miR-135a-5p alleviates cerebral ischemia-reperfusion injury by inhibiting pyroptosis mediated through the DDX3X/NLRP3 pathway

Abstract

MicroRNAs (miRNAs) are widely involved in signal transduction and regulation during cerebral ischemia-reperfusion injury (CIRI). This study investigates the molecular mechanisms of the specific miRNA/DDX3X/NLRP3 pathway in early-stage CIRI and explores its potential clinical applications. Through public database analysis, miR-135a-5p targeting DDX3X after CIRI was determined. The levels of DDX3X, NLRP3 inflammasome, and GSDMD-N were increased after MCAO/R. Upregulation of miR-135a-5p suppressed these protein levels. Upregulating miR-135a-5p also reduced infarct volume and neuronal pyroptosis, while improved neurological scores in MCAO/R mice. Co-IP confirmed protein interaction between DDX3X and NLRP3 in CIRI models. Furthermore, miR-135a-5p mimics alleviated pyroptosis and inhibited DDX3X/NLRP3 pathway activation after OGD/R cells, whereas miR-135a-5p inhibitor produced the opposite effect. The dual-luciferase reporter assay validated that DDX3X was a direct target of miR-135a-5p. Clinically, the serum level of miR-135a-5p was significantly lower in CIRI patients after thrombectomy compared to controls. The levels of DDX3X, NLRP3, and IL-18 were elevated in the CIRI group, while the difference of IL-1β levels between the two groups was not statistically significant (p = 0.055). Although an inverse correlation was observed between miR-135a-5p and DDX3X levels in CIRI patients, the linear regression analysis did not reach statistical significance (R² = 0.12, p = 0.061). This study indicated that miR-135a-5p/DDX3X/NLRP3 pathway is pivotal in early-stage CIRI. Upregulation of miR-135a-5p inhibits NLRP3-mediated neuronal pyroptosis by targeting DDX3X, thereby alleviating CIRI and improving neurological function. This signaling axis holds promise for future clinical applications in treating CIRI.