Influence of smoking habits and CARD15 mutations on the onset of Crohn's disease

Objective. Coeliac disease (CD), an autoimmune gluten-dependent enteropathy, can be associated with several extra-intestinal manifestations, including neurological disorders. At present, no data are available on the presence of hearing loss disorder in coeliac patients. The aim of the present study was to investigate the prevalence of hearing loss in coeliac patients compared with that in healthy controls. Material and methods. Twenty-four adult coeliac patients and 24 healthy subjects matched for gender, age, smoking and drinking habits were enrolled in the study. Among the coeliac patients, 6 were newly diagnosed and 18 patients were on a gluten-free diet for at least one year. Results. A hearing loss was found in 10 (47.1%) coeliac patients and 2 (9.1%) healthy controls. All CD patients with hearing loss presented a sensorineural hearing loss. The prevalence of hearing loss was significantly higher in coeliac patients than in healthy controls (p=0.01) but it was not significantly different between untreated (33.3%) and treated (44.4%) coeliac patients (p: NS). Conclusions. Despite the low number of subjects evaluated, the present study showed a higher prevalence of hearing loss in coeliac patients than in healthy controls, suggesting an association between CD and hearing loss. Immunological processes such as ear-specific and non-specific autoantibodies and vasculitis could be the basis of this association. Further longitudinal investigations on a larger sample size will be necessary to confirm the present data.     

CARD15 mutations in patients with Crohn's disease in a homogeneous Spanish population

OBJECTIVES: Three mutations in CARD15 have been repeatedly shown to be involved in Crohn's disease susceptibility, mainly in Caucasian individuals. However, those findings were not replicated in all populations studied so far. In this work, we studied the role of CARD15 mutations in a relatively homogeneous population from the Northwest of Spain, Galicia. METHODS: One hundred and sixty-five patients with Crohn's disease and a similar number of healthy controls were recruited from a single center in Galicia. All individuals were genotyped for the three main Crohn's disease associated CARD15 variants (R702W, G908R, and 1007fs). Association analyses were performed to study the influence of those mutations on Crohn's disease overall and on clinical subphenotypes. RESULTS: The allele frequencies of CARD15 variants were lower in this population than in most of the European populations studied so far. G908R and 1007fs were significantly associated with overall susceptibility to Crohn's disease. However, these associations were lost after stratification to clinical subgroups, probably due to the small number of cases in these subgroups. Significant associations were found between G908R and 1007fs and the behavior of Crohn's disease, but they were due to the influence of years of disease on the behavior of the disease rather than being the result of a direct effect of these mutations on disease behavior. CONCLUSIONS: The CARD15 variants G908R and 1007fs, but not R702W, are associated with susceptibility to Crohn's disease in Galicia. Interestingly, the frequency of these mutations appears to be lower than in other Caucasian populations studied so far.     

Clinical applications of NOD2/CARD15 mutations in Crohn's disease

The recent identification of the CARD15/NOD2 gene as a susceptibility locus for Crohn's disease represents an important step in the immunopathogenesis of inflammatory bowel disease. The gene explains about 20% of the genetic susceptibility CARD15 mutations are present in 30-50% of CD patients compared to 7-20% of healthy controls. The three risk alleles R702W, G908R and 1007fsInsC in NOD2 associated with susceptibility to Crohn's disease have demonstrated a remarkable amount of heterogeneity across ethnicities and populations, with regional variation across Europe. In non-Caucasian populations Crohn's disease continues to increase in incidence but this increase appears not to be a consequence of variation in NOD2. Genotype-phenotype analyses demonstrated an association of these mutations with ileum-specific disease and an increased incidence of the fibrostenotic phenotype. Although CARD15 variants do not predict response to the TNF alpha monoclonal antibodies, there are no data available on the possible influence of CARD15 mutations on response to other drugs. Screening for CARD15 mutations in order to identify high-risk individuals or to introduce an individualized disease management is therefore currently not recommended.     

