A Transcriptionally Distinct Intermediate Activation State Precedes Langerhans Cell Migration from the Epidermis

Langerhans cells (LC) are antigen-presenting cells that form dense epidermal networks for immune surveillance. Upon antigen uptake, LCs are activated and subsequently orchestrate immune responses by migrating into the lymphatic system. However, the regulation of transcriptional programs during activation and the behavior of LCs in vivo during this transition remain poorly understood. Therefore, this study combines single-cell transcriptomic analysis with intravital imaging to reconstruct the activation trajectory of epidermal LCs. In particular, the study generates a high-resolution single-cell transcriptomic dataset comprising over 22,000 high-quality epidermal LCs under both homeostatic and injury conditions. Notably, a distinct intermediate activated state that precedes LC migration was identified and characterized at the level of signaling pathways and transcription factors. The integration of this dataset with external datasets from homeostatic and injured skin reveals wound-specific fibroblasts as the major source of C3, which is the central component of the complement cascade. Moreover, intravital imaging of C3-deficient mice and pegcetacoplan treatment demonstrates that C3 contributes to LC accumulation adjacent to wound sites. Collectively, the study findings identify an activated epidermal LC population and implicate complement signaling in the regulation of LC behavior during skin injury.