Activated Macrophages in Repair of Skeletal Muscle Injuries

Macrophages (Mp) are essential for repair and regeneration in numerous tissues, but are also often associated with tissue destruction and fibrosis. These seemingly contradictory roles of macrophages may be related to the markedly different phenotypes that macrophages can assume upon exposure to different stimuli. Two of the best-characterized in vitro phenotypes of Mp are a pro-inflammatory “M1” phenotype produced by exposure to IFNγ and TNFα, and an anti-inflammatory “M2a” phenotype produced by exposure to interleukin IL-4 or IL-13. M2a Mp are frequently termed “wound healing” Mp, and are widely assumed to participate in tissue repair processes, though direct evidence for M2a Mp in vivo is lacking. The guiding hypothesis of this dissertation was that Mp after traumatic skeletal muscle injury exhibit a transition from an M1 to an M2a phenotype, and that modulation of Mp phenotype can be used to promote regeneration and reduce fibrosis in injured muscle. We found that macrophages accumulated in the lacerated mouse muscle for at least 21 days, accompanied by limited myofiber regeneration and persistent collagen deposition. However, muscle macrophages did not exhibit either an M1 or M2a phenotype, but instead upregulated both M1- and M2a-associated genes early after injury, followed by downregulation of all markers examined. Particularly, IL-10 mRNA and protein were markedly elevated in macrophages from 3 day injured muscle. Additionally, the early mixed M1/M2a gene expression was due to expression of both sets of markers by individual macrophages, rather than simultaneous presence of distinct M1 and M2a populations. Importantly, cell therapy with exogenous M1 macrophages produced a dose-dependent reduction in fibrosis in lacerated muscles but had no effect on myofiber regeneration, indicating that manipulating Mp function has potential to reduce fibrosis following traumatic injury. We also found that conditioned medium from M1 Mp increased proliferation of cultured myoblasts and decreased collagen expression in cultured fibroblasts, but these effects did not appear to be mediated by Mp production of the urokinase type plasminogen activator as we had hypothesized. An improved understanding of the reciprocal regulation of Mp phenotype and the tissue repair environment may assist in development of novel therapies to promote healing.