CD68+macrophages are associated with adipogenesis after murine muscle injury and fibrotic material implantation

Abstract Murphy roths large (MRL) mice are known to regenerate skin and skeletal muscle wounds without scar formation but are also susceptible to metabolic dysfunction and obesity. How biomaterials influence immune cell recruitment and immune-adipocyte interaction after muscle injury in MRL mice is yet to be delineated. To investigate this, we performed volumetric muscle loss surgeries (VML) in C57BL/6 (B6) and MRL mice and evaluated immune responses to injury, regenerative (decellularized extracellular matrix, ECM) and fibrotic (polyethylene, PE) material implants using multiparametric flow cytometry, RNA sequencing, and histopathology. In MRL but not B6 mice, we observed white fat deposition at the site of VML. This was correlated with enrichment of adipogenesis gene sets including Adipoq, Cebpa, Pparg and Acsl1. In addition, F4/80-CD68+ macrophages were significantly more abundant in MRL mice at 3 weeks post-injury in PE and saline groups. The same treatments also highly potentiated expression of the lipid associated macrophage gene signature comprising Trem2, Ly6c2, Cd9, Cd63 and Lyz2. Ly6G+ neutrophils were the second most abundant cell type (~ 25% of CD45+ cells) which was corroborated by increased abundance of Vav1, Ccl2, Adam8 and Rac2. In contrast, ECM implantation produced a largely SiglecF+ eosinophilic response (~ 60% of CD45+ cells) and inhibited CD68+ macrophage prevalence and also potentiated expression of the pro-regenerative Arg1, Chil3, Retnla and Gata3 in both B6 and MRL. Taken together, our results suggest a mechanistic role for CD68+ macrophages in promoting adipogenesis after VML. Moving forward, we will analyze adaptive immune responses to injury and material implants using abovementioned methods. Intramural Research Program, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health