Mutations in CARD15 and smoking confer susceptibility to Crohn's disease in the Danish population

OBJECTIVE: Three CAspase Recruitment Domain (CARD15) mutations have shown to predispose to Crohn's disease in Caucasian populations. The aim of this study was to investigate the mutation frequency in patients with inflammatory bowel disease and in healthy controls in Denmark. MATERIAL AND METHODS: Genotyping of the three common CARD15 mutations was carried out on 388 patients with Crohn's disease, 565 patients with ulcerative colitis and 796 healthy controls using real-time PCR. Allele and genotype frequencies in the three groups were compared. A possible additive effect of smoking on CARD15 mutations was also examined. RESULTS: Carrying at least one CARD15 mutation was significantly more common in patients with Crohn's disease compared with healthy controls (21% versus 10%; p <0.001). A gene-dosage effect was observed (ORadj.smoking 22.2; p <0.001 for carrying two CARD15 mutations versus ORadj.smoking 1.8; p=0.01 for carrying one CARD15 mutation). The 1007insC protein truncating mutation was the major contributing mutation. Ileal involvement was more common in Crohn's disease patients with CARD15 mutations as opposed to patients without CARD15 mutations (ORadj.smoking 3.6; p <0.001). Smoking was independently associated with Crohn's disease (OR 1.8; p <0.001), but no multiplicative effect of smoking on CARD15 genotypes was found. CONCLUSIONS: In the Danish population, CARD15 mutations were found to be associated with Crohn's disease, hence supporting the hypothesis of a genetic component contributing to the disease. Further research for other genes possibly involved in Crohn's disease may result in the use of genetic testing for diagnosis or treatment of Crohn's disease in the future.     

Mutations in CARD15 and smoking confer susceptibility to Crohn's disease in the Danish population

Objective. Three CAspase Recruitment Domain (CARD15) mutations have shown to predispose to Crohn's disease in Caucasian populations. The aim of this study was to investigate the mutation frequency in patients with inflammatory bowel disease and in healthy controls in Denmark. Material and methods. Genotyping of the three common CARD15 mutations was carried out on 388 patients with Crohn's disease, 565 patients with ulcerative colitis and 796 healthy controls using real-time PCR. Allele and genotype frequencies in the three groups were compared. A possible additive effect of smoking on CARD15 mutations was also examined. Results. Carrying at least one CARD15 mutation was significantly more common in patients with Crohn's disease compared with healthy controls (21% versus 10%; p <0.001). A gene–dosage effect was observed (OR_adj.smoking 22.2; p<0.001 for carrying two CARD15 mutations versus OR_adj.smoking 1.8; p=0.01 for carrying one CARD15 mutation). The 1007insC protein truncating mutation was the major contributing mutation. Ileal involvement was more common in Crohn's disease patients with CARD15 mutations as opposed to patients without CARD15 mutations (OR_adj.smoking 3.6; p<0.001). Smoking was independently associated with Crohn's disease (OR 1.8; p<0.001), but no multiplicative effect of smoking on CARD15 genotypes was found. Conclusions. In the Danish population, CARD15 mutations were found to be associated with Crohn's disease, hence supporting the hypothesis of a genetic component contributing to the disease. Further research for other genes possibly involved in Crohn's disease may result in the use of genetic testing for diagnosis or treatment of Crohn's disease in the future.     

CARD15 mutations are poorly related to Crohn's disease phenotypes in Asturias]

BACKGROUND: the association between the three common CARD15 gene mutations (R702W, G908R, L1007fs) and the genetic susceptibility to Crohn s disease (CD) have been confirmed by several studies, with some differences found, in relation to geographic areas and ethnic groups. OBJECTIVES: To analyze the prevalence of CARD15 gen and its polymorphisms in patients with CD in Asturias and its possible correlation with the different genotypes of the disease. METHODS: a total of 216 CD patients recruited from Asturias (North of Spain) and 86 ethnically matched healthy controls, were typed using Hybprobes on a LightCycler instrument for CARD15 mutations. Patients were subdivided according to Vienna classification. We have studied the frequency of these mutations in the different subgroups of CD patients and analyzed its contribution to the disease clinical characteristics and progression. RESULTS: carrier frequencies for CARD15 mutations in our CD patients were similar to controls (17.8 vs. 17.4%) respectively (NS). CD patients exhibited frequencies of 8.8, 3.0 and 6.0% for the R702, G908R and L1007fs polymorphisms respectively, whereas our control population had allele frequencies of 11.6, 2.3 and 3.5% for the three mutations respectively (NS).We did not find any relationship between CARD15 mutations and the different phenotypes of Crohn s disease, according to Vienna classification. CONCLUSIONS: in our CD population, other factors (i.e. environmental), in addition to genetics, must be mainly involved in the development of the disease.     

