cigarette smoke extract (cse) delays nod2 expression and affects nod2ripk2 interactions in intestinal epithelial cells香烟烟雾提取物(cse)延迟nod2表达和影响nod2ripk2在肠道上皮细胞的相互作用.pdfVIP

Cigarette Smoke Extract (CSE) Delays NOD2 Expression and Affects NOD2/RIPK2 Interactions in Intestinal Epithelial Cells 1 1 2 1 1 Marian C. Aldhous *, Kimberley Soo , Lesley A. Stark , Agata A. Ulanicka , Jennifer E. Easterbrook , Malcolm G. Dunlop2, Jack Satsangi1 1 Gastrointestinal Unit, Molecular Medicine Centre, Western General Hospital, University of Edinburgh, Edinburgh, Scotland, United Kingdom, 2 Edinburgh Cancer Centre and MRC Human Genetics Unit, Institute of Genetics and Molecular Medicine, Western General Hospital, Edinburgh, Scotland, United Kingdom Abstract Background: Genetic and environmental factors influence susceptibility to Crohn’s disease (CD): NOD2 is the strongest individual genetic determinant and smoking the best-characterised environmental factor. Carriage of NOD2 mutations predispose to small-intestinal, stricturing CD, a phenotype also associated with smoking. We hypothesised that cigarette smoke extract (CSE) altered NOD2 expression and function in intestinal epithelial cells. Methods and Findings: Intestinal epithelial cell-lines (SW480, HT29, HCT116) were stimulated with CSE and nicotine (to mimic smoking) 6TNFa (to mimic inflammation). NOD2 expression was measured by qRT-PCR and western blotting; NOD2- RIPK2 interactions by co-immunoprecipitation (CoIP); nuclear NFkB-p65 by ELISA; NFkB activity by luciferase reporter assays and chemokines (CCL20, IL8) in culture supernatants by ELISA. In SW480 and HT29 cells the TNFa-induced NOD2 expression at 4 hours was reduced by CSE (p = 0.0226), a response that was dose-dependent (p = 0.003) and time-dependent (p = 0.0004). Similar effects of CSE on NOD2 expression were seen in cultured ileal