Role of the CARD15 gene in the pathogenesis of Crohn disease: phenotypic classification and prognostic implications.

Crohn disease pathogenesis is the result of the complex interaction between environmental factors (i.e. micro-flora in the gut), immunologic and genetic predisposition (1, 2). The bacterial pathogenesis hypothesis in Crohn disease is based on experimental data and clinical observation. The first studies in rodent models, such as those in transgenic HLA-B27 rats and many others in mice, demonstrated differences in disease manifestations that lack disease in germ-free environments compared to experimental animals with normal enteric commensal microorganisms (2–5). Evidence for enhanced mucosal permeability, or absorption of potential inflammatory bacterial polymers from the small-bowel system based on overgrowth of luminal bacteria, has been documented (6). These pathogenic or normal luminal bacteria constantly stimulate the mucosal and systemic immune systems and perpetuate the inflammatory cascade (7, 8). Pioneer studies in Oxford by the late Sidney Truelove in the 1960s demonstrated the beneficial effect of diverting the luminal flow in severe colonic disease. Interestingly, the first studies showed better effect in Crohn disease than in ulcerative colitis. In 1983, Harper and co-workers reported an immediate but temporary clinical improvement in colonic Crohn disease patients who had a split or doublebarrelled ileostomy (9). When investigating the influence of the faecal stream in the pathogenesis of Crohn lesions in patients with an ileal resection, Rutgeerts and co-workers described important differences between Crohn disease patients with and those without a temporally diverting terminal ileostomy. Patients who underwent reanastomosis after 6 months did not present characteristic inflammatory lesions of Crohn disease in the isolated neoterminal ileum. However, recurrence of disease was observed 6 months after reanastomosis in these patients as well as in those with onestep surgery (10). The role of bacteria in the pathogenesis of Crohn disease has been the subject of speculation and debate. The fact that abnormal homeostatic mucosal immune responses to normal intestinal flora could predispose individuals to uncontrolled or pathological intestinal inflammation has further enhanced the search for pathogenic microorganisms in Crohn disease (11). One or more specific pathogens have been associated with Crohn disease, but it is clear that the disease is caused by more than just microbes. Familial aggregation and twin studies have suggested the role of genetics in determining the susceptibility to Crohn disease. In addition, the incomplete concordance for Crohn disease within monozygotic twins, the phenotypic variations and the observed familial pattern of non-Mendelian inheritance suggest the presence and importance of environmental factors in the pathogenesis of this disease or syndrome. Age appears to be a modifying factor and the genetic anticipation of the disease developing at an early age in families with multiple members affected by the disease has been discussed elsewhere (12–14). In 1996, Hugot (15) reported an association between the IBD1 locus on the pericentromeric region of chromosome 16 conferring susceptibility to Crohn disease in French families with multiple members affected. Ogura et al. in the United States and Hugot et al. in France (16, 17) identified the relevant gene on chromosome 16q12: CARD15. Using linkage disequilibrium mapping analysis on 1272 family members from 235 Crohn disease families, Hugot et al. identified numerous SNPs by sequencing. Subsequently, an association was demonstrated for Crohn disease with three of these identified SNPs—a frameshift mutation and two independent missense mutations—in the CARD15 gene (16). Ogura et al. independently identified the frameshift mutation in association with Crohn disease. By searching a genomic database for NOD1 homologues, the cDNA encoding the NOD2 protein coded by the CARD15 gene was identified by the G. Nuñez group in Michigan, USA (18). Sequence analyses predicted the protein and a variant. Transmission disequilibrium tests and case-control analysis on IBD families were used to associate this shorter protein variant with Crohn disease (17). These studies demonstrated that the CARD15 gene is the first locus causing susceptibility to Crohn disease in a Western population (15–21). NOD2, a member of the CED4/APAF1 superfamily of apoptosis regulators, is a cytoplasmatic bacterial patternrecognition receptor expressed primarily in monocytes (18, 22). This protein contains 1040 amino acids and is composed of three motifs: two amino-terminal caspase recruitment domains (CARD), a centrally located nucleotide binding domain (NBD) and 10 carboxy-terminal leucine-rich repeats (LRR) (see Fig. 1) (18).