From Constitution to Disease: MicroRNA Signatures for the Early Prediction and Targeted Prevention of Polycystic...

The association between PCOS and the TCM concept of phlegm-damp constitution is well-documented, but their connection via miRNA remains unclear. This study sought to identify key miRNAs implicated in the transition from phlegm-damp constitution to PCOS and to elucidate their associated regulatory networks.

We conducted miRNA sequencing on granulosa cells from three groups: healthy controls with a neutral constitution (NP), healthy individuals with a phlegm-damp constitution (NB), and PCOS patients (PC). Principal component analysis (PCA) and differential expression analysis were utilized to pinpoint candidate miRNAs, which were subsequently validated using quantitative real-time PCR (qRT-PCR) in larger cohorts. Bioinformatics, including target prediction, protein-protein interaction (PPI) network construction, and pathway analysis, were employed to delineate the regulatory networks involved.

PCA revealed distinct clustering patterns, with NB and PC groups exhibiting closer proximity compared to the NP group. Differential expression analysis identified 24 dysregulated miRNAs in the PC group (20 upregulated, 4 downregulated) and 14 in the NB group (11 upregulated, 3 downregulated), with both groups showing shared enrichment in metabolic, PI3K-Akt, and Ras pathways. Validation confirmed a progressive downregulation of hsa-miR-24-2-5p and hsa-miR-33a-5p across the NP, the NB, and PC groups (P<0.05), whereas hsa-miR-374b-5p exhibited differential expression exclusively between NB and PC groups. Network analysis identified five hub genes (EGFR, MYC, KRAS, CREB1, SMAD4) and nine risk pathways, including MAPK and Wnt signaling pathways. The constructed miRNA-hub gene-pathway network comprised five miRNA-hub gene pairs and thirteen functional modules.

These findings elucidate shared miRNA regulatory mechanisms between phlegm-damp constitution and PCOS, highlighting hsa-miR-24-2-5p and hsa-miR-33a-5p as pivotal mediators in this pathological transition. This provides molecular evidence for the TCM "constitution-disease correlation" theory and identifies potential targets for the prevention and treatment of PCOS.