Bergapten promotes angiogenesis by enhancing endothelial autophagy and reducing pyroptosis in ischemic skin flap.

BACKGROUND

Necrosis of distal flap tissue is often attributed to ischemic injury. Previous research has indicated that Bergapten (BeG), known for its anti-inflammatory and antioxidant activities, protects tissues from ischemic injury. This investigation aimed to ascertain the beneficial effects of BeG on ischemic flap survival and explore its underlying mechanisms.

METHODS

To assess the survival of ischemic skin flaps, analyses of flap viability were conducted utilizing survival rate evaluations and laser Doppler blood flow (LDBF) detection. RNA sequencing was performed to clarify the underlying molecular processes involved. In addition, angiogenesis, oxidative stress (OS), pyroptosis, transcription factor EB (TFEB)-mediated autophagy, and adenosine AMP-activated protein kinase (AMPK)-transient receptor potential mucolipin 1 (TRPML1)-calcineurin (CaN) signaling were assessed using molecular docking (MD), cellular thermal shift assay (CETSA), and surface plasmon resonance (SPR), Western blot (WB) assays, immunofluorescence, and dihydroethidium (DHE) staining.

RESULTS

The improvement in flap viability due to BeG was associated with the stimulation of autophagy, reduction of OS, and inhibition of pyroptosis. Notably, BeG-mediated enhancement of autophagic flux and increased resistance to OS were crucial for alleviating pyroptosis in vascular endothelial cells (VECs). BeG promoted autophagy flux and reduced endothelial oxidative stress by activating TFEB in ischemic flaps. However, the therapeutic effects of BeG were abolished by adeno-associated virus (AAV)-mediated TFEB knockdown. Additionally, BeG regulated TFEB activity through the AMPK-TRPML1-CaN pathway.

CONCLUSIONS

BeG enhences autophagy and alleviates OS through stimulation of the AMPK-TRPML1-CaN-TFEB signaling cascade, hence improving the survival of ischemic flaps and potentially offering significant clinical implications.