Decellularised skeletal muscles allow functional muscle regeneration by promoting host cell migration

Anna Urciuolo,Simon Eaton,Paolo De Coppi,Federico Scottoni,Stavros Loukogeorgakis,Silvia Torelli,Ketan Patel,Bert Blaauw,Carla Julia-Vilella,Elena Germinario,Henry Collins-Hooper,Silvia Perin,Luca Urbani,Panagiotis Maghsoudlou,Asllan Gjinovci,Athanasios Tyraskis,Mario Enrique Alvarez Fallas

doi:10.1038/s41598-018-26371-y

Abstract

Pathological conditions affecting skeletal muscle function may lead to irreversible volumetric muscle loss (VML). Therapeutic approaches involving acellular matrices represent an emerging and promising strategy to promote regeneration of skeletal muscle following injury. Here we investigated the ability of three different decellularised skeletal muscle scaffolds to support muscle regeneration in a xenogeneic immune-competent model of VML, in which the EDL muscle was surgically resected. All implanted acellular matrices, used to replace the resected muscles, were able to generate functional artificial muscles by promoting host myogenic cell migration and differentiation, as well as nervous fibres, vascular networks, and satellite cell (SC) homing. However, acellular tissue mainly composed of extracellular matrix (ECM) allowed better myofibre three-dimensional (3D) organization and the restoration of SC pool, when compared to scaffolds which also preserved muscular cytoskeletal structures. Finally, we showed that fibroblasts are indispensable to promote efficient migration and myogenesis by muscle stem cells across the scaffolds in vitro. This data strongly support the use of xenogeneic acellular muscles as device to treat VML conditions in absence of donor cell implementation, as well as in vitro model for studying cell interplay during myogenesis.

Highlights

Pathological conditions affecting skeletal muscle function may lead to irreversible volumetric muscle loss (VML)
For Latrunculin B protocol (LatB), one cycle was sufficient to remove the cellular compartment and preserve the extracellular matrix (ECM) structure, whilst three cycles for Detergent Enzymatic Treatment (DET) and 72 hours of Sodium Dodecyl Sulfate protocol (SDS) perfusion were needed to obtain the same level of decellularisation (Supplementary Fig. 1)
Compared to freshly isolated muscles, DNA content was significantly reduced by all decellularisation protocols (Fig. 1c), a finding confirmed by haematoxylin and eosin (H&E) staining and scanning electron microscopy (SEM)

Summary

Introduction

Pathological conditions affecting skeletal muscle function may lead to irreversible volumetric muscle loss (VML). We showed that fibroblasts are indispensable to promote efficient migration and myogenesis by muscle stem cells across the scaffolds in vitro This data strongly support the use of xenogeneic acellular muscles as device to treat VML conditions in absence of donor cell implementation, as well as in vitro model for studying cell interplay during myogenesis. Acellular tissues –such as pig urinary bladder ECM, have been clinically used to treat VML conditions[24], and only recently acellular skeletal muscle matrices have been tested for the same application in animal model of VML25–27 It still remains a matter of discussion whether the final in vivo outcome of acellular tissues can be influenced by the original tissue from which they are derived and by the specific protocol used for the decellularisation[5,28,29,30]. Preservation of ECM components and 3D topology was the sufficient requirement to drive host cells toward scaffold repopulation, which allowed proper muscular stem cell maintenance, cell differentiation and homing, as well as functional tissue formation

Methods

Results

Conclusion