NOD2/CARD15 genotype and common gastrointestinal diseases in 43 600 individuals

Abstract. Yazdanyar S, Nordestgaard BG (Herlev Hospital, Herlev; Copenhagen University Hospital, University of Copenhagen, Copenhagen; Denmark). NOD2/CARD15 genotype and common gastrointestinal diseases in 43 600 individuals. J Intern Med 2010; 267 228–236. Objectives. NOD2/CARD15 is involved in the innate immune response and three polymorphisms in this gene (Arg702Trp rs2066844, Gly908Arg rs2066845 and Leu1007fsinsC rs5743293) have been associated with risk of the rare Crohn’s disease. We tested the hypothesis that polymorphisms in NOD2/CARD15 associate with risk of nine common gastrointestinal diseases. Design and setting. We genotyped 43 596 white individuals from the Danish general population followed for 31 years, during which time 782 developed oesophagitis and reflux, 1395 ulcus ventriculi and duodeni, 1384 gastritis and dyspepsia, 1407 appendicitis, 646 irritable bowel syndrome, 1301 infectious diseases of the gastrointestinal tract, 681 anal fissure, fistula and abscess, 826 gastrointestinal cancer and 161 developed cancer in liver and pancreas. Results. Some 89% were non‐carriers, 11% heterozygotes, 0.15% homozygotes and 0.23% compound heterozygotes. Cumulative incidences differed by genotype for appendicitis (log‐rank P = 0.02), anal fissure, fistula and abscess (P = 0.003) and gastrointestinal cancer (P = 0.004), but not for any of the other endpoints. Compared with non‐carriers, age and sex adjusted hazard ratios were 2.7 (95%CI 1.4–5.5) for appendicitis amongst compound heterozygotes, 3.2 (1.3–7.8) for anal fissure, fistula and abscess amongst compound heterozygotes, and 3.8 (1.6–9.2) for gastrointestinal cancer amongst homozygotes, whilst other genotypes did not have increased risk. The increased risk of gastrointestinal cancer amongst homozygotes appeared to be similar amongst both men and women and amongst those below or above 60 years, and likely included both upper gastrointestinal cancer and colorectal cancer. Conclusions. NOD2/CARD15 polymorphisms are not major risk factors for common gastrointestinal diseases; however, we cannot completely exclude association with appendicitis, anal fissure, fistula and abscess, and gastrointestinal cancer.     

NOD2/CARD15 gene mutations in North Algerian patients with inflammatory bowel disease

AIM: To analyse allelic frequency of NOD2 gene variantsand to assess their correlation with inflammatory bowel disease(IBD) in Algeria.METHODS: We studied 132 unrelated patients diagnosed with IBD, 86 with Crohn's disease(CD) and 46 with ulcerative colitis(UC). Data was prospectively collected between January 2011 and December 2013. The demographic and clinical characteristics were recorded for all the patients. A group of 114 healthy unrelated individuals were selected as controls. All groups studied originated from different regions of North Algeria and confirmed the Algerian origin of their parents and grandparents. Informed and written consent was obtained from each of the participants. All individuals were genotyped for the three CDassociated NOD2 variants(p.Arg702 Trp, p.Gly908 Arg and p.Leu1007 fsins C mutations) using the polymerase chain reaction-restriction fragment length polymorphism method. Allele and genotype frequencies in patients and control subjects were compared by χ2 test and Fisher's exact test where appropriate. Odds ratios(OR) and 95% confidence intervals(95%CI) were also estimated. Association analyses were performed to study the influence of these variants on IBD and on clinical phenotypes.RESULTS: The p.Arg702 Trp mutation showed the highest frequency in CD patients(8%) compared to UC patients(2%)(P = 0.09, OR = 3.67, 95%CI: 0.48-4.87) and controls(5%)(P = 0.4, OR = 1.47, 95%CI: 0.65-3.31). In CD patients allelic frequencies of p.Gly908 Arg and p.Leu1007 fsins C variants compared to HC were 3% vs 2%(P = 0.5, OR = 1.67, 95%CI: 0.44-6.34); 2% vs 1%(P = 0.4 OR = 2.69 95%CI: 0.48-14.87 respectively). In UC patients, allelic frequencies of p.Gly908 Arg and p.Leu1007 fsins C variants compared to HC were 1% vs 2%(P = 1, OR = 1.62, 95%CI: 0.17-4.74) and 2% vs 1%(P = 0.32, OR = 0.39, 95%CI: 0.05-2.87). The total frequency of the mutated NOD2 chromosomes was higher in CD(13%), than in HC(8%) and UC(5%). In addition, NOD2 variants were linked to a particular clinical sub-phenotype in CD in this Algerian cohort. As expected, the three NOD2 variants showed a significant association with CD but did not reach statistical significance, despite the fact that the allele frequency of NOD2 variants was in the range found in most of the European populations. This might be due to the non-exposure of the NOD2 carriers to environmental factors, required for the expression of the disease.CONCLUSION: Further analyses are necessary to study genetic and environmental factors in IBD in the Algerian population, using larger patient groups.     

