Enhancing Myoblast Fusion and Myotube Diameter in Human 3D Skeletal Muscle Constructs by Electromagnetic Stimulation

Lisanne Terrie; Margherita Burattini; Sandra Van Vlierberghe; Lorenzo Fassina; Lieven Thorrez

doi:10.3389/fbioe.2022.892287

Enhancing Myoblast Fusion and Myotube Diameter in Human 3D Skeletal Muscle Constructs by Electromagnetic Stimulation

Front Bioeng Biotechnol. 2022 Jun 22:10:892287. doi: 10.3389/fbioe.2022.892287. eCollection 2022.

Authors

Lisanne Terrie¹, Margherita Burattini^{1

2}, Sandra Van Vlierberghe³, Lorenzo Fassina⁴, Lieven Thorrez¹

Affiliations

¹ Tissue Engineering Lab, Dep. Development and Regeneration, KU Leuven Kulak, Kortrijk, Belgium.
² Dept. of Surgical Sciences, Dentistry and Maternity, University of Verona, Verona, Italy.
³ Polymer Chemistry & Biomaterials Group, Centre of Macromolecular Chemistry, Dep. of Organic and Macromolecular Chemistry, Ghent University, Ghent, Belgium.
⁴ Dept. of Electrical, Computer and Biomedical Engineering, University of Pavia, Pavia, Italy.

Abstract

Skeletal muscle tissue engineering (SMTE) aims at the in vitro generation of 3D skeletal muscle engineered constructs which mimic the native muscle structure and function. Although native skeletal muscle is a highly dynamic tissue, most research approaches still focus on static cell culture methods, while research on stimulation protocols indicates a positive effect, especially on myogenesis. A more mature muscle construct may be needed especially for the potential applications for regenerative medicine purposes, disease or drug disposition models. Most efforts towards dynamic cell or tissue culture methods have been geared towards mechanical or electrical stimulation or a combination of those. In the context of dynamic methods, pulsed electromagnetic field (PEMF) stimulation has been extensively used in bone tissue engineering, but the impact of PEMF on skeletal muscle development is poorly explored. Here, we evaluated the effects of PEMF stimulation on human skeletal muscle cells both in 2D and 3D experiments. First, PEMF was applied on 2D cultures of human myoblasts during differentiation. In 2D, enhanced myogenesis was observed, as evidenced by an increased myotube diameter and fusion index. Second, 2D results were translated towards 3D bioartificial muscles (BAMs). BAMs were subjected to PEMF for varying exposure times, where a 2-h daily stimulation was found to be effective in enhancing 3D myotube formation. Third, applying this protocol for the entire 16-days culture period was compared to a stimulation starting at day 8, once the myotubes were formed. The latter was found to result in significantly higher myotube diameter, fusion index, and increased myosin heavy chain 1 expression. This work shows the potential of electromagnetic stimulation for enhancing myotube formation both in 2D and 3D, warranting its further consideration in dynamic culturing techniques.

Keywords: bioartificial muscle; biophysical stimuli; human myoblast; myotube; pulsed electromagnetic field; skeletal muscle; tissue engineering.