CD95 ligand drives abdominal aortic aneurysm progression through Caspase-8-mediated GSDMD-dependent endothelial pyroptosis: modulation by SRC kinase

Abdominal aortic aneurysm (AAA) progression is closely linked to inflammation and endothelial dysfunction. Our previous study has demonstrated that increased CD95 ligand (CD95L) and its downstream effector Caspase-8 in the aortic tissue, contributed to AAA by modulating inflammation. However, how the CD95L/Caspase-8 modulated aneurysmal inflammation remains poorly understood. This study investigates how CD95L/Caspase-8 signaling drives endothelial Pyroptosis to exacerbate AAA. Using a CaCl₂-induced AAA murine model and primary mouse aortic endothelial cells (MAECs), we demonstrate that CD95L triggers endothelial Pyroptosis, characterized by NLRP3 inflammasome activation, Gasdermin D N-terminal (GSDMD-N) cleavage, and Caspase-8/Caspase 1 activation. Electron microscopy confirmed pyroptotic morphology, while flow cytometry excluded Apoptosis or necrosis. CD95L elevated IL-1β/IL-18 secretion, which was abolished by Caspase-8 siRNA or inhibitor Z-IETD-FMK. Mechanistically, CD95L suppressed Caspase-8 phosphorylation at Tyr380, enabling its activation of GSDMD-dependent Pyroptosis. In vivo, CaCl₂-induced AAA mice exhibited aortic dilation, elastin degradation, and endothelial-specific Pyroptosis, all attenuated by endothelial-targeted Caspase-8 knockdown via AAV9-shRNA. This intervention reduced NLRP3 and GSDMD-N expression while preserving vascular integrity. Similarly, Src kinase activation mitigated Pyroptosis markers and aortic damage. These findings establish CD95L as a key mediator of endothelial Pyroptosis in AAA via Caspase-8 dephosphorylation and NLRP3/GSDMD-N activation. Targeting Caspase-8 or enhancing Src activity represents a promising therapeutic strategy to curb AAA progression by preserving endothelial homeostasis.