NOD2/CARD15, NOD1/CARD4, and ICAM-1 gene polymorphisms in Turkish patients with inflammatory bowel disease

Purpose The genetic susceptibility of people with certain NOD2/CARD15, NOD1/CARD4, and ICAM-1 gene variants to inflammatory bowel disease is still under investigation. The aim of this study was to investigate polymorphisms in the NOD2/CARD15 (R702W, G908R, and 3020insC), NOD1/CARD4 (E266K, D372N), and ICAM-1 (G241R, K469E) genes, which are known to be associated with inflammation, in Turkish patients with inflammatory bowel disease and healthy control groups. Methods The genotypes of 70 patients with endoscopically and histopathologically diagnosed Crohn's disease (38 men, 32 women; mean age, 38.8 ± 1.3), 120 patients with ulcerative colitis (67 men, 53 women; mean age, 41.7 ± 1.3) and 106 healthy control subjects (37 men, 69 women; mean age, 35.7 ± 1.4), who stated that they had never had any prior bowel disease history, were compared. A polymerase chain reaction-restriction fragment length polymorphism analysis was performed for two variants of the ICAM-1 gene, the three main variants of the NOD2/CARD15 gene, and the E266K variant of the NOD1/CARD4 gene, and DNA sequencing was used for the D372N polymorphism of the NOD1/CARD4 gene. Results In this study, the three previously described Crohn's disease-predisposing variants of the NOD2/CARD15 gene and the polymorphisms examined in the NOD1/CARD4 and ICAM-1 genes were not found to be associated with ulcerative colitis or Crohn's disease. Conclusions These findings suggest that the polymorphisms observed in the NOD2/CARD15, NOD1/CARD4, and ICAM-1 genes are not genetic susceptibility factors for Crohn's disease or ulcerative colitis in Turkey.     

Continuing low incidence of Crohn’s disease in Northwest Greece

AIM OF STUDY: The largest population-based study for inflammatory bowel disease in Northwest Greece. MATERIALS AND METHODS: A retrospective survey for the years 1982-1997. RESULTS: Of 400 patients, 334 had ulcerative colitis, 43 Crohn's disease and 23 indeterminate colitis. CONCLUSIONS: Crohn's disease still remains rare in Northwest Greece.     

