diseases, coupled with a lack of effective therapies, contribute to huge financial strains on healthcare systems. The focus of regenerative medicine has shifted to pluripotent stem cells (PSCs), namely, human embryonic stem cells and human-induced PSCs, due to the limited success of adult stem cell-based interventions. PSCs constitute a valuable cell source for musculoskeletal regeneration due to their capacity for unlimited self-renewal, ability to differentiate into all cell lineages of the three germ layers and perceived immunoprivileged characteristics. This review summarizes methods for chondrogenic, osteogenic, myogenic and adipogenic differentiation of PSCs and their potential for therapeutic applications.
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Affiliation(s)
- Lauren A Jevons
- Centre for Human Development, Stem Cells & Regeneration, Faculty of Medicine, University of Southampton, Southampton, SO16 6YD, UK
| | - Franchesca D Houghton
- Centre for Human Development, Stem Cells & Regeneration, Faculty of Medicine, University of Southampton, Southampton, SO16 6YD, UK
| | - Rahul S Tare
- Centre for Human Development, Stem Cells & Regeneration, Faculty of Medicine, University of Southampton, Southampton, SO16 6YD, UK.,Department of Mechanical Engineering, Faculty of Engineering & the Environment, University of Southampton, Southampton, SO17 1BJ, UK
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28
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Skeletal Muscle Cell Induction from Pluripotent Stem Cells. Stem Cells Int 2017; 2017:1376151. [PMID: 28529527 PMCID: PMC5424488 DOI: 10.1155/2017/1376151] [Citation(s) in RCA: 40] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/15/2017] [Accepted: 03/28/2017] [Indexed: 12/19/2022] Open
Abstract
Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) have the potential to differentiate into various types of cells including skeletal muscle cells. The approach of converting ESCs/iPSCs into skeletal muscle cells offers hope for patients afflicted with the skeletal muscle diseases such as the Duchenne muscular dystrophy (DMD). Patient-derived iPSCs are an especially ideal cell source to obtain an unlimited number of myogenic cells that escape immune rejection after engraftment. Currently, there are several approaches to induce differentiation of ESCs and iPSCs to skeletal muscle. A key to the generation of skeletal muscle cells from ESCs/iPSCs is the mimicking of embryonic mesodermal induction followed by myogenic induction. Thus, current approaches of skeletal muscle cell induction of ESCs/iPSCs utilize techniques including overexpression of myogenic transcription factors such as MyoD or Pax3, using small molecules to induce mesodermal cells followed by myogenic progenitor cells, and utilizing epigenetic myogenic memory existing in muscle cell-derived iPSCs. This review summarizes the current methods used in myogenic differentiation and highlights areas of recent improvement.
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29
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Helinska A, Krupa M, Archacka K, Czerwinska AM, Streminska W, Janczyk-Ilach K, Ciemerych MA, Grabowska I. Myogenic potential of mouse embryonic stem cells lacking functional Pax7 tested in vitro by 5-azacitidine treatment and in vivo in regenerating skeletal muscle. Eur J Cell Biol 2016; 96:47-60. [PMID: 28017376 DOI: 10.1016/j.ejcb.2016.12.001] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2016] [Revised: 12/05/2016] [Accepted: 12/05/2016] [Indexed: 12/25/2022] Open
Abstract
Regeneration of skeletal muscle relies on the presence of satellite cells. Satellite cells deficiency accompanying some degenerative diseases is the reason for the search for the "replacement cells" that can be used in the muscle therapies. Due to their unique properties embryonic stem cells (ESCs), as well as myogenic cells derived from them, are considered as a promising source of therapeutic cells. Among the factors crucial for the specification of myogenic precursor cells is Pax7 that sustains proper function of satellite cells. In our previous studies we showed that ESCs lacking functional Pax7 are able to form myoblasts in vitro when differentiated within embryoid bodies and their outgrowths. In the current study we showed that ESCs lacking functional Pax7, cultured in vitro in monolayer in the medium supplemented with horse serum and 5azaC, expressed higher levels of factors associated with myogenesis, such as Pdgfra, Pax3, Myf5, and MyoD. Importantly, skeletal myosin immunolocalization confirmed that myogenic differentiation of ESCs was more effective in case of cells lacking Pax7. Our in vivo studies showed that ESCs transplanted into regenerating skeletal muscles were detectable at day 7 of regeneration and the number of Pax7-/- ESCs detected was significantly higher than of control cells. Our results support the concept that lack of functional Pax7 promotes proliferation of differentiating ESCs and for this reason more of them can turn into myogenic lineage.
