Circ-USP9X interacts with EIF4A3 to promote endothelial cell pyroptosis by regulating GSDMD stability in atherosclerosis

Endothelial pyroptosis is a pathological mechanism of atherosclerosis (AS). Circular RNAs (circRNAs) are vital in AS progression by regulating endothelial cell functions. The study aimed to explore whether circ-USP9x regulated pyroptosis of endothelial cell to involve in AS development and the molecular mechanism. Pyroptosis was determined using lactate dehydrogenase (LDH) assay, enzyme linked immunosorbent assay (ELISA), flow cytometry, propidium iodide (PI) staining assay, and western blot. The mechanism of circ-USP9x was determined using RNA pull-down and RNA binding protein immunoprecipitation (RIP) assays. Results showed that circ-USP9x was upregulated in AS and oxidized low-density lipoprotein (ox-LDL)-treated human umbilical vein endothelial cells (HUVECs). Knockdown of circ-USP9x suppressed ox-LDL induced pyroptosis of HUVECs. Mechanically, circ-USP9x could bind to EIF4A3 in the cytoplasm. Moreover, EIF4A3 was bound to GSDMD and further affects GSDMD stability. Overexpression of EIF4A3 rescued cell pyroptosis induced by circ-USP9x depletion. In short, circ-USP9x interacted with EIF4A3 to enhance GSDMD stability, thus further promoting ox-LDL-induced pyroptosis of HUVECs. These findings suggested that circ-USP9x participates in AS progression and may be a potential therapeutic target for AS.

Automated summary

Endothelial pyroptosis is a pathological mechanism of atherosclerosis, and circular RNA (circRNA) plays a vital role in this progression. This study aimed to investigate the role of circ-USP9x in regulating pyroptosis of endothelial cells and its molecular mechanism in atherosclerosis development. The results showed that circ-USP9x was upregulated in atherosclerosis and ox-LDL-treated HUVECs. Knockdown of circ-USP9x suppressed ox-LDL-induced pyroptosis in HUVECs. Mechanistically, circ-USP9x interacted with EIF4A3 in the cytoplasm, which then affected the stability of GSDMD. Overexpression of EIF4A3 rescued cell pyroptosis induced by circ-USP9x depletion. In conclusion, circ-USP9x enhances GSDMD stability by interacting with EIF4A3 and promotes ox-LDL-induced pyroptosis in HUVECs, suggesting its potential as a therapeutic target for atherosclerosis.