Chronic wounds remain a major unmet healthcare challenge, associated with substantial morbidity and economic costs. Therefore, novel treatment strategies and therapeutic approaches need to be urgently developed. Yet, despite the increasingly recognized importance of neurohormonal signaling in skin physiology, the neuroendocrine regulation of cutaneous wound healing has received surprisingly little attention. Human skin, and its appendages, locally express the pleiotropic neurohormone prolactin (PRL), which not only regulates lactation but also hair follicle cycling, angiogenesis, keratinocyte proliferation, and epithelial stem cell functions. Therefore, we examined the effects of PRL in experimentally wounded female human skin organ culture. Overall, this revealed that PRL slightly, but significantly, inhibited epidermal regeneration (reepithelialisation), cytokeratin 6 protein expression and intraepidermal mitochondrial activity (MTCO1 expression), while it promoted keratinocyte terminal differentiation (i.e. involucrin expression) ex vivo. If the current pilot data are confirmed by further studies, PRL may serve as one of the—rarely studied—negative regulators of cutaneous wound healing that control excessive reepithelialisation. This raises the intriguing and clinically relevant question of whether PRL receptor antagonists could actually promote epidermal repair after human skin wounding.