METHODS: Reverse-transcription-polymerase chain reaction was employed to measure the expression of plasmacytoma variant translocation 1 (PVT1), microRNAs (miRNAs), and SIRT3, and the dual-luciferase assay was used to determine their interaction. Electron microscopy observes autophagosomes, green fluorescent protein-microtubule-associated protein 1 light chain 3 (GFP-LC3) staining, and immunoblot analysis with antibodies against LC3，beclin-1, and P62 were conducted to measure autophagy. Cellular senescence was determined using immunoblot analysis with anti-phosphorylated retinoblastoma and senescence-associated β-galactosidase staining.
RESULTS: Women with higher estrogen levels (during the 10-13th day of the menstrual cycle or premenopausal) exhibit markedly higher serum levels of PVT1 than women with lower estrogen levels (during the menstrual period or postmenopausal). The dual-luciferase assay showed that PVT1 acts as a sponge for miR-31, and miR-31 binds to its target gene, SIRT3. The 17β-E2 treatment increased the expression of PVT1 and SIRT3 and downregulated miR-31 expression in human umbilical vein endothelial cells (HUVECs). Consistently, PVT1 overexpression suppresses miR-31 expression, promotes 17β-E2-induced autophagy, and inhibits H2O2-induced senescence. miR-31 inhibitor increases SIRT3 expression and leads to activation of 17β-E2-induced autophagy and suppression of H2O2-induced senescence.
CONCLUSION: Our findings demonstrated that 17β-E2 upregulates PVT1 gene expression and PVT1 functions as a sponge to inhibit miR-31, resulting in the upregulation of SIRT3 expression and activation of autophagy and subsequent inhibition of H2O2-induced senescence in HUVECs.