Prediction of Crohn’s disease aggression through NOD2/CARD15 gene sequencing in an Australian cohort

AIM:To investigate the association between mutations in oligomerisation domain 2/caspase recruitment domains 15(NOD2/CARD15)and the natural history of Crohn’s disease(CD)to identify patients who would benefit from early aggressive medical intervention.METHODS:We recruited thirty consecutive unrelated CD patients with a history of ileo-caecal or small bowel resection during the period 1980-2000;Fifteen patients of these had post-operative relapse that required further surgery and fifteen did not.Full sequencing of the NOD2/CARD15 gene using dHPLC for exons 3,5,7,10 and 12 and direct sequencing for exons 2,4,6,8,9 and 11 was conducted.CD patients categorized as carrying variants were anyone with at least 1 variant of the NOD2/CARD15 gene.RESULTS:About 13.3%of the cohort(four patients)carried at least one mutant allele of 3020ins C of the NOD2/CARD15 gene.There were 20 males and 10females with a mean age of 43.3 years(range 25-69years).The mean follow up was 199.6 mo and a median of 189.5 mo.Sixteen sequence variations within the NOD2/CARD15 gene were identified,with 9 of them occurring with an allele frequency of greater than 10%.In this study,there was a trend to suggest that patients with the 3020ins C mutation have a higher frequency of operations compared to those without the mutation.Patients with the 3020insC mutation had a significantly shorter time between the diagnosis of CD and initial surgery.This study included Australian patients of ethnically heterogenous background unlike previous studies conducted in different countries.CONCLUSION:These findings suggest that patients carrying NOD2/CARD15 mutations follow a rapid and more aggressive form of Crohn’s disease showing a trend for multiple surgical interventions and significantly shorter time to early surgery.     

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.     

Association between polymorphisms in the Toll-like receptor 4, CD14, and CARD15／NOD2 and inflammatory bowel disease in the Greek population

AIM: Crohn's disease (CD) and ulcerative colitis (DC) are multifactorial diseases with a significant genetic background. Apart from CARD15/NOD2 gene, evidence is accumulating that molecules related to the innate immune response such as CD14 or Toll-like receptor 4 (TLR4), are involved in their pathogenesis. In further exploring the genetic background of these diseases, we investigated the variations in the CARD15/NOD2 gene (Arg702Trp, Gly908Arg and Leu1007fsinsC), and polymorphisms in the TLR4 gene (Asp299Gly and Thr399Ile) as well as in the promoter of the CD14 gene (T/C at position -159) in Greek patients with CD and UC. METHODS: DNA was obtained from 120 patients with CD, 85 with UC and 100 healthy individuals. Genotyping was performed by allele specific PCR or by PCR-RFLP analysis. RESULTS: The 299Gly allele frequency of the TLR4 gene and the T allele and TT genotype frequencies of the CD14 promoter were significantly higher in CD patients only compared to healthy individuals (P= 0.026<0.05; P= 0.0048<0.01 and P= 0.047<0.05 respectively). Concerning the NOD2/CARD15 mutations the overall presence in CD patients was significantly higher than that: in UC patients or in controls. Additionally, 51.67% of the CD patients were carriers of a TLR4 and/or CD14 polymorphic allele and at least one variant of the NOD2/CARD15, compared to 27% of the UC patients. It should be pointed out that both frequencies significantly increased as compared with the 10% frequency of multiple carriers found in healthy controls. A possible interaction of the NOD2/CARD15 with TLR4 and especially CD14, increased the risk of developing inflammatory bowel disease (IBD). CONCLUSION: Our results indicate that co-existence of a mutation in either the TLR4 or CD14 gene, and in NOD2/CARD15 is associated with an increased susceptibility to developing CD compared to UC, and to developing either CD or UC compared to healthy individuals.     

Crohn’s disease incidence evolution in North-western Greece is not associated with alteration of NOD2/CARD15 variants

AIM To assess the trends in the incidence of inflammatory bowel disease (IBD) over 23 years in the same area and to identify genetic factors related to incidence evolution.METHODS Patients with IBD arising from Northwestern Greece were systematically recorded through the 1983-2005 period. Trends in disease incidence and genetic patterns related to CARD15 variants were documented and correlated.RESULTS A total of 447 patients with IBD were recorded (23.5% Crohn's disease, 72.7% Ulcerative colitis and 3.8% indeterminate colitis). Mean annual incidence rates of CD and UC were 0.9/100000 (95% CI 0.1-1.7) and 2.7/100000 (95% CI 1.7-4.1) inhabitants,respectively. There was a statistically significant increase of CD incidence (P ＜ 0.01) during the study period, in contrast to the UC incidence. There were no statistical differences in CARD15 variants over the study period.CONCLUSION The incidence of CD in North-western Greece has risen disproportionately to that of UC in the 21st century. This is not related to alterations of genetic background though.     

