MicroRNA-30d-5p promotes ovarian granulosa cell apoptosis by targeting Smad2.

Abstract

Polycystic ovarian syndrome (PCOS) is one of the leading causes of female infertility. MicroRNA-30d-5p (miR-30d-5p) has been reported to be significantly increased during follicle stimulating hormone (FSH)-mediated progesterone secretion of cultured granulosa cells. However, its role in the proliferation and apoptosis of ovarian granulosa cells is unclear. The present study aimed to investigate the role of miR-30d-5p in the proliferation and apoptosis of ovarian granulosa cells. Bioinformatic analysis and dual-luciferase reporter assay were used to predict and confirm the direct target of miR-30d-5p. The levels of miR-30d-5p were detected via reverse transcription-quantitative PCR (RT-qPCR), cell proliferation was detected via an MTT assay and cell apoptosis was measured via flow cytometry. The levels of phosphorylated (p)-Smad2, Smad2, p-Smad3 and Smad3 were detected by performing a western blot assay or RT-qPCR. In the present results, Smad2 was identified as the direct and functional target of miR-30d-5p. Compared with the control and control plasmid groups, the Smad2 plasmid significantly enhanced Smad2 mRNA levels in rat ovarian granulosa cells, enhanced rat ovarian granulosa cell viability and reduced cell apoptosis. In addition, the results demonstrated that overexpression of miR-30d-5p significantly decreased the level of Smad2, the effect of which was reversed by the Smad2-plasmid. Furthermore, it was demonstrated that the enhanced expression of miR-30d-5p significantly inhibited ovarian granulosa cell proliferation and promoted cell apoptosis. Restoration of Smad2 reversed the effect of miR-30d-5p on ovarian granulosa cell proliferation and apoptosis. Transfection with miR-30d-5p mimics significantly decreased the expression of Smad2 and increased the relative p-Smad2/Smad2 and p-Smad3/Smad3 levels in ovarian granulosa cells, which was reversed by overexpressing Smad2. The present study demonstrated that the overexpression of miR-30d-5p reduced proliferation and induced the apoptosis of granulosa cells by targeting Smad2. The molecular mechanism of ovarian granulosa cell apoptosis may therefore be explained by the newly identified miR-30d-5p/Smad2 axis, which represents a novel potential treatment target for PCOS.