Skeletal muscle injury-responsive macrophage subsets in larval zebrafish

Single-cell RNA-sequencing (scRNA-seq) was applied to identify and characterise macrophage subsets that responded to larval zebrafish muscle injury. The Tg(mpeg1:mCherry) transgenic zebrafish line was utilised to isolate mCherry-expressing macrophages by FACS. Following needle-stab muscle injury of a 4 days post fertilisation (dpf) larvae, the wound site was dissected out at 1, 2, and 3 days post injury (dpi) for macrophage isolation. Macrophages isolated from 4 dpf-uninjured larvae were also included. This analysis led to the identification of 8 discrete clusters of macrophages, one of which corresponded to uninjured macrophages. The 7 wound-present macrophage subsets highlighted greater macrophage heterogeneity than previously described in an in vivo skeletal muscle injury context. Total of 4 different time points were assayed, including a 4 dpf uninjured control and 3 time points during injury-resolution (1-3 dpi).