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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Once escaped from the quiescence niche, precursor cells interact with stromal components that support their survival, proliferation, and differentiation. We examined interplays between human myogenic precursor cells (mpc) and monocyte/macrophages (MP), the main stromal cell type observed at site of muscle regeneration. mpc selectively and specifically attracted monocytes in vitro after their release from quiescence, chemotaxis declining with differentiation. A DNA macroarray–based strategy identified five chemotactic factors accounting for 77% of chemotaxis: MP-derived chemokine, monocyte chemoattractant protein-1, fractalkine, VEGF, and the urokinase system. MP showed lower constitutive chemotactic activity than mpc, but attracted monocytes much strongly than mpc upon cross-stimulation, suggesting mpc-induced and predominantly MP-supported amplification of monocyte recruitment. Determination of [3H]thymidine incorporation, oligosomal DNA levels and annexin-V binding showed that MP stimulate mpc proliferation by soluble factors, and rescue mpc from apoptosis by direct contacts. We conclude that once activated, mpc, which are located close by capillaries, initiate monocyte recruitment and interplay with MP to amplify chemotaxis and enhance muscle growth.