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MiR?132 is upregulated in polycystic ovarian syndrome and inhibits granulosa cells viability by targeting Foxa1.

ABSTRACT: Polycystic ovary syndrome (PCOS) is one of the most common endocrine metabolic disorders characterized by hyperandrogenism, polycystic ovaries and ovulatory dysfunction. Several studies have suggested that the aberrant expression of microRNAs (miRNAs/miRs) plays an important role in the pathogenesis of PCOS; however, the role and underlying mechanisms of miR?132 in the development of PCOS remain unclear. In the present study, the expression of miR?132 in granulosa cells (GCs) derived from 26 patients with PCOS and 30 healthy controls was detected by reverse transcription?quantitative PCR (RT?qPCR). The apoptosis of GCs was examined using a TUNEL assay. The human ovarian granulosa?like tumor cell line, KGN, was cultured for Cell Counting Kit?8 assays following the overexpression or knockdown of miR?132. TargetScan was applied to identify the potential targets of miR?132, which was further verified by a luciferase assay, RT?qPCR and western blotting. The expression of miR?132 was decreased in GCs from patients with PCOS. Moreover, the GCs of patients with PCOS exhibited significantly increased apoptotic nuclei. Furthermore, the overexpression of miR?132 inhibited the viability of KGN cells. In addition, the results verified that miR?132 directly targeted forkhead box protein A1 (Foxa1), the knockdown of which suppressed KGN cell viability. On the whole, the findings of the present study demonstrated that miR?132 inhibited cell viability and induced apoptosis by directly interacting with Foxa1. Thus, miR?132 may be a potential target for the treatment of patients with PCOS.

SUBMITTER: Cui X

PROVIDER: S-EPMC7646966 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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miR‑132 is upregulated in polycystic ovarian syndrome and inhibits granulosa cells viability by targeting Foxa1.

Cui Xiangrong X Jing Xuan X Liu Junfen J Bi Xingyu X Wu Xueqing X

Molecular medicine reports 20201014 6

Polycystic ovary syndrome (PCOS) is one of the most common endocrine metabolic disorders characterized by hyperandrogenism, polycystic ovaries and ovulatory dysfunction. Several studies have suggested that the aberrant expression of microRNAs (miRNAs/miRs) plays an important role in the pathogenesis of PCOS; however, the role and underlying mechanisms of miR‑132 in the development of PCOS remain unclear. In the present study, the expression of miR‑132 in granulosa cells (GCs) derived from 26 pat ...[more]

PMID: 33174054

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Project description:Vitamin D (VD) is not only associated with bone growth and development, but is also closely associated with numerous other pathological conditions. The present study aimed to investigate the effect of microRNA (miRNA/miR)‑378d on ovarian granulosa cells by regulating the round spermatid basic protein 1 (Rsbn1) in the absence of VD. The abnormal expression of miRNAs in ovarian tissues of the VD‑deficient mouse was analyzed using transcriptome sequencing. miR‑378d, glucose transporter 4 (Glut4) and aromatase (Cyp19a) expression levels were examined via reverse transcription‑quantitative (RT‑q)PCR and western blotting. The expression levels of Rsbn1, Glut4 and Cyp19a were detected in transfected mouse ovarian granulosa cells. The targeting regulation between miR‑378d and Rsbn1 was verified using double reporter gene assay and functional rescue experiments. Among the 672 miRNAs that were differentially expressed, cluster analysis revealed that 17 were significantly upregulated and 16 were significantly downregulated. Moreover, miR‑378d showed significant upregulation, which was further verified via RT‑qPCR. It was identified that the protein expression level of Rsbn1 was significantly downregulated. Furthermore, Glut4 mRNA expression was significantly decreased in the mimic group but markedly increased in the inhibitor group. By contrast, the mRNA expression levels of Rsbn1 and Cyp19a did not demonstrate any significant difference. The western blotting results indicated that the protein expression levels of Rsbn1 and Glut4 were decreased and increased, respectively, while Cyp19a did not show any significant change. In addition, the double reporter gene experiments confirmed that Rsbn1 was the target gene of miR‑378d. Collectively, the present results demonstrated that miR‑378d was abnormally overexpressed in the ovarian tissues of the VD‑deficient mice, and that miR‑378d could inhibit Glut4 production by targeting Rsbn1, which may lead to insulin resistance.

Project description:BackgroundEstrogen synthesis is an important function of the mammalian ovary. Estrogen plays important roles in many biological processes, including follicular development, oocyte maturation and endometrial proliferation, and dysfunctions in estrogen synthesis contribute to the development of polycystic ovary syndrome and premature ovarian failure. Classical signaling cascades triggered by follicle-stimulating hormone induce estrogen synthesis via the upregulation of Cyp19a1 in granulosa cells (GCs). This study aimed to determine the effect of microRNA-132 (miR-132) on estradiol synthesis in GCs.MethodsPrimary mouse GCs were collected from ovaries of 21-day-old immature ICR mice through follicle puncture. GCs were cultured and treated with the stable cyclic adenosine monophosphate analog 8-Br-cAMP or transfected with miR-132 mimics, Nurr1-specific small interfering RNA oligonucleotides and Flag-Nurr1 plasmids. Concentrations of estradiol and progesterone in culture medium were determined by an automated chemiluminescence-based assay. Quantitative real time PCR and western blot were performed to identify the effect of miR-132 on Cyp19a1, Cyp11a1 and an orphan nuclear receptor-Nurr1 expression in GCs. Direct suppression of Nurr1 via its 3'-untranslated region by miR-132 were further verified using luciferase reporter assays.ResultsThe expression level of miR-132 in cultured mouse GCs was significantly elevated during 48 h of treatment with 8-Br-cAMP. The synthesis of estradiol increased after the overexpression of miR-132 in mouse GCs. The real-time PCR results demonstrated that miR-132 induced the expression of Cyp19a1 significantly. Nurr1, an orphan nuclear receptor that suppresses Cyp19a1 expression, was found to be a direct target of miR-132. Nurr1 was suppressed by miR-132, as indicated by a luciferase assay and Western blotting. The knockdown of Nurr1 primarily elevated the synthesis of estradiol and partially attenuated the miR-132-induced estradiol elevation, and the ectopic expression of Flag-Nurr1 abrogated the stimulatory effect of miR-132 on estradiol synthesis in mouse GCs.ConclusionsOur findings suggest that miR-132 is involved in the cAMP signaling pathway and promotes estradiol synthesis via the translational repression of Nurr1 in ovarian GCs.

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MiR?132 is upregulated in polycystic ovarian syndrome and inhibits granulosa cells viability by targeting Foxa1.

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miR‑132 is upregulated in polycystic ovarian syndrome and inhibits granulosa cells viability by targeting Foxa1.

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