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Affiliation(s)
- Anita Helinska
- Department of Cytology, Institute of Zoology, Faculty of Biology, University of Warsaw, Poland
| | - Maciej Krupa
- Department of Cytology, Institute of Zoology, Faculty of Biology, University of Warsaw, Poland
| | - Karolina Archacka
- Department of Cytology, Institute of Zoology, Faculty of Biology, University of Warsaw, Poland
| | - Areta M Czerwinska
- Department of Cytology, Institute of Zoology, Faculty of Biology, University of Warsaw, Poland
| | - Wladyslawa Streminska
- Department of Cytology, Institute of Zoology, Faculty of Biology, University of Warsaw, Poland
| | - Katarzyna Janczyk-Ilach
- Department of Cytology, Institute of Zoology, Faculty of Biology, University of Warsaw, Poland
| | - Maria A Ciemerych
- Department of Cytology, Institute of Zoology, Faculty of Biology, University of Warsaw, Poland
| | - Iwona Grabowska
- Department of Cytology, Institute of Zoology, Faculty of Biology, University of Warsaw, Poland.
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30
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Chal J, Al Tanoury Z, Hestin M, Gobert B, Aivio S, Hick A, Cherrier T, Nesmith AP, Parker KK, Pourquié O. Generation of human muscle fibers and satellite-like cells from human pluripotent stem cells in vitro. Nat Protoc 2016; 11:1833-50. [PMID: 27583644 DOI: 10.1038/nprot.2016.110] [Citation(s) in RCA: 187] [Impact Index Per Article: 20.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Progress toward finding a cure for muscle diseases has been slow because of the absence of relevant cellular models and the lack of a reliable source of muscle progenitors for biomedical investigation. Here we report an optimized serum-free differentiation protocol to efficiently produce striated, millimeter-long muscle fibers together with satellite-like cells from human pluripotent stem cells (hPSCs) in vitro. By mimicking key signaling events leading to muscle formation in the embryo, in particular the dual modulation of Wnt and bone morphogenetic protein (BMP) pathway signaling, this directed differentiation protocol avoids the requirement for genetic modifications or cell sorting. Robust myogenesis can be achieved in vitro within 1 month by personnel experienced in hPSC culture. The differentiating culture can be subcultured to produce large amounts of myogenic progenitors amenable to numerous downstream applications. Beyond the study of myogenesis, this differentiation method offers an attractive platform for the development of relevant in vitro models of muscle dystrophies and drug screening strategies, as well as providing a source of cells for tissue engineering and cell therapy approaches.
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Affiliation(s)
- Jérome Chal
- Institut de Génétique et de Biologie Moléculaireet Cellulaire (IGBMC), CNRS (UMR 7104), Inserm U964, Université de Strasbourg, Illkirch-Graffenstaden, France
- Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts, USA
- Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA
- Harvard Stem Cell Institute, Boston, Massachusetts, USA
| | - Ziad Al Tanoury
- Institut de Génétique et de Biologie Moléculaireet Cellulaire (IGBMC), CNRS (UMR 7104), Inserm U964, Université de Strasbourg, Illkirch-Graffenstaden, France
| | - Marie Hestin
- Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts, USA
- Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA
- Harvard Stem Cell Institute, Boston, Massachusetts, USA
| | - Bénédicte Gobert
- Institut de Génétique et de Biologie Moléculaireet Cellulaire (IGBMC), CNRS (UMR 7104), Inserm U964, Université de Strasbourg, Illkirch-Graffenstaden, France
| | - Suvi Aivio
- Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts, USA
- Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA
- Harvard Stem Cell Institute, Boston, Massachusetts, USA
| | - Aurore Hick
- Anagenesis Biotechnologies, Parc d'innovation, Illkirch-Graffenstaden, France
| | - Thomas Cherrier
- Institut de Génétique et de Biologie Moléculaireet Cellulaire (IGBMC), CNRS (UMR 7104), Inserm U964, Université de Strasbourg, Illkirch-Graffenstaden, France
| | - Alexander P Nesmith
- Disease Biophysics Group, Wyss Institute for Biologically Inspired Engineering, School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts, USA
