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Combined Effects of Electrical Stimulation and Protein Coatings on Myotube Formation in a Soft Porous Scaffold

Annals of Biomedical Engineering volume 48, pages 734–746 (2020)Cite this article

Abstract

Compared to two-dimensional cell cultures, three-dimensional ones potentially allow recreating natural tissue environments with higher accuracy. The three-dimensional approach is being investigated in the field of tissue engineering targeting the reconstruction of various tissues, among which skeletal muscle. Skeletal muscle is an electroactive tissue which strongly relies upon interactions with the extracellular matrix for internal organization and mechanical function. Studying the optimization of myogenesis in vitro implies focusing on appropriate biomimetic stimuli, as biochemical and electrical ones. Here we present a three-dimensional polyurethane-based soft porous scaffold (porosity ~ 86%) with a Young’s modulus in wet conditions close to the one of natural skeletal muscle tissue (~ 9 kPa). To study the effect of external stimuli on muscle cells, we functionalized the scaffold with extracellular matrix components (laminin and fibronectin) and observed an increase in myoblast proliferation over three days. Furthermore, the combination between laminin coating and electrical stimulation resulted in more spread and thicker myotubes compared to non-stimulated samples and samples receiving the single (non-combined) inputs. These results pave the way to the development of mature muscle tissue within three-dimensional soft scaffolds, through the combination of biochemical and electrical stimuli.

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Abbreviations

3D:

Three-dimensional

DMEM:

Dulbecco’s Modified Eagle’s Medium

DMSO:

Dimethylsulfoxide

ECM:

Extracellular matrix

ES:

Electrical stimulation

EtOH:

Ethanol

FBS:

Fetal bovine serum

FN:

Fibronectin

HMDI:

Hexamethylene diisocyanate

IQR:

Interquartile range

LMN:

Laminin

MDM:

Myoblast differentiation medium

MGM:

Myoblast growth medium

MSM:

Myoblast seeding medium

PBS:

Phosphate buffered saline

PEG6K:

Polyethylene glycol 6000 g/mol

Pt:

Platinum

PU:

Polyurethane

SEM:

Scanning electron microscopy

SM:

Skeletal muscle

SMTE:

Skeletal muscle tissue engineering

TRITC:

Tetramethylrhodamine

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Acknowledgments

The authors would like to thank Federica Elliot for English language editing and review.

Conflict of interest

The authors declare no competing financial interest.

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Authors and Affiliations

  1. The BioRobotics Institute, Scuola Superiore Sant’Anna, Piazza Martiri della Libertà 33, 56127, Pisa, Italy

    Federica Iberite, Lorenzo Vannozzi & Leonardo Ricotti

  2. Tensive S.r.l, Via Timavo 34, Milan, 20124, Italy

    Irini Gerges

  3. Smart Bio-Interfaces, Istituto Italiano di Tecnologia, Viale Rinaldo Piaggio 34, Pontedera, Pisa, 56025, Italy

    Attilio Marino

  4. Interdisciplinary Centre for Nanostructured Materials and Interfaces (CIMaINa) and Department of Physics, Università degli Studi di Milano, Via Celoria 16, Milan, 20133, Italy

    Marco Piazzoni, Tommaso Santaniello & Cristina Lenardi

  5. Department of Excellence in Robotics & AI, Scuola Superiore Sant’Anna, Piazza Martiri della Libertà 33, 56127, Pisa, Italy

    Federica Iberite, Lorenzo Vannozzi & Leonardo Ricotti

Authors
  1. Federica Iberite
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  2. Irini Gerges
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  3. Lorenzo Vannozzi
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Correspondence to Federica Iberite.

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Iberite, F., Gerges, I., Vannozzi, L. et al. Combined Effects of Electrical Stimulation and Protein Coatings on Myotube Formation in a Soft Porous Scaffold. Ann Biomed Eng 48, 734–746 (2020). https://doi.org/10.1007/s10439-019-02397-9

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  • DOI: https://doi.org/10.1007/s10439-019-02397-9

Keywords

  • Three-dimensional scaffold
  • Biophysical stimulation
  • Skeletal muscle
  • Tissue engineering
  • Polyurethane scaffold