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Incorporation of macrophages into engineered skeletal muscle enables enhanced muscle regeneration

Abstract

Adult skeletal muscle has a robust capacity for self-repair, owing to synergies between muscle satellite cells and the immune system. In vitro models of muscle self-repair would facilitate the basic understanding of muscle regeneration and the screening of therapies for muscle disease. Here, we show that the incorporation of macrophages into muscle tissues engineered from adult-rat myogenic cells enables near-complete structural and functional repair after cardiotoxic injury in vitro. First, we show that—in contrast with injured neonatal-derived engineered muscle—adult-derived engineered muscle fails to properly self-repair after injury, even when treated with pro-regenerative cytokines. We then show that rat bone-marrow-derived macrophages or human blood-derived macrophages resident within the in vitro engineered tissues stimulate muscle satellite cell-mediated myogenesis while significantly limiting myofibre apoptosis and degeneration. Moreover, bone-marrow-derived macrophages within engineered tissues implanted in a mouse dorsal window-chamber model augmented blood vessel ingrowth, cell survival, muscle regeneration and contractile function.

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Fig. 1: Function and injury response of adult-rat-derived skeletal muscle constructs.
Fig. 2: Structure, function and Ca2+ transient injury response of adult-derived Mu–BMDM constructs.
Fig. 3: Structural, functional and myogenic recovery from CTX-induced in vitro injury in Mu–BMDM constructs.
Fig. 4: Anti-apoptotic effect of BMDMs leads to enhanced regeneration in vitro.
Fig. 5: In vitro regeneration of muscle constructs under shared-media conditions following CTX injury.
Fig. 6: Cytokine analysis following injury of engineered muscle and the effects of TNFα inhibition on recovery.
Fig. 7: Effect of BMDMs on implanted engineered muscle vascularization, function and survival.

Data availability

All the data supporting the findings of this study are available within the paper and its Supplementary Information.

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Acknowledgements

We acknowledge C. Jackman, A. Khodabukus, L. Li, I. Shadrin, A. Ganapathi, G. Palmer and G. Hanna for technical assistance, and the Light Microscopy and Optical Molecular Imaging and Analysis core facilities at Duke University for use of their resources. We also thank B. Chazaud for granting a protocol for human macrophage derivation. This study was supported by the National Science Foundation’s Graduate Research Fellowship to M.J., and grants AR070543 and AR065873 from the National Institute of Arthritis and Musculoskeletal and Skin Diseases to N.B.

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M.J. and N.B. conceived and designed the research. M.J., N.A., J.T.W., J.Y., Z.S. and C.S. performed the experiments. Y.Q. performed the implantation surgery. M.J., N.A., J.Y., Z.S. and N.B. analysed the results. M.J. and N.B. wrote the manuscript.

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Correspondence to Nenad Bursac.

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Supplementary Video 1

Real-time assessment of engineered muscle regeneration in vitro.

Supplementary Video 2

Perfused ingrown blood vessels within engineered muscle implants.

Supplementary Video 3

In vivo spontaneous calcium transients in engineered muscle implants.

Supplementary Video 4

Ex vivo electrically induced calcium transients in engineered muscle explants.

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Juhas, M., Abutaleb, N., Wang, J.T. et al. Incorporation of macrophages into engineered skeletal muscle enables enhanced muscle regeneration. Nat Biomed Eng 2, 942–954 (2018). https://doi.org/10.1038/s41551-018-0290-2

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