| | - Kevin K Parker
- Disease Biophysics Group, Wyss Institute for Biologically Inspired Engineering, School of Engineering and Applied Sciences, Harvard University, Cambridge, Massachusetts, USA
| | - Olivier Pourquié
- Institut de Génétique et de Biologie Moléculaireet Cellulaire (IGBMC), CNRS (UMR 7104), Inserm U964, Université de Strasbourg, Illkirch-Graffenstaden, France
- Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts, USA
- Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA
- Harvard Stem Cell Institute, Boston, Massachusetts, USA
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31
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Lenzi J, Pagani F, De Santis R, Limatola C, Bozzoni I, Di Angelantonio S, Rosa A. Differentiation of control and ALS mutant human iPSCs into functional skeletal muscle cells, a tool for the study of neuromuscolar diseases. Stem Cell Res 2016; 17:140-7. [PMID: 27318155 PMCID: PMC5009183 DOI: 10.1016/j.scr.2016.06.003] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/09/2015] [Revised: 05/20/2016] [Accepted: 06/07/2016] [Indexed: 12/20/2022] Open
Abstract
Amyotrophic Lateral Sclerosis (ALS) is a severe and fatal neurodegenerative disease characterized by progressive loss of motoneurons, muscle atrophy and paralysis. Recent evidence suggests that ALS should be considered as a multi-systemic disease, in which several cell types contribute to motoneuron degeneration. In this view, mutations in ALS linked genes in other neural and non-neural cell types may exert non-cell autonomous effects on motoneuron survival and function. Induced Pluripotent Stem Cells (iPSCs) have been recently derived from several patients with ALS mutations and it has been shown that they can generate motoneurons in vitro, providing a valuable tool to study ALS. However, the potential of iPSCs could be further valorized by generating other cell types that may be relevant to the pathology. In this paper, by taking advantage of a novel inducible system for MyoD expression, we show that both control iPSCs and iPSCs carrying mutations in ALS genes can generate skeletal muscle cells. We provide evidence that both control and mutant iPSC-derived myotubes are functionally active. This in vitro system will be instrumental to dissect the molecular and cellular pathways impairing the complex motoneuron microenvironment in ALS.
A novel method for inducing iPSCs differentiation into muscle is presented Multiple inducible lines can be easily generated by a new transposable vector Both control and iPSCs carrying ALS mutations can generate functional muscle fibers This system will be instrumental to study non-cell autonomous contributions to ALS
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Affiliation(s)
- Jessica Lenzi
- Center for Life Nano Science, Istituto Italiano di Tecnologia, Viale Regina Elena 291, 00161 Rome, Italy; Department of Biology and Biotechnology Charles Darwin, Sapienza University of Rome, P.le A. Moro 5, 00185 Rome, Italy
| | - Francesca Pagani
- Center for Life Nano Science, Istituto Italiano di Tecnologia, Viale Regina Elena 291, 00161 Rome, Italy
| | - Riccardo De Santis
- Center for Life Nano Science, Istituto Italiano di Tecnologia, Viale Regina Elena 291, 00161 Rome, Italy; Department of Biology and Biotechnology Charles Darwin, Sapienza University of Rome, P.le A. Moro 5, 00185 Rome, Italy
| | - Cristina Limatola
- Department of Physiology and Pharmacology, Sapienza University of Rome, P.le A. Moro 5, 00185 Rome, Italy
| | - Irene Bozzoni
- Center for Life Nano Science, Istituto Italiano di Tecnologia, Viale Regina Elena 291, 00161 Rome, Italy; Department of Biology and Biotechnology Charles Darwin, Sapienza University of Rome, P.le A. Moro 5, 00185 Rome, Italy; Institute Pasteur Fondazione Cenci-Bolognetti, Sapienza University of Rome, P.le A. Moro 5, 00185 Rome, Italy
| | - Silvia Di Angelantonio
- Center for Life Nano Science, Istituto Italiano di Tecnologia, Viale Regina Elena 291, 00161 Rome, Italy; Department of Physiology and Pharmacology, Sapienza University of Rome, P.le A. Moro 5, 00185 Rome, Italy
| | - Alessandro Rosa
- Department of Biology and Biotechnology Charles Darwin, Sapienza University of Rome, P.le A. Moro 5, 00185 Rome, Italy.
