Melatonin-engineered MSCs-exosomes deliver USP4 to stabilise ARNTL and inhibit clock rhythmic ferroptosis for enhanced flap survival.

BACKGROUND: This study investigates the impact of sleep restriction (SR) on flap viability and its underlying mechanisms. It reveals that SR triggers clock rhythmic ferroptosis, which leads to impaired skin barrier function and increased flap necrosis. METHODS: A retrospective analysis of sleep quality in 344 patients undergoing flap surgery proved that SR is a risk factor for flap necrosis. Further research demonstrated that SR increases the level of ferroptosis, disrupts the circadian rhythm of ferroptosis and exacerbates flap damage in human and murine models. RESULTS: In order to address this clinical issue, the use of melatonin (MT)-preconditioned bone marrow mesenchymal stromal cells-derived exosomes (MEXOs) was found to enhance the therapeutic efficacy of flap repair by mitigating clock rhythmic ferroptosis. Mechanistically, MT increased m6A modification to stabilise and enhance the translation of ubiquitin-specific protease 4 (USP4) mRNA within MEXOs. USP4 delivered by MEXOs directly interacted with and deubiquitinated ARNTL, a core circadian regulator, stabilising its protein levels and suppressing ferroptosis in flap. CONCLUSIONS: These findings identify SR-induced clock rhythmic ferroptosis as a critical pathological driver of flap failure and propose a novel exosome-based strategy targeting the USP4-ARNTL axis to enhance skin barrier integrity and flap survival, offering translational potential for clinical reconstructive surgery. KEY POINTS: This study identifies SR-induced clock rhythmic ferroptosis as a pivotal pathological process in flap necrosis. We reveal a potential therapeutic mechanism in which USP4-enriched MEXOs can effectively repair SR-induced flap necrosis. USP4-enriched MEXOs represent a novel therapy for SR-induced flap necrosis by stabilizing ARNTL to inhibit clock rhythmic ferroptosis.