Chronic macrophage activation derails muscle repair by disrupting mannose-receptor-linked plasticity revealed by endogenous irg1/acod1 tracking

Macrophages are central drivers of chronic inflammation, yet how a sustained inflammatory state alters their function remains unclear. Using GFP knock-in zebrafish targeting irg1/acod1 that marks macrophage activation, we track the dynamic transitions of macrophage states during acute muscle injury under homeostatic and chronically inflamed conditions, induced by genetic mutation of nlrc3l. In the chronic inflammation model, muscle repair is impaired and expression of the mannose receptor mrc1b/cd206 is severely downregulated in a myd88-dependent manner. Two reparative macrophage subtypes, defined by their cellular behavior and single-cell transcriptomics profile, clustering and muscle-encasing, are lost. A chronic infection model recapitulates these defects, underscoring the link to macrophage mrc1b repression. Depleting either mrc1b or macrophages impairs muscle repair. Reinstating normal macrophage states by restoring macrophage nlrc3l expression or ablating myd88-mediated inflammatory pathways rescues muscle repair in nlrc3l mutants. Contrary to conventional discrete states, we identify hybrid M1/M2 macrophage states post-injury. While transient during normal injury response, a pro-inflammatory hybrid state persists during chronic activation, which restricts macrophage heterogeneity, represses mrc1b, and inhibits intracellular cathepsin K accumulation, a hallmark of reparative subtypes. Thus, our study provides mechanistic insight into the dynamics of macrophage activation during muscle injury and repair, and how these processes are modulated under chronic inflammation.