Comparison between methotrexate and azathioprine in the treatment of chronic active Crohn’s disease: a randomised, investigator-blind study

BACKGROUND AND AIMS: The efficacy of azathioprine in the treatment of chronic active Crohn's disease is well established. However, this drug has a long onset of action. Methotrexate has also been shown to be effective in chronic active Crohn's disease. The aim of this study was to evaluate the efficacy and safety of methotrexate in comparison with azathioprine, and to establish whether methotrexate has a shorter onset of action in this setting. METHODS: Patients with chronic active Crohn's disease were admitted to this investigator-blind study. Chronicity was defined as the need for steroid therapy of > or = 10 mg/day for at least 4 months during the preceding 12 months, with at least one attempt to discontinue treatment. The disease had to be clinically active at entry, with a Crohn's Disease Activity Index of > or = 200. Six patients treated with azathioprine and methotrexate, respectively, were found to have enterocutaneous and perianal fistulas. At entry, all patients received prednisolone (40 mg once a day) which was tapered over a period of 12 weeks unless their clinical condition deteriorated. All patients were randomised to receive i.v. methotrexate 25 mg/week, or oral azathioprine 2 mg/kg per day, for a 6-month follow-up period. After the first 3 months, methotrexate was switched to oral administration maintaining the same dose. The primary efficacy outcome considered was the proportion of patients entering first remission after 3 and 6 months of therapy. Clinical remission was defined as the lack of need for steroid treatment and a Crohn's Disease Activity Index score of < or = 150 points at each scheduled visit. RESULTS: In the 54 patients (26 F, 28 M, mean age 34 years, range 18-60) randomly assigned to methotrexate (n=27) or azathioprine (n=27), no statistically significant difference was found between the two treatment regimens with respect to remission rate after 3 (methotrexate 44%, azathioprine 33%, p=0.28, (95% CI, 0.369-0.147), and 6 months (methotrexate 56%, azathioprine 63%, p=0.39, 95% CI, 0.187-0.335), respectively. Six patients withdrew from therapy due to adverse events: 3/27 (11%) in methotrexate and 3/27 (11%) in azathioprine. Drug-related adverse events (asthenia, nausea and vomiting) that did not require withdrawal from therapy were more frequent in the methotrexate group (azathioprine: 2/27 (7%); methotrexate: 12/27 (44%), p=0.00009). The frequency of these adverse events was comparable during the intravenous or oral administration of the drug. CONCLUSIONS: This study confirms that methotrexate is effective in inducing remission in patients with chronic active Crohn's disease, therapeutic efficacy being comparable, but not faster, than that of azathioprine.     

NOD2/CARD15基因多态性与克罗恩病患者相关性研究

目的NOD2/CARD15基因是人类的第一个克罗恩病(CD)易感基因,其间的3个单核苷酸多态性(SNPs)与白种人CD有显著性相关,但与日本人无关。本研究旨在证实这3个SNPs是否与浙江地区人群的CD易感性有关。方法血样来自浙江地区32例CD患者,110例溃疡性结肠炎患者及292例健康对照者。通过PCRSSP方法直接检测野生型及NOD2/CARD15基因的3个多态性(Arg702Trp,Gly908Arg,Leu1007fsinsC)。结果没有发现1例CD患者纯合子或杂合子的SNPs突变,同样在溃疡性结肠炎患者和健康人中也未能检测到。结论本研究表明一些存在于特定人群的CD易感基因可能在其他人群中不存在,白种人CD患者相关的易感基因NOD2/CARD15常见的3个SNPs与浙江地区CD人群无关。     

