title: Cigarette Smoke Modulates NOD1 Signal Pathway and Human β Defensins Expression in Human Oral Mucosa.
abstract: OBJECTIVE
  Nucleotide binding oligomerization domain 1 (NOD1) signal pathway and human β defensins (hBDs) play crucial roles in innate immune. Cigarette smoke has been confirmed to dampen innate immune in some human tissues, such as oral mucosa. The aim of this study was to evaluate potential effects of smoking on NOD1 signaling and hBDs expression in oral mucosa.
  
  
  METHODS
  Tissue specimens of normal oral mucosa were collected from donors undergoing routine surgical treatment. All 20 participants were classified equally as two groups: non-smokers and smokers. By using Western blotting and immunohistochemistry, we investigated differential expression of crucial molecules in NOD1 signal pathway, hBD-1, -2, and -3 in oral mucosa tissues between non-smokers and smokers. Immortalized human oral mucosal epithelial (Leuk-1) cells were treated with various concentrations of cigarette smoke extract (CSE) for 24h. Western blotting and immunofluorescence assays were performed to study CSE-induced alteration of protein expression. Leuk-1 cells were treated with 4% CSE, iE-DAP (NOD1 agonist), CSE + iE-DAP, BAY 11-7082 (NF-κB inhibitor), 4% CSE + BAY 11-7082, respectively. Real-time PCR and ELISA were performed to detect the mRNA levels and secretion of hBD-1, -2, and -3, respectively.
  
  
  RESULTS
  The levels of NOD1, NF-κB, hBD-1 and hBD-3 significantly reduced in oral mucosa tissues of smokers compared with non-smokers. The levels of RIP2 (receptor-interacting protein 2), phospho-NF-κB (P-NF-κB) and hBD-2 remarkably enhanced in oral mucosal tissues of smokers. CSE treatment suppressed NOD1 and NF-κB expression and activated RIP2 and P-NF-κB expression in Leuk-1 cells. The mRNA and secretory levels of hBD-1 and -3 were down-regulated by CSE, while the mRNA and secretory level of hBD-2 were up-regulated by CSE. The iE-DAP or BAY 11-7082 treatment reversed the regulatory effects of CSE on levels of hBDs.
  
  
  CONCLUSIONS
  The present study indicated that cigarette smoke could potentially modulate the expression of crucial molecules of NOD1 signal pathway and hBDs in human oral mucosal epithelium. NOD1 signal pathway could play an important role in the regulatory effects of CSE on hBDs levels in oral mucosal epithelial cells.
authors.affiliation: Department of Oral Medicine, Nanjing Stomatological Hospital, Medical School of Nanjing University, Nanjing, China.
authors.forename: Ya-Jie
authors.initials: YJ
authors.lastname: Qian
authors.forename: Xiang
authors.initials: X
authors.lastname: Wang
authors.forename: Ya-Fan
authors.initials: YF
authors.lastname: Gao
authors.forename: Ning
authors.initials: N
authors.lastname: Duan
authors.forename: Xiao-Feng
authors.initials: XF
authors.lastname: Huang
authors.forename: Fang-Fang
authors.initials: FF
authors.lastname: Sun
authors.forename: Xiao-Dong
authors.initials: XD
authors.lastname: Han
authors.forename: Wen-Mei
authors.initials: WM
authors.lastname: Wang
mesh_terms: D000328:Adult; D002460:Cell Line; D002470:Cell Survival; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D008297:Male; D008875:Middle Aged; D009061:Mouth Mucosa / Q000276:immunology* / Q000378:metabolism / Q000473:pathology; D016328:NF-kappa B / Q000032:analysis / Q000235:genetics / Q000276:immunology; D053474:Nod1 Signaling Adaptor Protein / Q000032:analysis / Q000235:genetics / Q000276:immunology*; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2 / Q000032:analysis / Q000235:genetics / Q000276:immunology; D015398:Signal Transduction; D012907:Smoking / Q000235:genetics / Q000276:immunology* / Q000473:pathology; D055815:Young Adult; D023083:beta-Defensins / Q000032:analysis / Q000235:genetics / Q000276:immunology*
chemical_list: D016328:NF-kappa B; C119067:NOD1 protein, human; D053474:Nod1 Signaling Adaptor Protein; D023083:beta-Defensins; C506253:RIPK2 protein, human; D053475:Receptor-Interacting Protein Serine-Threonine Kinase 2