title: Mmu-miR-125b overexpression suppresses NO production in activated macrophages by targeting eEF2K and CCNA2.
abstract: BACKGROUND
  MicroRNAs have been shown to be important regulators of the immune response and the development of the immune system. It was reported that microRNA-125b (miR-125b) was down-regulated in macrophages challenged with endotoxin. However, little is known about the function and mechanism of action of miR-125b in macrophage activation. Macrophages use L-arginine to synthesize nitric oxide (NO) through inducible NO synthase (iNOS), and the released NO contributes to the tumoricidal activity of macrophages.
  
  
  METHODS
  Luciferase reporter assays were employed to validate regulation of a putative target of miR-125b. The effect of miR-125b on endogenous levels of this target were subsequently confirmed via Western blot. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was performed to determine the expression level of miR-125b in macrophage. MTS assays were conducted to explore the impact of miR-125b overexpression on the cell viability of 4T1 cells.
  
  
  RESULTS
  Here, we demonstrate that mmu-miR-125b overexpression suppresses NO production in activated macrophages and that LPS-activated macrophages with overexpressed mmu-miR-125b promote 4T1 tumor cell proliferation in vitro and 4T1 tumor growth in vivo. CCNA2 and eEF2K are the direct and functional targets of mmu-miR-125b in macrophages; CCNA2 and eEF2K expression was knocked down, which mimicked the mmu-miR-125b overexpression phenotype.
  
  
  CONCLUSIONS
  These data suggest that mmu-miR-125b decreases NO production in activated macrophages at least partially by suppressing eEF2K and CCNA2 expression.
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100005, China.
authors.forename: Zhenbiao
authors.initials: Z
authors.lastname: Xu
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100005, China.
authors.forename: Lianmei
authors.initials: L
authors.lastname: Zhao
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100005, China.
authors.forename: Xin
authors.initials: X
authors.lastname: Yang
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100005, China.
authors.forename: Sisi
authors.initials: S
authors.lastname: Ma
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100005, China.
authors.forename: Yehua
authors.initials: Y
authors.lastname: Ge
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100005, China.
authors.forename: Yanxin
authors.initials: Y
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100005, China.
authors.forename: Shilian
authors.initials: S
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100005, China. shijuantt@163.com.
authors.forename: Juan
authors.initials: J
authors.lastname: Shi
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100005, China. zhengdx@pumc.edu.cn.
authors.forename: Dexian
authors.initials: D
authors.lastname: Zheng
mesh_terms: D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000235:genetics; D056751:Cyclin A2 / Q000096:biosynthesis / Q000235:genetics*; D054736:Elongation Factor 2 Kinase / Q000096:biosynthesis / Q000235:genetics*; D004731:Endotoxins / Q000633:toxicity; D005786:Gene Expression Regulation; D006801:Humans; D008262:Macrophage Activation / Q000235:genetics / Q000276:immunology; D008264:Macrophages / Q000187:drug effects / Q000276:immunology / Q000473:pathology; D035683:MicroRNAs / Q000096:biosynthesis* / Q000276:immunology; D009569:Nitric Oxide / Q000096:biosynthesis* / Q000276:immunology; D052247:Nitric Oxide Synthase Type II / Q000235:genetics
chemical_list: C508184:CCNA2 protein, mouse; D056751:Cyclin A2; D004731:Endotoxins; D035683:MicroRNAs; C527468:Mirn125 microRNA, mouse; D009569:Nitric Oxide; C496319:NOS2 protein, human; D052247:Nitric Oxide Synthase Type II; C517690:Eef2k protein, mouse; D054736:Elongation Factor 2 Kinase
keywords: CCNA2; Macrophages; Mmu-miR-125; Nitric oxide; eEF2K