NOD2/CARD15基因突变与中国人克罗恩病相关性研究

背景 NOD21CARDl5 基因是人类克罗恩病(Crohn's disease,CD)第一个易感基因,既往研究发现P268S可能与中国人CD发病及临床特征相关.目的 本研究旨在证实P268S与中国人CD发病及其临床特征的相关性.方法 血样来自临床确诊的50例CD患者,60例溃疡性结肠炎(ulcerative colitis,uc) 患者及100 例健康体检者(healthy controls,HC).提取人血白细胞基因组DNA,PCR 扩增目的片段,PCR-RFLP发现突变位点,DNA 测序证实突变位点.结果 共有8例CD患者发现有P268S改变,而在UC患者和 HC 中分别发现2例和3例P268S改变,CD组明显高于UC和HC组(x2=10.829,P=0.004),而UC组和HC组无明显差异.8例有P268S改变的CD患者临床特征包括病变多位于回肠.发病年龄轻(6例<20岁),常并发肠腔狭窄而需手术治疗,中一重度患者比例高.结论 P268S可能是NOD21CARDl5 基因中与中国人CD相关的SNP.P268S与CD患者发病年龄、病变部位及并发症及病情严重程度可能相关.     

NOD2／CARDl5基因突变与中国人克罗恩病相关性研究

背景NOD2／CARDl5基因是人类克罗恩病（Crohn＇s disease，CD）第一个易感基因，既往研究发现P268S可能与中国人CD发病及临床特征相关。目的本研究旨在证实P268S与中国人CD发病及其临床特征的相关性。方法血样来自临床确诊的50例CD患者，60例溃疡性结肠炎（ulcerativecolitis，uc）患者及100例健康体检者（healthy controls，HC）。提取人血白细胞基因组DNA．PCR扩增目的片段，PCR—RFLP发现突变位点，DNA测序证实突变位点。结果共有8例CD患者发现有P268S改变，而在UC患者和HC中分别发现2例和3例P268S改变，CD组明显高于UC和HC组（X^2=10．829，P=0．004），而UC组和HC组无明显差异。8例有P268S改变的CD患者临床特征包括病变多位于回肠．发病年龄轻（6例〈20岁），常并发肠腔狭窄而需手术治疗，中一重度患者比例高。结论P268S可能是NOD2/CARDl5基因中与中国人CD相关的SNP。P268S与CD患者发病年龄、病变部位及并发症及病情严重程度可能相关。     

Infliximab heals intestinal inflammatory lesions and restores growth in children with Crohn’s disease

BACKGROUND: Infliximab has recently emerged as an efficacious agent for patients with severe Crohn's disease. There are only few studies on the use of infliximab in children with Crohn's disease: most of them are retrospective and deal only with the clinical response to the drug. AIM: We aimed at assessing the efficacy of infliximab in children and adolescents with severe Crohn's disease recruited consecutively and followed up prospectively at a single centre. Clinical response, intestinal inflammation and growth pattern were evaluated. PATIENTS: Eighteen patients entered into the trial (median age: 13 years, range: 6-18). They were referred because of severe symptoms with unsatisfactory response to conventional drugs. METHODS: All patients received a baseline schedule of three intravenous infusions of infliximab (0, 2 and 6 weeks), 5 mg/kg. Paediatric Crohn's Disease Activity Index, nutritional and activity serum variables, and ileocolonoscopy (with histology) were evaluated before and 8 weeks after beginning the therapy. All patients had long-term administration of azathioprine (2 mg/kg per day). After the baseline schedule, eight patients had a retreatment infusion of infliximab (5 mg/kg) every 8 weeks. Weight and height Z scores were measured before starting the baseline infusion programme and after 6 months. RESULTS: After 8 weeks of therapy, there was a dramatic improvement in Paediatric Crohn's Disease Activity Index, in nutritional and activity blood parameters, as well as in endoscopic and histological scores; 10 patients had a clinical remission (Paediatric Crohn's Disease Activity Index < or = 10), 12 patients had an inflammatory remission (decrease in both endoscopic and histological scores for > or = 50% as compared to baseline values). In all patients corticosteroids were stopped within 4 weeks after beginning infliximab therapy. After 6 months of therapy, Paediatric Crohn's Disease Activity Index was markedly lower than the pre-treatment value; however, it was significantly lower in patients on retreatment than in those who received only three infusions of infliximab. Furthermore, a significant increase in both weight and height Z scores was observed 6 months after beginning of the baseline infusion programme. Moreover, weight and height gain was significantly higher in patients on retreatment rather than in those treated only with three baseline infusions of infliximab. Mild infusion reactions controlled by slowing infusion rate were observed in four patients. No delayed hypersensitivity-like reactions were seen. CONCLUSIONS: In children with severe Crohn's disease, infliximab is a safe and valuable treatment in inducing remission, in healing inflammatory lesions of the gut, as documented by endoscopy and histology, and in promoting growth. Retreatment infusions of infliximab may be suggested in childhood-onset Crohn's disease to maintain remission and reverse growth failure.     

