title: MicroRNA-495 inhibits proliferation of glioblastoma multiforme cells by downregulating cyclin-dependent kinase 6.
abstract: BACKGROUND
  Glioblastoma multiforme (GBM) is the most aggressive type of glioma and carries the poorest chances of survival. There is therefore an urgent need to understand the mechanisms of glioma tumorigenesis and develop or improve therapeutics. The aim of this study was to assess the possible prognostic value of cyclin-dependent kinase 6 (CDK6) and the effects of microRNA-495 (miR-495) manipulation on CDK6 expression and cell survival in glioma cells.
  
  
  METHODS
  Analyses of clinical specimens from GBM patients were used. Expression of CDK6 was analyzed by real-time polymerase chain reaction (RT-PCR), Western blotting, and immunohistochemistry. Expression of CDK6 was also analyzed after over-expression of miR-495 in T98 cells; both cell proliferation and RB phosphorylation were examined. Cell proliferation, cell cycle distribution, and RB phosphorylation were also examined after knockdown of CDK6 in U87-MG and T98 cells.
  
  
  RESULTS
  Analyses of clinical specimens from GBM patients identified that CDK6 is significantly expressed in gliomas. CDK6 antigen expression was higher in tumor cores and margins than in adjacent normal brain tissues, and higher levels of CDK6 expression in the tumor margin correlated with decreased survival. Over-expression of miR-495 in T98 cells downregulated the expression of CDK6 and inhibited retinoblastoma phosphorylation, and knockdown of CDK6 in U87-MG and T98 cells by siRNAs resulted in cell cycle arrest at the G1/S transition and inhibition of cell proliferation.
  
  
  CONCLUSIONS
  This study revealed miR-495 is down-regulated in glioma tissues. Furthermore, miR-495 regulated CDK6 expression and involved in glioma cell growth inhibition, which indicated the possible role of miR-495 in tumor progression.
authors.affiliation: Graduate Institute of Clinical Medical Sciences, College of Medicine, Chang Gung University, No.259 Wen-Hwa 1st Rd, Kweishan 333, Taoyuan, Taiwan.
authors.forename: Shu-Mei
authors.initials: SM
authors.lastname: Chen
authors.forename: Hua-Chien
authors.initials: HC
authors.lastname: Chen
authors.forename: Shu-Jen
authors.initials: SJ
authors.lastname: Chen
authors.forename: Chiung-Yin
authors.initials: CY
authors.lastname: Huang
authors.forename: Pin-Yuan
authors.initials: PY
authors.lastname: Chen
authors.forename: Tai-Wei Erich
authors.initials: TW
authors.lastname: Wu
authors.forename: Ly-Ying
authors.initials: LY
authors.lastname: Feng
authors.forename: Hong-Chieh
authors.initials: HC
authors.lastname: Tsai
authors.forename: Tai-Ngar
authors.initials: TN
authors.lastname: Lui
authors.forename: Chuen
authors.initials: C
authors.lastname: Hsueh
authors.forename: Kuo-Chen
authors.initials: KC
authors.lastname: Wei
mesh_terms: D017209:Apoptosis; D015153:Blotting, Western; D001932:Brain Neoplasms / Q000235:genetics / Q000401:mortality / Q000473:pathology*; D059447:Cell Cycle Checkpoints; D049109:Cell Proliferation; D051361:Cyclin-Dependent Kinase 6 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D005260:Female; D005500:Follow-Up Studies; D015972:Gene Expression Regulation, Neoplastic; D005909:Glioblastoma / Q000235:genetics / Q000401:mortality / Q000473:pathology*; D006801:Humans; D007124:Immunoenzyme Techniques; D008297:Male; D035683:MicroRNAs / Q000235:genetics*; D008875:Middle Aged; D009367:Neoplasm Staging; D010766:Phosphorylation; D011379:Prognosis; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D016160:Retinoblastoma Protein / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015996:Survival Rate; D014407:Tumor Cells, Cultured
chemical_list: C577768:MIRN495 microRNA, human; D035683:MicroRNAs; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D016160:Retinoblastoma Protein; C495911:CDK6 protein, human; D051361:Cyclin-Dependent Kinase 6