Generation of Skeletal Muscle Organoids from Human Pluripotent Stem Cells to Model Myogenesis and Muscle Regeneration
Abstract
:1. Introduction
2. Results
2.1. Generation and Characterization of hSkMOs from hPSCs
2.2. Identifying and Characterizing Different Types of Skeletal Muscle Stem/Progenitor Cells during Myogenesis in hSkMOs
2.3. Maturation of Skeletal Muscle Cells in hSkMOs
2.4. Toxin-Induced Muscular Degeneration and Regeneration Capacity of hSkMOs
3. Discussion
4. Materials and Methods
4.1. Culture of Human Pluripotent Stem Cells (hPSCs)
4.2. Generation of Human Skeletal Muscle Organoids
4.3. Immunofluorescence Analyses Using Cryosection
4.4. RNA Extraction, Brachyury Reverse Transcription, Quantitative Real-Time RT-PCR Analysis
4.5. Scanning Electron Microscopy (SEM)
4.6. Muscle Injury Procedures and Regeneration
4.7. Quantification and Statistical Analyses
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Primary Antibody Name | Catalogue Number | Company | Host Species | Dilution |
---|---|---|---|---|
Ki67 | 550609 | BD | Mouse | 1:200 |
Ki67 | ab15580 | Abcam | Rabbit | 1:200 |
PAX3 | PAX3 | DSHB | Mouse | 1:100 |
PAX7 | PAX7 | DSHB | Mouse | 1:200 |
MAP2 | ab5392 | Abcam | Chicken | 1:2000 |
MyHC | 5-6-s | DSHB | Mouse | 1:25 |
MYOD | ab133627 | Abcam | Rabbit | 1:500 |
MYOG | ab124800 | Abcam | Rabbit | 1:500 |
TBX6 | AF4744 | R&D Systems | Goat | 1:50 |
TITIN | 9D10 | DSHB | Mouse | 1:200 |
Gene | Forward Primer | Reverse Primer |
---|---|---|
GAPDH | CAAGATCATCAGCAATGCCTCCTG | GCCTGCTTCACCACCTTCTTGA |
OCT4 | GTGGAGGAAGCTGACAACAA | ATTCTCCAGGTTGCCTCTCA |
NANOG | TTTGTGGGCCTGAAGAAAACT | AGGGCTGTCCTGAATAAGCAG |
Brachyury | TTCATAGCGGTGACTGCTTATCA | CACCCCCATTGGGAGTACC |
MSGN1 | CTGCACACCCTCCGGAATT | CTCTGCCGCGGTTAAGGAG |
TBX6 | CATCCACGAGAATTGTACCCG | AGCAATCCAGTTTAGGGGTGT |
PAX3 | AGCTCGGCGGTGTTTTTATCA | CTGCACAGGATCTTGGAGACG |
PAX7 | CGTGCTCAGAATCAAGTTCG | GTCAGGTTCCGACTCCACAT |
MYOD | ACTTTCTGGAGCCCTCCTGGCA | TTTGTTGCACTACACAGCATG |
MYOG | GCCAACCCAGGGGATCAT | CCCGGCTTGGAAGACAATCT |
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Shin, M.-K.; Bang, J.S.; Lee, J.E.; Tran, H.-D.; Park, G.; Lee, D.R.; Jo, J. Generation of Skeletal Muscle Organoids from Human Pluripotent Stem Cells to Model Myogenesis and Muscle Regeneration. Int. J. Mol. Sci. 2022, 23, 5108. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms23095108
Shin M-K, Bang JS, Lee JE, Tran H-D, Park G, Lee DR, Jo J. Generation of Skeletal Muscle Organoids from Human Pluripotent Stem Cells to Model Myogenesis and Muscle Regeneration. International Journal of Molecular Sciences. 2022; 23(9):5108. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms23095108
Chicago/Turabian StyleShin, Min-Kyoung, Jin Seok Bang, Jeoung Eun Lee, Hoang-Dai Tran, Genehong Park, Dong Ryul Lee, and Junghyun Jo. 2022. "Generation of Skeletal Muscle Organoids from Human Pluripotent Stem Cells to Model Myogenesis and Muscle Regeneration" International Journal of Molecular Sciences 23, no. 9: 5108. https://0-doi-org.brum.beds.ac.uk/10.3390/ijms23095108