Macrophages (MPs) are important for skeletal muscle regeneration in vivo and may exert beneficial effects on myogenic cell growth through mitogenic and antiapoptotic activities in vitro. However, MPs are highly versatile and may exert various, and even opposite, functions depending on their activation state. We studied monocyte (MO)/MP phenotypes and functions during skeletal muscle repair. Selective labeling of circulating MOs by latex beads in CX3CR1GFP/+ mice showed that injured muscle recruited only CX3CR1lo/Ly-6C+ MOs from blood that exhibited a nondividing, F4/80lo, proinflammatory profile. Then, within muscle, these cells switched their phenotype to become proliferating antiinflammatory CX3CR1hi/Ly-6C− cells that further differentiated into F4/80hi MPs. In vitro, phagocytosis of muscle cell debris induced a switch of proinflammatory MPs toward an antiinflammatory phenotype releasing transforming growth factor β1. In co-cultures, inflammatory MPs stimulated myogenic cell proliferation, whereas antiinflammatory MPs exhibited differentiating activity, assessed by both myogenin expression and fusion into myotubes. Finally, depletion of circulating MOs in CD11b–diphtheria toxin receptor mice at the time of injury totally prevented muscle regeneration, whereas depletion of intramuscular F4/80hi MPs at later stages reduced the diameter of regenerating fibers. In conclusion, injured skeletal muscle recruits MOs exhibiting inflammatory profiles that operate phagocytosis and rapidly convert to antiinflammatory MPs that stimulate myogenesis and fiber growth.

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Objective. Skeletal muscle may be the site of a variety of poorly understood immune reactions, particularly after myofiber injury, which is typically observed in inflammatory myopathies. This study was undertaken to explore both the cell dynamics and functions of resident macrophages and dendritic cells (DCs) in damaged muscle, using a mouse model of notexininduced myoinjury to study innate immune cell reactions.Methods. The myeloid cell reaction to notexininduced myoinjury was analyzed by microscopy and flow cytometry. Bone marrow (BM) transplantation studies were used to discriminate resident from exudate monocyte/macrophages. Functional tests included cytokine screening and an alloantigenic mixed leukocyte reaction to assess the antigen-presenting cell (APC) function. Selective resident macrophage depletion was obtained by injection of diphtheria toxin (DT) into CD11b-DT receptor-transgenic mice transplanted with DT-insensitive BM.Results. The connective tissue surrounding mouse muscle/fascicle tissue (the epimysium/perimysium) after deep muscle injury displayed a resident macrophage population of CD11b؉F4/80؉CD11c؊Ly-6C؊ CX3CR1؊ cells, which concentrated first in the epimysium. These resident macrophages were being used by leukocytes as a centripetal migration pathway, and were found to selectively release 2 chemokines, cytokineinduced neutrophil chemoattractant and monocyte chemoattractant protein 1, and to crucially contribute to massive recruitment of neutrophils and monocytes from the blood. Early epimysial inflammation consisted of a predominance of Ly-6C high CX3CR1 Conclusion. The results in this mouse model show that resident macrophages in the muscle epimysium/ perimysium orchestrate the innate immune response to myoinjury, which is linked to adaptive immunity through the formation of inflammatory DCs.Skeletal muscle is the site of immune reactions in the inflammatory myopathies, muscular dystrophy, graft-versus-host disease, intramuscular vaccination, and therapeutic cell or gene transfer. Since muscle tissue lacks major histocompatibility complex (MHC) expression under physiologic conditions, it forms an unusual microenvironment in which immunopathologic mecha-