Clinical significance of NOD2/CARD15 and Toll-like receptor 4 gene single nucleotide polymorphisms in inflammatory bowel disease.

AIM: To evaluate the role of genetic factors in the pathogenesis of Crohn's disease (CD) and ulcerative colitis (UC), we investigated the single nucleotide polymorphisms (SNPs) of NOD2/CARD15 (R702W, G908R and L1007finsC), and Toll-like receptor 4 (TLR4) genes (D299G and T399I) in a selected inflammatory bowel disease (IBD) population coming from Southern Italy. METHODS: Allele and genotype frequencies of NOD2/CARD15 (R702W, G908R and L1007finsC) and TLR4 (D299G and T399I) SNPs were examined in 133 CD patients, in 45 UC patients, and in 103 healthy controls. A genotype-phenotype correlation was performed. RESULTS: NOD2/CARD15 R702W mutation was significantly more frequent in CD (9.8%) than in controls (2.4%, P = 0.001) and in UC (2.3%, P = 0.03). No significant difference was found between UC patients and control group (P > 0.05). In CD and UC patients, no significant association with G908R variant was found. L1007finsC SNP showed an association with CD (9.8%) compared with controls (2.9%, P = 0.002) and UC patients (2.3%, P = 0.01). Moreover, in CD patients, G908R and L1007finsC mutations were significantly associated with different phenotypes compared to CD wild-type patients. No association of IBD with the TLR4 SNPs was found in either cohort (allele frequencies: D299G-controls 3.9%, CD 3.7%, UC 3.4%, P > 0.05; T399I-controls 2.9%, CD 3.0%, UC 3.4%, P > 0.05). CONCLUSION: These findings confirm that, in our IBD patients selected from Southern Italy, the NOD2/CARD15, but not TLR4 SNPs, are associated with increased risk of CD.

Intestinal permeability and genetic determinants in patients, first-degree relatives, and controls in a high-incidence area of Crohn's disease in Southern Italy.

OBJECTIVE: A defect of gastrointestinal barrier function is considered to represent an important step in the pathogenesis of Crohn's disease (CD) but the mechanisms leading to an increased intestinal permeability (IP) are poorly understood. Since IP is influenced by pro-inflammatory mediators, it seems likely that a genetically determined abnormal immune response may lead to a loss of barrier function. METHODS: In a geographic area in Southern Italy with high incidence of CD we investigated IP (lactulose/mannitol testing) together with the three main mutations of the NOD2/CARD15 and the D299G polymorphism of the toll-like receptor (TLR)-4 gene in 23 families of CD patients (patients and first-degree relatives). RESULTS: Forty-eight percent of CD patients and 40% of their healthy relatives were found to have an abnormal IP compared to 5% of an appropriate control population (p < 0.0001). IP, however, was not associated with the L1007finsC mutation of the NOD2/CARD15 or the D299G variant of the TLR-4 gene. Allele frequency of the only L1007finsC mutation of CARD15 was significantly increased in patients (8.7%, p < 0.003) and in relatives (8.3%, p < 0.024) compared with controls (2.4%), whereas the D299G variant of the TLR-4 gene was found to be increased only in relatives (8.3%, p < 0.022), but not in patients (4.3%) compared with the control population (1.7%). CONCLUSIONS: There was no association between IP and genetic markers. Our findings showed a very high proportion of healthy first-degree relatives to bare alterations suggested to constitute determinants of CD. Mutations of NOD2/CARD15 or TLR-4, however, do not lead to permeability defects emphasizing the importance of additional environmental and/or genetic factors for pathogenesis.

A protective effect of the synthetic coumarine derivative Cloricromene against DNB-colitis in the rat.

Biologic therapies, namely antibodies against tumor necrosis factor-alpha (TNF- alpha) or its receptors, have been recently introduced for the treatment of patients with inflammatory bowel disease (IBD). In the present study the effects of cloricromene, an agent with known antithrombotic actions and with demonstrated anti-TNF- alpha activity were investigated in a rat model of experimental colitis induced with dinitrobenzenesulphonic acid (DNB)/ethanol. We investigated three experimental groups: (i) sham-colitis with vehicle-treatment (controls, n = 6), (ii) colitis with vehicle-treatment (saline, 0.1 ml s.c., daily) (DNB-V, n = 7), (iii) colitis with cloricromene-treatment (10 mg/kg/day s.c.; DNB-C, n = 8). After 7 days, the weight gain, colon wet weight, macroscopic damage score, coagulation parameters, colon mucosal myeloperoxidase activity (MPO), and tissue concentrations of TNF- alpha and of macrophage inhibitory peptide-2 (MIP-2) were assessed. The macroscopic damage scores, colon wet weights, and tissue MIP-2 levels were significantly increased in untreated and in cloricromene-treated rats compared with controls. Cloricromene treatment was associated with a minor body weight loss (p < 0.025) and significantly reduced tissue concentrations of MPO and TNF-alpha (p < 0.02, both). Blood coagulation parameters were not affected by treatment. In the DNB-model treatment with cloricromene effectively reduces tissue levels of TNF- alpha and of myeloperoxidase, whereas MIP-2 concentrations were not influenced. Blood coagulation parameters remained unchanged indicating safety of treatment. Since biological therapies frequently fail to improve disease course of IBD, other therapies with similar targets should be further investigated.