CARD15 gene mutations and risk for early surgery in pediatric-onset Crohn's disease.

BACKGROUND & AIMS: The risk for Crohn's disease (CD) is determined in part by genetic factors. Three recently described mutations in the CARD15(NOD2) gene have been associated with adult-onset CD. We investigated the effect of CARD15 mutations on disease manifestation, disease progression, and the risk for early surgery in childhood-onset CD. METHODS: Genotyping for 3 CARD15 mutations: R702W, G908R, and 3020insC, was performed in 186 children with CD from a prospective cohort. A transmission-disequilibrium test was used to test for association with CD. Genotype with disease location and behavior was tested with logistic regression analysis. The effect of mutations on surgical outcome was evaluated using a Cox proportional hazard analysis. RESULTS: The mean age at CD diagnosis was 12.4 years. The frequency of allelic mutations observed was 6.6% for R702W, 6% for G908R, and 13.1% for 3020insC. Of Caucasian CD children, 42% had at least one CARD15 mutation. None of the non-Caucasian children with CD had any CARD15 mutation. A significant association was detected for 3020insC (P = .0045). Ileal location (odds ratio, 4.3; P = .003) and stricturing disease (odds ratio, 6.6; P = .0001) was more frequent and the risk for surgery was higher (hazard ratio, 5.8; P < .0001) and surgery occurred earlier (hazard ratio, 2.24) in those children with 3020insC mutation compared with those without 3020insC. CONCLUSIONS: In children with pediatric-onset CD, early development of stricturing behavior leading to surgical resection is influenced by ileal location and 3020insC variant of the CARD15 mutation. Genetic testing may identify children with CD who are at risk for early surgery.     

Identification of NOD2/CARD15 mutations in Malaysian patients with Crohn's disease

OBJECTIVE: The NOD2/CARD15 gene has been identified as an important susceptibility gene for Crohn's disease (CD) but the three common disease predisposing mutations (DPM) found in developed countries have not been identified in Asian populations. The aim of our study was to look for the DPM in our multiracial population and to discover whether there were any differences in the three major ethnic groups; Malay, Chinese and Indian. METHODS: Blood samples from consecutive CD patients and healthy controls were obtained and analyzed for the three common mutations (R702W, G908R, 1007fs) but in addition to this, we also looked for the SNP5 and JW1 variants which are associated with CD in Ashkenazi Jews. A polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) technique was used to identify the mutations, which was confirmed by sequencing. The baseline socio‐demography and clinical characteristics of the CD patients were recorded. RESULTS: Overall 45 patients (three Malays, 15 Chinese, 26 Indians and one other) with confirmed CD and 300 controls were recruited. The three common DPM were not observed in either the CD patients or the controls. Neither the SNP5 nor the JW1 mutation was found in any of the controls. However, the SNP5 mutation was identified in six (13.3%) Indian CD patients and the JW1 mutation in eight CD patients who are different from those carrying the SNP5 mutation: one Malay (33.3%), two Chinese (13.3%), one other (Portuguese) and four Indians (15.4%). The presence of SNP5 was strongly associated with CD in the Indian population and that of JW1 was strongly associated with CD overall and in each of the major ethnic groups. There was a trend towards a younger age of onset and stricturing disease in patients carrying the JW1 mutation. CONCLUSION: These findings suggest the presence of novel DPM in the NOD2/CARD15 gene in Asian patients with CD.     

