Safranal accelerates diabetic wound healing by suppressing ferroptosis through modulation of transcription factor Forkhead Box O3

Background: Diabetic wounds (DW), which represent a serious complication of diabetes mellitus, present significant clinical challenges owing to the paucity of effective therapeutic interventions. Safranal (Saf), a principal bioactive constituent of saffron (Crocus sativus L.), has significant pharmacological properties, such as anti-inflammatory and antioxidant effects. However, its therapeutic potential and mechanism in DW remain unknown.

Purpose: This investigation aimed to explore whether Saf can improve DW and to elucidate its underlying mechanisms.

Methods: The therapeutic effects of Saf on DW were investigated using an established disease model. The mechanism through which Saf ameliorates DW was elucidated by integrated transcriptomic analysis, network pharmacology, and experimental validation. Endothelial cells were identified as key mediators of DW healing through single-cell Sequencing analysis. The DW microenvironment was recapitulated using high glucose-stimulated Human Umbilical Vein Endothelial Cells (HUVECs), and the molecular mechanisms underlying Saf's therapeutic action were systematically deciphered through molecular docking simulations and targeted gene knockout approaches. Finally, potential therapeutic targets were predicted and validated using adeno-associated virus 9 (AAV9)-mediated gene silencing experiments.

Results: In vivo studies demonstrated that Saf significantly enhanced DW healing. Transcriptomic analysis further revealed its potent anti-ferroptotic effects. Mechanistic studies demonstrated that in vitro experiments confirmed Saf exerts its therapeutic effects by activating transcription factor Forkhead Box O3 (FOXO3)-mediated transcriptional activity, thereby effectively suppressing Ferroptosis. Subsequent studies established the FOXO3/ Solute Carrier Family 7 Member 11 (SLC7A11) pathway as the central pathway mediating Saf's anti-ferroptotic effects in DW. Importantly, both in vitro FOXO3 genetic knockdown and in vivo AAV9-mediated FOXO3 silencing through wound site injection completely abolished Saf's dual therapeutic benefits: Ferroptosis inhibition and wound healing promotion.

Conclusion: This study provides the first evidence that Saf promotes DW healing by inhibiting Ferroptosis through the FOXO3/SLC7A11 pathway. These findings offer novel mechanistic insights into Saf's therapeutic potential and highlight its translational promise for clinical applications in DW management.

Angiogenesis; Diabetic wounds; FOXO3/SLC7A11 pathway; Ferroptosis; Natural product; Safranal.