title: Role of mast cell-miR-490-5p in irritable bowel syndrome.
abstract: OBJECTIVE
  To determine the functional role of miR-490-5p in mast cell proliferation and apoptosis, and in the mast cell tryptase/PAR-2 signal pathway.
  
  
  METHODS
  The 3rdgeneration of lentivirus vector systems containing enhanced green fluorescent protein (EGFP) (Ruisai Inc., Shanghai, China), which acts as a reporter gene was used to construct the mmu-miR-490-5p lentivirus expression vector pEGFP-antagomiR-490-5p, and the lentivirus vector pEGFP-negative was used as a negative control. The stably transfected mast cell line p815 was then constructed. GFP positive cells were successfully transfected cells. We determined the expression of miR-490-5p in p815 mast cells before and after transfection using quantitative real-time PCR (qRT-PCR). In addition, after transduction with the lentivirus vectors, the role of miR-490-5p in mast cell proliferation and apoptosis was investigated using the CCK-8 assay and flow cytometry, respectively. The mRNA levels of tryptase and PAR-2 were detected by qRT-PCR and the protein levels were detected by Western blot.
  
  
  RESULTS
  The inhibition of miR-490-5p expression promoted apoptosis and inhibited proliferation of p815 mast cells. The mRNA levels of tryptase and PAR-2 were significantly increased after transfection compared with the control group, tryptase (P= 0.721, normalvsnull;P= 0.001, siRNAvsnormal;P= 0.002, siRNAvsnull) and PAR-2 (P= 0.027, siRNAvsnull;P= 0.353, normalvsnull;P= 0.105, siRNAvsnormal). The protein levels of tryptase and PAR2 were slightly higher in the siRNA group than those in the control group, but the difference was not statistically significant (P> 0.05).
  
  
  CONCLUSIONS
  miR-490-5p plays a vital role in the pathogenesis of irritable bowel syndrome by affecting mast cell proliferation and apoptosis; with down-regulation of miR-490-5p, the mRNA level of mast cell tryptase and PAR-2 increased, and the protein level increased, but the difference was not statistically significant.
authors.affiliation: Hai-Xia Ren, He-Sheng Luo, Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei Province, China.
authors.forename: Hai-Xia
authors.initials: HX
authors.lastname: Ren
authors.affiliation: Hai-Xia Ren, He-Sheng Luo, Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei Province, China.
authors.forename: Fa-Can
authors.initials: FC
authors.lastname: Zhang
authors.affiliation: Hai-Xia Ren, He-Sheng Luo, Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei Province, China.
authors.forename: He-Sheng
authors.initials: HS
authors.lastname: Luo
authors.affiliation: Hai-Xia Ren, He-Sheng Luo, Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei Province, China.
authors.forename: Guo
authors.initials: G
authors.lastname: Zhang
authors.affiliation: Hai-Xia Ren, He-Sheng Luo, Department of Gastroenterology, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei Province, China.
authors.forename: Lie-Xin
authors.initials: LX
authors.lastname: Liang
mesh_terms: D000818:Animals; D000070416:Antagomirs / Q000235:genetics; D017209:Apoptosis / Q000235:genetics; D015415:Biomarkers / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000235:genetics; D015536:Down-Regulation; D005434:Flow Cytometry; D017930:Genes, Reporter / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000378:metabolism; D043183:Irritable Bowel Syndrome / Q000235:genetics / Q000378:metabolism*; D016086:Lentivirus / Q000235:genetics; D008407:Mast Cells / Q000201:enzymology / Q000378:metabolism*; D051379:Mice; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D044464:Receptor, PAR-2 / Q000378:metabolism*; D015398:Signal Transduction; D014162:Transfection; D053802:Tryptases / Q000378:metabolism*
chemical_list: D000070416:Antagomirs; D015415:Biomarkers; C000589011:MIRN490 microRNA, mouse; D035683:MicroRNAs; D012333:RNA, Messenger; D044464:Receptor, PAR-2; D049452:Green Fluorescent Proteins; D053802:Tryptases
keywords: Irritable bowel syndrome; Mast cell tryptase; PAR-2; miR-490-5p