Which Comes First: Angiogenesis or Myogenesis Following Punch Biopsy Injury?

Skeletal muscle is integral to metabolic homeostasis, movement, and breathing. The microcirculation supplies oxygen and nutrients to myofibers while removing their metabolic byproducts. Following skeletal muscle injury, damaged myofibers and capillaries regenerate to restore tissue structure and function. However, it is unknown if angiogenesis or myogenesis is preferred when blood vessels and myofibers are removed simultaneously and permitted to regenerate. Congruent with the dependence of myofiber function and survival on their microvascular supply, we hypothesized that angiogenesis precedes myogenesis during skeletal muscle regeneration following punch biopsy. Methods The left gluteus maximus muscle (GM) of anesthetized male and female Cdh5‐mTmG reporter mice (age, 4‐5 months) was injured by creating a hole (diameter, 2 mm) through the muscle with a biopsy punch. Mice were studied at four timepoints: uninjured control [0 days post injury (dpi)], 7 dpi, 14 dpi, 21 dpi. Results Using intravital microscopy to observe the GM in anesthetized mice, regenerating microvessels grew into the biopsy zone by 7 dpi with networks traversing the residual gap by 14 dpi; nascent microvessels were perfused at 14 dpi as confirmed with fluorescent markers injected into the circulation. In contrast, myofibers were absent from the injured area at 7 dpi, began regenerating at wound edges by 14 dpi, and spanned the gap at 21 dpi. Confocal imaging and histology confirmed the presence of microvessels and absence of myofibers at 14 dpi, with both tissue components reoccupying the injured area at 21 dpi. Conclusion Angiogenesis precedes myogenesis during regeneration following punch biopsy of skeletal muscle.