Skeletal muscle regeneration involves macrophage-myoblast bonding

Abstract

Regeneration in adult skeletal muscle relies on the activation, proliferation, and fusion of myogenic precursor cells (MPC), mostly resident satellite cells (SC). However, the regulatory mechanism during this process is still under evaluation, with the final aim to manipulate regeneration when the intrinsic mechanism is corrupted. Furthermore, intercellular connections during skeletal muscle regeneration have not been previously thoroughly documented. Our hypothesis was that a direct and close cellular interaction between SC/MPC and invading myeloid cells is a key step to control regeneration. We tested this hypothesis during different steps of skeletal muscle regeneration: (a) the recruitment of activated SC; (b) the differentiation of MPC; (c) myotubes growth, in a mouse model of crush injury. Samples harvested (3 and 5 days) post-injury were screened by light and confocal microscopy. Ultrastructural analysis was performed by conventional transmission electron microscopy (TEM) and scanning transmission electron microscopy (STEM) followed by 3D modeling of electron tomography (ET) data. This revealed a new type of interaction between macrophages and myogenic cells by direct heterocellular surface apposition over large areas and long linear distances. In the analyzed volume, regions spaced below 20 nm, within molecular range, represented 31% of the macrophage membrane surface and more than 27% of the myotube membrane. The constant interaction throughout all stages of myogenesis suggests a potential new type of regulatory mechanism for the myogenic process. Thus, deciphering structural and molecular mechanisms of SC-macrophage interaction following injury might open promising perspectives for improving muscle healing.

Age‐related ultrastructural changes of the basement membrane in the mouse blood‐brain barrier

“…The 1 mm 3 fragments of brain tissue were fixed by immersion in 4% glutaraldehyde, buffered with 0.1 mol L −1 sodium cacodylate (pH 7.3) at 4°C overnight and 25 pieces were further processed for Epon‐embedding (Agar100 resin, Agar Scientific, Essex, UK) as previously described . Epon‐embedded murine cerebral tissue fragments were sectioned using a Leica EM UC7 ultramicrotome (Leica Microsystems GmbH, Vienna, Austria).…”

“…The 1 mm 3 fragments of brain tissue were fixed by immersion in 4% glutaraldehyde, buffered with 0.1 mol L −1 sodium cacodylate (pH 7.3) at 4°C overnight and 25 pieces were further processed for Epon-embedding (Agar100 resin, Agar Scientific, Essex, UK) as previously described. 15 Epon-embedded murine cerebral tissue fragments were sectioned using a Leica EM UC7 ultramicrotome (Leica Microsystems GmbH, Vienna, Austria). Light microscopy was done on 1-μm-thick sections stained with 1% toluidine blue and representative images were recorded under a Nikon 600 light microscope performed with a 100× objective (oil immersion) with a 11-megapixel camera (AxioCam HRc, Zeiss, Germany).…”

Immunohistochemical Identification of Human Skeletal Muscle Macrophages

“…Recent work nicely details fiber repair in human skeletal muscle in vivo , showing the presence of macrophages, using a pan-macrophage intracellular marker (CD68+), in regenerating zones along injured fibers (Mackey and Kjaer, 2017). Direct interaction between macrophages and satellite cells (Dumont and Frenette, 2013; Ceafalan et al , 2017; Du et al , 2017; Wehling-Henricks et al , 2018), and defects in skeletal muscle regeneration in the absence of macrophage participation (Arnold et al , 2007; Melton et al , 2016), highlight the necessity of these cells for skeletal muscle repair. In vitro , M1 macrophages promote skeletal muscle cell proliferation and M2 macrophages promote differentiation, suggesting that macrophages may play a role in skeletal muscle growth adaptations, as well as repair (Arnold et al , 2007; Saclier et al , 2013b).…”

Stem cells in skeletal muscle growth and regeneration in amniotes and teleosts: Emerging themes

“…In line with these results, Ceafalan et al () used ultrastructural analysis to demonstrate that following muscle crush injury in mice, macrophages and myogenic cells in the wound displayed heterocellular surface appositions (Ceafalan et al, ). The authors suggest that this form of interaction may be an alternative macrophage‐driven myogenic regulatory process (Ceafalan et al, ).…”