Muscle damage elicits a sterile immune response that facilitates complete regeneration. Here, we used mass spectrometry–based lipidomics to map the mediator lipidome during the transition from inflammation to resolution and regeneration in skeletal muscle injury. We observed temporal regulation of glycerophospholipids and production of pro-inflammatory lipid mediators (for example, leukotrienes and prostaglandins) and specialized pro-resolving lipid mediators (for example, resolvins and lipoxins) that were modulated by ibuprofen. These time-dependent profiles were recapitulated in sorted neutrophils and Ly6C^hi and Ly6C^lo muscle-infiltrating macrophages, with a distinct pro-resolving signature observed in Ly6C^lo macrophages. RNA sequencing of macrophages stimulated with resolvin D2 showed similarities to transcriptional changes found during the temporal transition from Ly6C^hi macrophage to Ly6C^lo macrophage. In vivo, resolvin D2 increased Ly6C^lo macrophages and functional improvement of the regenerating muscle. These results reveal dynamic lipid mediator signatures of innate immune cells and provide a proof of concept for their exploitable effector roles in muscle regeneration.