| Autor: |
Vonk LA; Department of Physiology, Amsterdam UMC., Esen O; Department of Physiology, Amsterdam UMC., Yuen M; Department of Physiology, Amsterdam UMC; Discipline of Child and Adolescent Health, Faculty of Health and Medicine, University of Sydney., Kirby TJ; Department of Physiology, Amsterdam UMC; Amsterdam Cardiovascular Sciences, Heart Failures and Arrhythmias, Amsterdam UMC; Amsterdam Movement Sciences, Tissue Function and Regeneration, Amsterdam UMC; t.kirby@amsterdamumc.nl. |
| Jazyk: |
angličtina |
| Zdroj: |
Journal of visualized experiments : JoVE [J Vis Exp] 2023 May 05 (195). Date of Electronic Publication: 2023 May 05. |
| DOI: |
10.3791/65103 |
| Abstrakt: |
In vitro cell culture is a powerful tool to assess cellular processes and test therapeutic strategies. For skeletal muscle, the most common approaches involve either differentiating myogenic progenitor cells into immature myotubes or the short-term ex vivo culture of isolated individual muscle fibers. A key benefit of ex vivo culture over in vitro is the retention of the complex cellular architecture and contractile characteristics. Here, we detail an experimental protocol for the isolation of intact flexor digitorum brevis muscle fibers from mice and their subsequent ex vivo culture. In this protocol, muscle fibers are embedded in a fibrin-based and basement membrane matrix hydrogel to immobilize the fibers and maintain their contractile function. We then describe methods to assess the muscle fiber contractile function using an optics-based, high-throughput contractility system. The embedded muscle fibers are electrically stimulated to induce contractions, after which their functional properties, such as sarcomere shortening and contractile velocity, are assessed using optics-based quantification. Coupling muscle fiber culture with this system allows for high-throughput testing of the effects of pharmacological agents on contractile function and ex vivo studies of genetic muscle disorders. Finally, this protocol can also be adapted to study dynamic cellular processes in muscle fibers using live-cell microscopy. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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