An assessment of serum vitamin B12 and folate in patients with Crohn’s disease

Crohn’s disease is a chronic inflammatory condition that can involve any area in the gastrointestinal tract often involving the distal ileum where vitamin B12 is specifically absorbed. The aim of this study was to ascertain serum vitamin B12 and folate levels in order to investigate the correlation among these vitamin levels and disease activation, localization, duration and age at the onset of the disease.Study population included 103 patients with Crohn’s disease and a healthy control group of 114 individuals. C-reactive protein, vitamin B12, folate levels were studied along with hemogram analyses. The results were evaluated in statistical comparisons. While serum vitamin B12 levels and serum folate levels were 161.9 ± 63.2(73–496) pg/mL and 4.9 ± 1.4(1.2–9.4) ng/mL in the Crohn’s patient group respectively, they were 321.7 ± 126.3(85–680) pg/mL and 7.6 ± 3.8(3–25.1) ng/mL in the control group respectively. Vitamin B12 and folate levels were distinctly lower in patients with Chron’s disease than those of the control group (P < .001). The intragroup analysis of the patient group revealed that low vitamin B12 levels were significantly lower in the moderate group classified according to the Crohn’s Disease Activity Index (P < .001), along with those in the L1 group with terminal/distal ileal involvement (P < .001).Vitamin B12 and folate deficiencies are quite prevalent in patients with Crohn’s disease while this condition can lead to various complications and they prove to be important risk factors associated especially with thrombosis and its complications. Patients must be regularly followed-up for vitamin B12 and folate levels to supplement them where needed.     

克罗恩病CARD15/NOD2基因突变的研究

目的 发现与中国人克罗恩病（CD）发病相关的单核苷酸多态性（SNP）及其与临床特点的关系.方法 临床资料完整的CD、溃疡性结肠炎（UC）患者及健康体检者各30例.提取人血白细胞基因组DNA,PCR扩增NOD2基因第4、8、11外显子,纯化后直接测序.结果 5例CD患者有SNP改变,其中2例为P268S,1例为R459R,2例为P268S和R459R,而在UC患者和健康人中各检测到1例R459R.所有研究对象未发现R702W、G908R及3020insC改变.CD有4例P268S,与UC和健康体检者比较差异有统计学意义（χ^2=8.037,P＜0.05）.4例P268S CD患者病变均在回肠（χ^2=9.231,P=0.01）,发病年龄小于20岁（χ^2=10.769,P＜0.01）,并发肠腔狭窄而需手术（χ^2=7.972,P＜0.01）,2例有P268S和R459R的患者病情较重,多次复发.结论 P268S可能是CARD15/NOD2基因中与中国人CD相关的SNP,与患者发病年龄、病变部位及肠腔狭窄明显相关,与患者性别及病变严重程度无关.     

