miR-146a/b-5p-IRAK1-NF-κB Axis Regulates Unexplained Recurrent Spontaneous Abortion

Unexplained recurrent spontaneous abortion (URSA) remains a prevalent obstetric issue, the underlying causes of which are still not fully understood. MicroRNAs (miRNAs) have the potential to influence URSA through their regulation of gene expression. This study aimed to screen key miRNAs and to verify their regulatory roles and potential molecular mechanisms in URSA. Differentially expressed miRNAs and mRNAs in URSA were identified using GEO datasets (GSE178619 and GSE113790). Weighted gene co-expression network analysis (WGCNA) was performed to identify hub miRNAs and mRNAs, followed by construction of a miRNA-mRNA interaction network. Functional enrichment was analyzed using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. An oxidized low-density lipoprotein (OX-LDL)-induced trophoblast cell model was established after concentration screening, and gain- and loss-of-function experiments involving miR-146a/b-5p and IRAK1 were conducted to evaluate cellular functions. The targeting relationship between miR-146a/b-5p and IRAK1 was examined using dual-luciferase reporter assays. In vivo, a URSA mouse model was constructed, and lentiviral overexpression of miR-146a/b-5p was applied to assess pregnancy outcomes and NF-κB pathway-related molecular changes. Results showed that GSE178619 identified 34 differentially expressed miRNAs, while GSE113790 revealed 1056 differentially expressed mRNAs. WGCNA highlighted the MEbrown module (112 hub miRNAs) and the MEdarkorange module (673 hub mRNAs) as closely associated with URSA. Eighteen core miRNAs, including miR-146a/b-5p, and 21 core genes, including IRAK1, were identified, with IRAK1 upregulated in URSA and linked to the NF-κB signaling pathway. In vitro, OX-LDL treatment impaired trophoblast cell proliferation and invasion, promoted apoptosis and oxidative stress, reduced miR-146a/b-5p expression, and activated NF-κB signaling through IRAK1. Overexpression of miR-146a/b-5p alleviated these alterations, whereas inhibition of miR-146a/b-5p produced opposite effects that were partially reversed by IRAK1 knockdown. In URSA mice, miR-146a/b-5p overexpression improved pregnancy outcomes, alleviated placental pathological changes, reduced IRAK1 expression, and attenuated NF-κB pathway activation. Together, miR-146a/b-5p regulates URSA progression by targeting IRAK1 and modulating the NF-κB signaling pathway, suggesting its potential as a therapeutic target for URSA.