A comparison of human skeletal muscle cell maturation in 2D versus 3D culture: A quantitative proteomic study
1MRC-Versus Arthritis Centre for Integrated Research into Musculoskeletal Ageing (CIMA), Department of Musculoskeletal and Ageing Science, Institute of Life Course and Medical Sciences, University of Liverpool, Liverpool, UK.
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Summary
Three-dimensional (3D) muscle cultures mature over time, developing a slow-twitch phenotype by day 8 and a fast-twitch phenotype by day 21. Optimal contractile force in 3D constructs is achieved at 8 days for muscle research.
Area of Science:
- Biomedical Engineering
- Cell Biology
- Muscle Physiology
Background:
- Three-dimensional (3D) culture systems more accurately represent native muscle tissue compared to traditional 2D monolayers.
- Understanding the impact of culture duration and conditions on muscle cell maturity in 3D is vital for advancing research.
Purpose of the Study:
- To compare the differentiation, maturity, and functional characteristics of human skeletal muscle cells cultured in 2D versus 3D environments.
- To identify optimal culture durations for 3D muscle constructs used in functional studies.
Main Methods:
- Human skeletal muscle cells were cultured in 2D and 3D hydrogel models for up to 21 days.
- Quantitative proteomic analysis and functional measurements (passive tension, contractile force) were performed.
- Myoblast differentiation and myotube development were assessed.
Main Results:
- Myoblasts differentiated into myotubes by day 8 in both 2D and 3D cultures.
- 3D constructs showed a slow-twitch phenotype at day 8, shifting to a fast-twitch phenotype by day 21.
- By day 21, 3D cultures exhibited enhanced mitochondrial maturity and extracellular matrix remodeling, but reduced contractile forces compared to earlier time points.
Conclusions:
- 3D culture promotes skeletal muscle myotube maturity, with distinct phenotypic and functional changes over 21 days.
- Culture duration significantly impacts the characteristics of 3D muscle constructs.
- 8 days of differentiation in 3D constructs appears optimal for peak contractile force, offering valuable models for interventions targeting muscle function preservation.