title: Characterization of microRNA profile in human cumulus granulosa cells: Identification of microRNAs that regulate Notch signaling and are associated with PCOS.
abstract: BACKGROUND
  Polycystic ovary syndrome (PCOS), a complex and heterogeneous endocrine condition, is characterized by polycystic ovaries, hyperandrogenism, insulin resistance and chronic anovulation. Cumulus granulosa cells surrounding the oocyte are involved in different aspects of PCOS pathology. Several studies suggested that miRNAs play an important regulatory role at the post-transcriptional level in cumulus granulosa cells.
  
  
  OBJECTIVE
  Our objective was to describe the altered miRNA expression profiles and miRNA targeted signaling pathways in PCOS.
  
  
  METHODS
  Case-control study that involved 21 women with PCOS and 20 women without the disease (controls). The miRNA expression profiles of human cumulus granulosa cells were determined using next generation sequencing by Illumina Hiseq 2000. The differentially expressed miRNAs and novel miRNAs were validated by quantitative real-time PCR. The Notch3 and MAPK3 were demonstrated to be targeted by miR-483-5p based on quantitative real-time PCR, western blot and luciferase activity assay.
  
  
  RESULTS
  Compared with controls, a total of 59 known miRNA were identified that differentially expressed in PCOS cumulus granulosa cells, including 21 miRNAs increase and 38 miRNAs decrease. Moreover, the novel miRNAs were predicted in PCOS and control cumulus granulosa cells. The potential regulating roles of miRNA in pathophysiology of PCOS were analyzed by GO and KEGG pathway annotation, and several important processes were identified to be targeted by the differentially expressed miRNAs, such as Notch signaling, regulation of hormone, and energy metabolism. Furthermore, Notch3 and MAPK3, the members of Notch signaling and ERK-MAPK pathway, were demonstrated to be regulated by miR-483-5p based on negative expression correlation validation and detection of Notch3/MAPK3 expression after miR-483-5p mimics transfection. Dual luciferase activity assay suggested that Notch3 and MAPK3 were directly targeted by miR-483-5p.
  
  
  CONCLUSIONS
  Our data suggested that miRNAs and their targeted pathways (e.g. Notch signaling pathway) play important roles in the etiology and pathophysiology of PCOS, and provides novel candidates for molecular biomarkers or treatment targets in the research of female infertility associated with PCOS.
authors.affiliation: Center for Reproductive Medicine, Anhui Provincial Hospital Affiliated to Anhui Medical University, Hefei 230001, China. Electronic address: bio_xubo@163.com.
authors.forename: Bo
authors.initials: B
authors.lastname: Xu
authors.affiliation: Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, Hefei 230027, China.
authors.forename: Yuan-Wei
authors.initials: YW
authors.lastname: Zhang
authors.affiliation: Center for Reproductive Medicine, Anhui Provincial Hospital Affiliated to Anhui Medical University, Hefei 230001, China.
authors.forename: Xian-Hong
authors.initials: XH
authors.lastname: Tong
authors.affiliation: Center for Reproductive Medicine, Anhui Provincial Hospital Affiliated to Anhui Medical University, Hefei 230001, China.
authors.forename: Yu-Sheng
authors.initials: YS
authors.lastname: Liu
mesh_terms: D016022:Case-Control Studies; D002478:Cells, Cultured; D054885:Cumulus Cells / Q000166:cytology* / Q000473:pathology; D005260:Female; D020869:Gene Expression Profiling; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics*; D011085:Polycystic Ovary Syndrome / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D000071656:Receptor, Notch3; D051880:Receptors, Notch / Q000235:genetics* / Q000378:metabolism; D017423:Sequence Analysis, RNA; D015398:Signal Transduction
chemical_list: C548912:MIRN483 microRNA, human; D035683:MicroRNAs; C499374:NOTCH3 protein, human; D000071656:Receptor, Notch3; D051880:Receptors, Notch; D048052:Mitogen-Activated Protein Kinase 3
keywords: Cumulus granulosa cells; Next-generation sequencing; Notch signaling; Polycystic ovary syndrome; microRNA profile