Uncovering Sex-Related Differences in Skin Macrophage Polarization During Wound Healing in Diabetic Mice

Background: Chronic wounds, such as diabetes-related foot ulcers, arise from delayed wound healing and create significant health and economic burdens. Macrophages regulate healing by shifting between pro- and anti-inflammatory phenotypes, known as macrophage polarization. Sex and diabetes can impair wound healing, but their influence on macrophage phenotype in skin tissue during wound healing remains unclear, which was investigated in this study using a novel two-sex diabetic mouse model. Methods: Diabetes was induced in male and female C57BL/6J mice using low-dose streptozotocin injections and high-fat diet feeding, with chow-fed mice as controls. After 18 weeks, each mouse received four circular full-thickness dorsal skin wounds. The macrophage phenotypes in wounded skin tissues at Day 0 and Day 10 post-wounding were analyzed using mass cytometry with manual gating and automated computational clustering. Results: Male diabetic mice exhibited more severe hyperglycemia and insulin resistance compared to females. Although diabetic mice did not display delayed wound healing, male mice had a greater proportion of total macrophages than females, especially a higher proportion of pro-inflammatory matrix metalloproteinase-9 (MMP-9)+ macrophages and a lower proportion of anti-inflammatory adiponectin receptor 1 (AdipoR1)+ macrophages in male diabetic mice compared to females, indicating an imbalanced polarization towards a pro-inflammatory phenotype that could result in poorer wound healing. Interestingly, computational clustering identified a new pro-inflammatory, pro-healing phenotype (Ly6C+AdipoR1+CD163–CD206–) more abundant in females than males, suggesting this phenotype may play a role in the transition from the inflammatory to the proliferative stage of wound healing. Conclusions: This study demonstrated a significant sex-based difference in macrophage populations, with male diabetic mice showing a pro-inflammatory bias that may impair wound healing, while a unique pro-inflammatory, pro-healing macrophage population more abundant in females could facilitate recovery. Further research is needed to investigate the role of these newly identified phenotypes in regulating impaired wound healing.