Introduction: Gestational diabetes mellitus (GDM) is the glucose intolerance that occurs during pregnancy and associated with short and long-term consequences to mother and offspring. In this study, we identify the role of miRNAs in small extracellular vesicles (sEVs) from placenta and their role in regulation of insulin sensitivity in GDM.
Method: sEVs were isolated from conditioned media of primary trophoblast cultures from normal glucose tolerant (NGT) and GDM patients and miRNA profile in sEVs analysed using next generation sequencing. The miRNAs were subjected to motif analysis using MDS2 algorithm. RBPs specifically interacting with the miRNAs were identified by biotin pull down assay and mass spectrometry. RBPs were knockdown in placental cells and cocultured with skeletal muscle cells and insulin stimulated glucose uptake assay was performed.
Results: 262 miRNAs were highly enriched in sEVs compared to placental cells in NGT and 187 miRNAs were highly enriched in sEVs from placental cells in GDM. An unbiased search for sequences identified overexpressed miRNA motifs and based on their coverage, statistical significance and uniqueness candidate motifs in NGT and GDM were chosen. Four candidate miRNAs were chosen (miR-1246, miR-1285-5p, miR-150-5p, miR-486-5p) and identified that proteins ELF4B, FASTKD2, GRSF1, HNRNPH2, PTBP3, HDLBP, YBX3 and DIS3 were specifically interacting with the candidate miRNAs. Further, knockdown of DIS3 protein in donor placental cells leads to changes in insulin stimulated glucose uptake in recipient skeletal muscle cells.
Conclusion: RBPs are involved in the packaging of miRNAs in sEVs which when delivered in target cells might regulate insulin sensitivity.