title: MicroRNA-204-5p-Mediated Regulation of SIRT1 Contributes to the Delay of Epithelial Cell Cycle Traversal in Diabetic Corneas.
abstract: OBJECTIVE
  We investigated how the microRNA (miRNA) modifies the expression of silent mating type information regulation 2 homolog 1 (SIRT1) in diabetic corneas.
  
  
  METHODS
  The bioinformatic assay was used to predict which miRNAs might regulate the expression of SIRT1. A lipid transfection protocol was used to upregulate or knockdown the miRNA expression in TKE2 cells. Adenovirus-expressing short interfering RNA was used to knockdown the expression of SIRT1 in TKE2 cells and Ins2(Akita/+) mice were used to evaluate how miRNA promotes diabetic corneal epithelial wound healing. Cell cycle status was determined by flow cytometry assay and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to analyze the cell viability.
  
  
  RESULTS
  Nine miRNAs were selected for quantitative PCR (qPCR) detection after bioinformatics analysis. The miR-204-5p merited further investigation, because it was increased almost 5-fold in diabetic corneal epithelia compared to nondiabetic control corneal epithelia. Using luciferase activity assay, we identified SIRT1 was a direct target of miR-204-5p. The results of flow cytometry and MTT assay demonstrated that downregulation of miR-204-5p increased TKE2 cell growth and restored cell cycle progression in high glucose (HG) conditions by the regulation of Cyclin D1 and p16. Furthermore, we showed downregulation of miR-204-5p promoted HG attenuation of corneal epithelial wound healing via upregulation of SIRT1 in Ins2(Akita/+) mice.
  
  
  CONCLUSIONS
  Our data provide firm evidence of a role for miR-204-5p in the direct regulation of SIRT1 in diabetic corneas and identified the miR-204-5p-mediated regulation of SIRT1 contributes to the delay of epithelial cell cycle traversal in diabetic keratopathy. : Chinese Abstract.
authors.affiliation: Department of Ophthalmology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, Shandong Eye Institute, Shandong Academy of Medical Sciences, Qingdao, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Gao
authors.affiliation: State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, Shandong Eye Institute, Shandong Academy of Medical Sciences, Qingdao, China.
authors.forename: Ye
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, Shandong Eye Institute, Shandong Academy of Medical Sciences, Qingdao, China.
authors.forename: Xiaowen
authors.initials: X
authors.lastname: Zhao
authors.affiliation: State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, Shandong Eye Institute, Shandong Academy of Medical Sciences, Qingdao, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Chen
authors.affiliation: Department of Ophthalmology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, Shandong Eye Institute, Shandong Academy of Medical Sciences, Qingdao, China.
authors.forename: Lixin
authors.initials: L
authors.lastname: Xie
mesh_terms: D000818:Animals; D002453:Cell Cycle / Q000235:genetics*; D019295:Computational Biology; D003316:Corneal Diseases / Q000235:genetics* / Q000503:physiopathology*; D048909:Diabetes Complications / Q000235:genetics* / Q000503:physiopathology*; D003921:Diabetes Mellitus, Experimental / Q000235:genetics / Q000503:physiopathology; D004847:Epithelial Cells / Q000502:physiology*; D019573:Epithelium, Corneal / Q000503:physiopathology*; D005434:Flow Cytometry; D005786:Gene Expression Regulation / Q000235:genetics*; D008810:Mice, Inbred C57BL; D008815:Mice, Inbred Strains; D035683:MicroRNAs / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D056564:Sirtuin 1 / Q000235:genetics*; D014945:Wound Healing / Q000235:genetics
chemical_list: D035683:MicroRNAs; C470545:Sirt1 protein, mouse; D056564:Sirtuin 1
keywords: SIRT1; cell cycle; corneal wound healing; diabetic keratopathy; miRNA