Crohn's disease is a chronic inflammatory disorder of the gastrointestinal tract, which is thought to result from the effect of environmental factors in a genetically predisposed host. A gene location in the pericentromeric region of chromosome 16, IBD1, that contributes to susceptibility to Crohn's disease has been established through multiple linkage studies, but the specific gene(s) has not been identified. NOD2, a gene that encodes a protein with homology to plant disease resistance gene products is located in the peak region of linkage on chromosome 16 (ref. 7). Here we show, by using the transmission disequilibium test and case-control analysis, that a frameshift mutation caused by a cytosine insertion, 3020insC, which is expected to encode a truncated NOD2 protein, is associated with Crohn's disease. Wild-type NOD2 activates nuclear factor NF-kappaB, making it responsive to bacterial lipopolysaccharides; however, this induction was deficient in mutant NOD2. These results implicate NOD2 in susceptibility to Crohn's disease, and suggest a link between an innate immune response to bacterial components and development of disease.

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Crohn's disease (CD) is a chronic inflammation affecting the gastrointestinal tract. Three mutations (Arg702Trp, Gly908Arg and Leu1007fsinsC) within the NOD2/CARD15 gene increase CD susceptibility. Here, we define cytokine regulation in primary human mononuclear cells, with muramyl dipeptide (MDP), the minimal NOD2/CARD15 activating component of peptidoglycan. By microarray, MDP induces a broad array of transcripts, including interleukin 1beta (IL-1beta) and interleukin 8 (IL-8). Leu1007fsinsC homozygotes demonstrated decreased transcriptional response to MDP. Electromobility shift assay demonstrated that MDP-induced NF-kappaB activation is mediated via p50 and p65 subunits, but not RelB or c-Rel. In wild-type individuals, MDP-induced IL-8 protein expression with a greater response to high dose (1 micro g/ml) compared with low-dose (10 ng/ml) MDP. At low MDP doses, in all homozygotes, we observed no induction of IL-8 protein. With high doses of MDP, Leu1007fsinsC homozygotes showed no induction. Modest induction of IL-8 protein was observed in Gly908Arg and Arg702Trp homozygotes, indicating varying MDP sensitivity of the CD-associated mutations. In wild-type healthy control, CD and ulcerative colitis individuals, low-dose MDP and TNFalpha alone results in only modest IL-1beta protein induction. With MDP plus TNFalpha, there is a synergistic induction of IL-1beta secretion. In Leu1007fsinsC homozygotes, there is a profound defect in IL-1beta secretion, despite marked induction of IL-1beta mRNA. These findings demonstrate post-transcriptional dependency on the NOD2/CARD15 pathway for IL-1beta secretion with MDP and TNFalpha treatment. Taken together, these studies suggest that a signaling defect of innate immunity to MDP may be an essential underlying defect in the pathogenesis of some CD patients.

Carrying two NOD2 mutations predicts youthful onset, ileal disease involvement, and development of stricturing or non-perianal fistulizing complications. Smoking and early onset independently influence ileal site and time to surgery.

Crohn disease (CD) and ulcerative colitis (UC) are overlapping chronic inflammatory bowel diseases (IBDs). Suggestive evidence for linkage at chromosome 7q has been reported for both CD and UC. Contained within this region is the gene for MDR1 (multidrug resistance), a membrane transport protein for which human polymorphisms have been reported in Ala893Ser/Thr and C3435T that alter pharmacokinetic profiles for a variety of drugs. Because mdr1 knockout mice spontaneously develop colitis, exonic regions were resequenced and tested for IBD association in a large, multicenter North American cohort. Two missense mutations, Asn21Asp and Ala893Ser/Thr, as well as the expression-associated polymorphism C3435T, described elsewhere, were genotyped in the entire cohort. Significant association of Ala893 with IBD was observed by both case-control analysis (P=.002) and the pedigree disequilibrium test (PDT [P=.00020-.00030]) but not for the Asn21Asp or C3435T polymorphisms. Significant association by PDT was observed within the subset with CD (P=.0014-.00090), with similar, nonsignificant trends in a smaller subset with UC. The Ala893Ser/Thr variant is triallelic, and the associated, common allele is Ala893, with undertransmission of the 893Ser (common) and the 893Thr (rare) variants. The Ala893 variant has decreased activity compared with the 893Ser variant; therefore, the association with human IBD is consistent with the murine model of mdr1 deficiency. Taken together, these data support the association of the common Ala893 polymorphism with IBD specifically and, more broadly, provides additional support for its contribution to interindividual pharmacogenetic variation.