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Almalki SG, Agrawal DK. Key transcription factors in the differentiation of mesenchymal stem cells. Differentiation 2016; 92:41-51. [PMID: 27012163 DOI: 10.1016/j.diff.2016.02.005] [Citation(s) in RCA: 298] [Impact Index Per Article: 33.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2015] [Revised: 02/15/2016] [Accepted: 02/25/2016] [Indexed: 11/15/2022]
Abstract
Mesenchymal stem cells (MSCs) are multipotent cells that represent a promising source for regenerative medicine. MSCs are capable of osteogenic, chondrogenic, adipogenic and myogenic differentiation. Efficacy of differentiated MSCs to regenerate cells in the injured tissues requires the ability to maintain the differentiation toward the desired cell fate. Since MSCs represent an attractive source for autologous transplantation, cellular and molecular signaling pathways and micro-environmental changes have been studied in order to understand the role of cytokines, chemokines, and transcription factors on the differentiation of MSCs. The differentiation of MSC into a mesenchymal lineage is genetically manipulated and promoted by specific transcription factors associated with a particular cell lineage. Recent studies have explored the integration of transcription factors, including Runx2, Sox9, PPARγ, MyoD, GATA4, and GATA6 in the differentiation of MSCs. Therefore, the overexpression of a single transcription factor in MSCs may promote trans-differentiation into specific cell lineage, which can be used for treatment of some diseases. In this review, we critically discussed and evaluated the role of transcription factors and related signaling pathways that affect the differentiation of MSCs toward adipocytes, chondrocytes, osteocytes, skeletal muscle cells, cardiomyocytes, and smooth muscle cells.
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Affiliation(s)
- Sami G Almalki
- Departments of Biomedical Sciences, Creighton University School of Medicine, Omaha, NE, USA
| | - Devendra K Agrawal
- Clinical and Translational Science, Creighton University School of Medicine, Omaha, NE, USA.
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33
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Świerczek B, Ciemerych MA, Archacka K. From pluripotency to myogenesis: a multistep process in the dish. J Muscle Res Cell Motil 2015; 36:363-75. [PMID: 26715014 PMCID: PMC4762919 DOI: 10.1007/s10974-015-9436-y] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/14/2015] [Accepted: 11/30/2015] [Indexed: 12/11/2022]
Abstract
Pluripotent stem cells (PSCs), such as embryonic stem cells or induced pluripotent stem cells are a promising source of cells for regenerative medicine as they can differentiate into all cell types building a mammalian body. However, protocols leading to efficient and safe in vitro generation of desired cell types must be perfected before PSCs can be used in cell therapies or tissue engineering. In vivo, i.e. in developing mouse embryo or teratoma, PSCs can differentiate into skeletal muscle, but in vitro their spontaneous differentiation into myogenic cells is inefficient. Numerous attempts have been undertaken to enhance this process. Many of them involved mimicking the interactions occurring during embryonic myogenesis. The key regulators of embryonic myogenesis, such as Wnts proteins, fibroblast growth factor 2, and retinoic acid, have been tested to improve the frequency of in vitro myogenic differentiation of PSCs. This review summarizes the current state of the art, comparing spontaneous and directed myogenic differentiation of PSCs as well as the protocols developed this far to facilitate this process.
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Affiliation(s)
- Barbara Świerczek
- Department of Cytology, Faculty of Biology, University of Warsaw, Miecznikowa 1, 02-096, Warsaw, Poland
| | - Maria A Ciemerych
- Department of Cytology, Faculty of Biology, University of Warsaw, Miecznikowa 1, 02-096, Warsaw, Poland
| | - Karolina Archacka
- Department of Cytology, Faculty of Biology, University of Warsaw, Miecznikowa 1, 02-096, Warsaw, Poland.
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