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Stiff matrices enhance myoblast proliferation, reduce differentiation, and alter the response to fluid shear stress in vitro

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Abstract

During myofiber regeneration, myoblasts are continuously subjected to shear stress. It is currently not known whether shear stress affects the regenerative capacity of myoblasts when extracellular matrix (ECM) stiffness increases (e.g. upon aging). Therefore, we aimed to assess (1) whether matrix stiffness and pulsating fluid shear stress affect myoblast proliferation and/or expression of differentiation-associated genes in myoblasts, and (2) whether matrix stiffness modulates the mechanoresponse of myoblasts to pulsating fluid shear stress. Myoblasts were seeded on matrigel-coated polyacrylamide gel matrices of varying stiffness, mimicking young (“soft”, 0.5 kPa) and old ECM (“stiff”, 20 kPa), as well as on matrigel-coated glass matrices with very high stiffness (40 ϺPa), and subjected to 1 h pulsating fluid shear stress (3 Pa/s or 4 Pa/s, 1 Hz). We found enhanced proliferation of myoblasts on stiff matrices, but reduced differentiation compared to myoblasts on soft matrices. Pulsating fluid shear stress significantly upregulated gene expression of proliferation-associated genes C-fos and Il-6, as well as expression of cytoskeletal α-actin in myoblasts seeded on glass. In contrast, pulsating fluid shear stress significantly downregulated gene expression of α-actin and Myogenin in myoblasts seeded on soft matrices. In conclusion, these results suggest that age and disease-associated increased ECM stiffness may contribute to declined regenerative capacity of myoblasts, by reducing their capacity to differentiate into new muscular tissue, at least in the absence of mechanical stimulation.

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Data availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request. All graphs and plots corresponding to the datasets used in this study are included in this published article.

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Acknowledgements

We thank Carla Offringa, Guus Baan and Gerard de Wit from the Laboratory for Myology of the Vrije Universiteit Amsterdam for technical support with experiments. We thank Mohammad Haroon for his technical support on PFF experiments.

Author contributions

All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by Victor van Santen. The first draft of the manuscript was written by Victor van Santen and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

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  1. These authors contributed equally: Astrid D. Bakker, Richard T. Jaspers

Authors and Affiliations

  1. Laboratory for Myology, Department of Human Movement Sciences, Faculty of Behavioral and Movement Sciences, Vrije Universiteit Amsterdam, Amsterdam Movement Sciences, Amsterdam, The Netherlands

    Victor J. B. van Santen & Richard T. Jaspers

  2. Department of Oral Cell Biology, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and Vrije Universiteit Amsterdam, Amsterdam Movement Sciences, Amsterdam, The Netherlands

    Victor J. B. van Santen, Jenneke Klein-Nulend & Astrid D. Bakker

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van Santen, V.J.B., Klein-Nulend, J., Bakker, A.D. et al. Stiff matrices enhance myoblast proliferation, reduce differentiation, and alter the response to fluid shear stress in vitro. Cell Biochem Biophys 80, 161–170 (2022). https://doi.org/10.1007/s12013-021-01050-4

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