title: Mindin is upregulated during colitis and may activate NF-kappaB in a TLR-9 mediated manner.
abstract: OBJECTIVE
  To investigate the regulation of mindin expression and the signaling pathway involved during inflammation.
  
  
  METHODS
  C57BL/6 mice were treated with 3% dextran sulfate sodium (DSS) in drinking water for 6 d to induce acute colitis, and then the colon was harvested for histological analysis or for RNA isolation. mRNA expression of mindin and nuclear factor (NF)-kappaB p65 was analyzed by quantitative real time polymerase chain reaction (RT-PCR) and mindin expression construct was confirmed by Western blotting. Mouse macrophage and intestinal epithelial lineage cells were stimulated with different cytokines and toll-like receptor (TLR) ligands, before pNF-kappaB-luciferase activity was assessed using the Dual-Luciferase reporter assay system.
  
  
  RESULTS
  mRNA expression of mindin was upregulated 4.7 + or - 1.1 fold compared with the baseline during DSS-induced intestinal inflammation in the mice. Stimulation with CpG-ODN (a known TLR-9 ligand) induced 4.2 + or - 0.3 fold upregulation of mindin expression in RAW 264.7 cells. Full-length of mindin was cloned from cDNA of mouse mesenteric lymph node, then the pCMV-Mindin-Flag expression vector was established and the protein expression level was confirmed. Transfection of the mindin construct and stimulation with CpG-ODN significantly increased the NF-kappaB-luciferase activity by 2.5 + or - 0.3 and 4.5 + or - 0.5 fold in RAW264.7 and CMT93 cells, respectively (P < 0.01).
  
  
  CONCLUSIONS
  Mindin expression is upregulated during intestinal inflammation and may induce NF-kappaB promoter activation in a TLR-9 mediated manner.
authors.affiliation: Division of Gastroenterology, Zhongshan Hospital Affiliated to Xiamen University, Xiamen 361004, Fujian Province, China.
authors.forename: Bayasi
authors.initials: B
authors.lastname: Guleng
authors.forename: Ya-Mei
authors.initials: YM
authors.lastname: Lian
authors.forename: Jian-Lin
authors.initials: JL
authors.lastname: Ren
mesh_terms: D000208:Acute Disease; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D003001:Cloning, Molecular; D003092:Colitis / Q000139:chemically induced / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D003106:Colon / Q000276:immunology / Q000378:metabolism*; D016264:Dextran Sulfate; D004195:Disease Models, Animal; D016326:Extracellular Matrix Proteins / Q000235:genetics / Q000378:metabolism*; D017930:Genes, Reporter; D008024:Ligands; D008264:Macrophages / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D009838:Oligodeoxyribonucleotides / Q000276:immunology; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D051217:Toll-Like Receptor 9 / Q000378:metabolism*; D051996:Transcription Factor RelA / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D015854:Up-Regulation
chemical_list: C408982:CPG-oligonucleotide; D016326:Extracellular Matrix Proteins; D008024:Ligands; D009838:Oligodeoxyribonucleotides; D012333:RNA, Messenger; C498144:Rela protein, mouse; C495370:Tlr9 protein, mouse; D051217:Toll-Like Receptor 9; D051996:Transcription Factor RelA; C121491:mindin; D016264:Dextran Sulfate