NOD2/CARD15 mutations and presence of granulomas in pediatric and adult Crohn's disease

OBJECTIVES: The etiology and mechanism leading to granuloma formation in patients with Crohn's disease (CD) are presently unknown. The first susceptibility gene to be identified as a risk factor for CD is the NOD2/CARD15 gene on Chromosome 16. Mutations in NOD2 could affect the intracellular response to bacterial products and may eventually lead to granuloma formation. The association between NOD2 and granulomas has not been previously explored. We evaluated a possible association between NOD2 mutations and granuloma formation, and compared the prevalence of granulomas in both pediatric and adult cohorts. METHODS: Patients were consecutively recruited through pediatric gastroenterology and adult gastroenterology programs. Patients were eligible if CD was confirmed, and they had undergone full colonoscopy with biopsy and/or surgical resection. Patients underwent genotyping for NOD2 disease-associated mutations. RESULTS: A total of 230 patients were enrolled into the study, of whom 169 patients met all inclusion/exclusion criteria (Group 1, 77 patients [age range 1-16 years]; Group 2, 92 patients [age range 17-68 years]). Surgical resection was performed more often in adults (P < 0.005), and gastroscopy was performed more frequently in children (P < 0.001). Granulomas were found in 34% of the patients studied. The prevalence of granulomas did not differ by age, age group, or gender. A disease-associated NOD2 mutation was found in 37.8% of patients. Granulomas were found in 39% of patients with NOD2 mutations compared with 31% of those without NOD 2 mutations (difference was not significant). In addition, no difference was noted for the specific mutations. CONCLUSIONS: We did not find any correlation between NOD2 mutations and granuloma formation. The cause of granulomas in CD remains elusive.     

Association of NOD2/CARD15 mutations with previous surgical procedures in Crohn's disease.

OBJECTIVE: The aim of this study is to assess the importance of NOD2/CARD15 gene mutations as prognostic factors for surgical indications in Crohn's disease. PATIENTS AND EXPERIMENTAL DESIGN: A total of 165 Crohn s disease patients were studied, considering previous surgery related to Crohn's disease. We analyzed for previous surgery in global procedures as well as separately for the two main surgical indications: ileal resection and fistula treatment. The need for appendectomy was also studied. All patients were genotyped for the three CARD15 mutations, and association studies were developed using Chi-square statistics and Fisher's exact test whenever appropriate. RESULTS: Carriers of the G908R or 1007fs mutation needed surgery more frequently, both for ileal resection and fistula repair. In contrast, appendectomy was not associated with CARD15 mutations. CONCLUSIONS: As previously reported in this population, the R702W mutation does influence parameters of disease or need of surgery. The need for Crohn's disease-related surgery is higher in carriers of the G908R or 1007fs CARD15 mutation in the Galician population. Nevertheless, the frequency of these mutations does not allow their use to predict the course of disease.     

Association of NOD2/CARD15 mutations on Crohn's disease phenotype in an Italian population

AIMS: To confirm the prevalence of NOD2/CARD15 mutations in Italian inflammatory bowel disease patients and to define the role of the different mutations on Crohn's disease phenotype. PATIENTS AND METHODS: A total of 177 patients with Crohn's disease and 92 patients with ulcerative colitis and 164 control participants were investigated for the presence of Arg702Trp, Gly908Arg and Leu1007fsinsC NOD2/CARD15 mutations. Allele frequencies in Crohn's disease and ulcerative colitis patients were compared with those observed in the control population. Genotype-phenotype correlations with the major clinical features were also established and estimated risks (odds ratio with 95% confidence interval) for the mutations were calculated by logistic regression and multiple correspondent analysis. RESULTS: Gly908Arg and Leu1007fsinsC mutations were significantly more frequent in Crohn's disease patients compared with healthy controls (P<0.01 and <0.003 respectively). Indeed, using a logistic regression model adding terms for age (differently distributed between cases and controls) and sex, a significantly increased risk of having Crohn's disease compared with healthy controls was found for all NOD2 mutations: Leu1007fsinsC (odds ratio=7.35; 95% confidence interval: 1.73-31.3), Gly908Arg (odds ratio=5.70; 95% confidence interval: 1.37-23.7) and Arg702Trp (odds ratio=2.45; 95% confidence interval: 1.10-5.47). As far as the genotype-phenotype correlations are concerned, by multivariate conditional logistic regression methods, we found a significant association between Gly908Arg mutations and familial history of inflammatory bowel disease, between Leu1007fsinsC mutations and appendectomy and between Arg702Trp mutations and fibrostenotic phenotype of Crohn's disease. A nonsignificant association between Arg702Trp variants and ileal disease was also found (odds ratio=8, 95% confidence interval: 0.99-64.9). CONCLUSIONS: The results of the study confirm a significant association of CARD15 gene mutations in our Italian Crohn's disease population and the impact of different NOD2/CARD15 mutations on specific disease phenotypes.     

