Thyroid autoimmunity is associated with fertilization impairment and follicular fluid exosomal miRNA alterations in euthyroid women undergoing IVF-ET.

Objective: To investigate the impact of thyroid autoimmunity (TAI) on fertilization outcomes in euthyroid infertile women undergoing Methods: A total of 4,332 infertile women aged 20-40 years undergoing IVF-ET at the First People's Hospital of Yunnan Province between January and October 2024 were retrospectively enrolled. TAI was defined as TPOAb >34 IU/mL and/or TgAb >115 IU/mL. Fertilization rates were compared using the Wilcoxon rank-sum test, and the association between TAI and fertilization outcomes was assessed using generalized linear models. For mechanistic analysis, follicular fluid samples from 10 euthyroid TAI-positive and 10 euthyroid TAI-negative women were subjected to exosomal miRNA sequencing. TAI-related miRNAs were identified by integrating WGCNA and differential expression analysis (edgeR), followed by GO and KEGG enrichment analyses. Key miRNAs were screened using LASSO and SVM, and their diagnostic performance was evaluated by ROC analysis. Target genes were predicted using miRWalk and used to construct a miRNA-mRNA regulatory network with subsequent functional enrichment analysis. Results: Among the 4,332 infertile women undergoing IVF-ET, 565 were TAI-positive and 3,767 were TAI-negative. The fertilization rate was significantly lower in the TAI-positive group, and TAI was identified as an independent risk factor for impaired fertilization in euthyroid women. Exosomal miRNA sequencing of follicular fluid identified 70 differentially expressed miRNAs, and 17 candidate miRNAs were obtained by integrating WGCNA results. Further screening using LASSO and SVM identified four key miRNAs: hsa-miR-6799-3p, hsa-miR-6836-3p, hsa-miR-4449, and hsa-miR-3162-5p, all of which demonstrated good diagnostic performance (AUC > 0.7). Functional enrichment analysis of target genes suggested that these miRNAs may be involved in TAI-related reproductive impairment through pathways such as cell-matrix adhesion and mTOR signaling. Conclusion: TAI is independently associated with impaired fertilization outcomes in euthyroid infertile women undergoing IVF-ET and is accompanied by altered expression profiles of follicular fluid exosomal miRNAs. hsa-miR-6799-3p, hsa-miR-6836-3p, hsa-miR-4449, and hsa-miR-3162-5p may serve as potential molecular biomarkers for TAI-related reproductive dysfunction, providing insights into underlying mechanisms and potential diagnostic targets.