Macrophages play important roles in tissue homeostasis, host defense, response to injury, tissue repair, and resolution of inflammation. The long term goals of this project are to understand how macrophage functional phenotypes are regulated to balance host defense and inflammation with homeostatic functions that resolve inflammation and restore tissue integrity. An associated goal is to therapeutically modulate macrophages to promote tissue repair while minimizing inflammatory pathology. Macrophages assume various phenotypes dependent on ontogeny, tissue-specific factors, epigenetic programming, and (micro)environmental challenges. Inflammatory macrophages produce cytokines such as TNF and IL-1 that are important for host defense and inflammation, while reparative and pro-resolution macrophages produce suppressive and growth factors such as IL-10, VEGF and PDGF important for tissue repair, wound healing and resolution of inflammation. Recent high dimensional analyses using flow/mass cytometry and single cell RNA sequencing (scRNAseq) have revealed distinct macrophage populations and functional phenotypes in different tissues and disease states. Wound healing typically progresses through inflammatory, tissue repair, and tissue remodeling stages. Both inflammatory and repair phases, and a well-regulated balance and transition between them, are important for effective wound healing. Excessive inflammation, such as occurs in chronic TNF-driven inflammatory diseases such as rheumatoid arthritis (RA), results in ineffective repair and chronic wounds. Immune cells including macrophages play important roles in the first two phases of wound healing, and modulation of macrophage function to improve tissue repair is an emerging therapeutic strategy. In the previous project period we investigated mechanisms of crossregulation between the inflammatory factor TNF and the repair factor IL-4, using primary human macrophages and mouse models of wound healing. While IL-4 suppressed components of the TNF inflammatory response, the two cytokines synergized to induce genes associated with resolution of inflammation. The balance between an inflammatory and reparative macrophage phenotype was regulated in part by SREBP transcription factors. Myeloid inactivation or pharmacological inhibition of SREBPs accelerated wound healing. scRNAseq identified novel macrophage populations induced by injury and associated with tissue repair. These data provide insights into mechanisms that regulate the transition from inflammation to repair, and support our overarching hypothesis that selective modulation of the immune system can be utilized to improve tissue repair. In this project we will investigate mechanisms that regulate the balance and transition between inflammation and tissue repair. We anticipate that our studies will yield insights that can be used to develop novel therapeutic strategies to enhance wound healing and restore tissue function.