NOD2/CARD15 mutations in Croatian patients with Crohn's disease: prevalence and genotype-phenotype relationship

BACKGROUND: Crohn's disease (CD) is a chronic inflammatory disorder of the gastrointestinal tract with variations in localization and behaviour. Mutations in the NOD2/CARD15 gene on chromosome 16q have been implicated in the pathogenesis of the disease and three main sequence variants, all single nucleotide polymorphisms (SNPs), have been identified in North American and European populations. AIMS AND METHODS: As no data exist in the Croatian population, we consecutively collected a cohort of 136 CD patients and 91 healthy controls to determine the prevalence of NOD2/CARD15 mutations and their association with phenotypic expression of the disease. All patients and controls were genotyped for Arg702Trp (Hugot SNP8), Gly908Arg (Hugot SNP12), and Leu1007fsinsC (Hugot SNP13) and allele frequencies were compared between the Crohn's patients and controls. The correlation of NOD2/CARD15 genotypes with the phenotypic expression of Crohn's disease was further assessed by logistic regression analysis. RESULTS: NOD2/CARD15 variants were found in 38/136 CD patients (27.9%) compared to 10/91 (10.9%) healthy controls (P = 0.0022). Allele frequencies in patients with CD were 13.97%, 4.4% and 11.76%, respectively, for SNP8, 12 and 13, compared to 5.49%, 1.12% and 4.40% in controls (P = 0.041, P = 0.162, P = 0.055). Six CD patients carried double mutations and, remarkably, we identified two homozygous mutants amongst the healthy control group. Surgery over the course of the disease and a younger age at onset of the disease were significantly more frequent in patients who were carriers of NOD2/CARD15 mutations. CONCLUSIONS: This report on NOD2/CARD15 mutations in Croatian patients with CD demonstrates that this gene is also implicated in susceptibility to CD in the Croatian population. Phenotypic association showed a younger age at diagnosis and a higher need for surgery in patients carrying NOD2/CARD15 mutations. However, the prevalence is somewhat lower compared to other reports, likely due to a more prominent colonic inflammation.     

NOD2/CARD15 disease associations other than Crohn's disease

At this moment, few confirmed associations between NOD2 mutations and diseases other than Crohn's disease (CD) and Blau syndrome (BS) have been reported, but research is ongoing in several fields where a genetic susceptibility factor and/or a role for the innate immune system is suspected. Whether the Crohn's-associated CARD15 mutations lead to a loss or gain of function of the NOD2 receptor is subject to controversy, and by which mechanisms this change in function might increase the susceptibility to CD is still under investigation. The possible involvement of NOD2/CARD15 in the pathogenesis of certain diseases with already (partially) unraveled pathophysiologic mechanisms might contribute to our further understanding of NOD2/CARD15 and its function in CD. We review studies on the association of CARD15 variants with diseases other than CD. The association of NOD2/CARD15 mutations with CD and BS, and possibly also early onset sarcoidosis, suggests a role for the gene in the development of granulomata and granulomatous diseases, possibly by inappropriate activation of the immune system. The data from the oncology field suggest that this inappropriate activation might even lead to uncontrolled proliferation of certain cell types. The studies in allergic diseases and atopy are the largest so far, and the association of NOD2/CARDI5 mutations with atopic phenotypes might be an indication that CARD15 also plays a role in the Th2 pathway. Finally, transplantation studies indicate that the genetic background of a patient should be taken into account when considering hematopoietic stem cell transplantation, given the increased risk of mortality and graft versus host disease observed. Whether NOD2 variants are also associated with an increased risk for infections and sepsis in patients receiving immunosuppressive therapies is unclear.