Association between NOD2/CARD15 gene polymorphisms and Crohn’s disease in Chinese Zhuang patients

AIM:To assess the relationship between the P268S,JW1 and N852S polymorphisms and Crohn’s disease(CD)susceptibility in Zhuang patients in Guangxi,China.METHODS:Intestinal tissues from 102 Zhuang[48CD and 54 ulcerative colitis(UC)]and 100 Han(50 CD and 50 UC)unrelated patients with inflammatory bowel disease and 72 Zhuang and 78 Han unrelated healthy individuals were collected in the Guangxi Zhuang Autonomous Region from January 2009 to March 2013.Genomic DNA was extracted using the phenol chloroform method.The P268S,JW1 and N852S polymorphisms were amplified using polymerase chain reaction(PCR),detected by restriction fragment length polymorphism(RFLP),and verified by gene sequencing.RESULTS:Heterozygous mutation of P268S in the NOD2/CARD15 gene was detected in 10 CD cases(six Zhuang and four Han),two Han UC cases,and one Zhuang healthy control,and P268S was strongly associated with the Chinese Zhuang and Han CD populations(P=0.016 and 0.022,respectively).No homozygous mutant P268S was detected in any of the groups.No significant difference was found in P268S genotype and allele frequencies between UC and control groups(P>0.05).Patients with CD who carried P268S were likely to be≤40 years of age(P=0.040),but were not significantly different with regard to race,lesion site,complications,and other clinical features(P>0.05).Neither JW1 nor N852S polymorphisms of the NOD2/CARD15gene were found in any of the subjects(P>0.05).CONCLUSION:P268S polymorphism may be associated with CD susceptibility in the Zhuang population in the Guangxi Zhuang Autonomous Region,China.In contrast,JW1 and N852S polymorphisms may not be related to CD susceptibility in these patients.     

NOD2/CARD15基因R702W、G908R及L1007fs多态性与广西壮族人群炎症性肠病的相关性

目的:探讨我国广西壮族人群NOD2/CARD15基因R702W、G908R及L1007fs的遗传多态性与炎症性肠病的相关性.方法:分别收集2007-02/2010-10在广西地区无亲缘关系的壮族(n=70)和汉族(n=76)IBD患者及壮族(n=80)和汉族(n=84)正常对照者的肠黏膜组织.采用酚氯仿法提取各组织样本DNA,采用聚合酶链反应-限制性片段长度多态性分析(PCR-RFLP)方法对NOD2/CARD15基因R702W、G908R及L1007fs进行检测,统计基因型及等位基因频率,分析上述3个多态性位点与广西壮族人群炎症性肠病的相关性.结果:广西壮族和汉族IBD患者与正常对照者均未发现NOD2/CARD15基因R702W、G908R及L1007fs突变型基因型,所有多态性位点上的基因型全部为野生型纯合子,其基因型频率和等位基因频率分布在IBD患者和正常对照者中差异无统计学意义(P>0.05).结论:NOD2/CARD15基因R702W、G908R及L1007fs多态性与广西壮族人群炎症性肠病无明显相关性.     

Disease location, anti-Saccharomyces cerevisiae antibody, and NOD2/CARD15 genotype influence the progression of disease behavior in Crohn's disease

BACKGROUND: Crohn's disease (CD) is characterized by heterogeneity of phenotype. The Vienna classification can be used to classify CD, and recent data illustrate that behavior evolves over the course of the disease. Clinical and biological influences on disease progression remain unclear. We examined the associations of CD disease progression at diagnosis and for up to 20 years of follow-up. METHODS: Two hundred thirty-one well-characterized CD patients were studied. Demographic, clinical, and NOD2/CARD15 data were collected. Disease behavior according to the Vienna classification was assessed at diagnosis and for up to 20 years following diagnosis. RESULTS: At diagnosis, 70% of patients had inflammatory disease, 9% stricturing, and 21% penetrating. Early age at diagnosis was associated with ileocolonic and upper GI disease (p = 0.015), and positive anti-Saccharomyces cerevisiae antibody (ASCA) was associated with ileal involvement (p = 0.008). Smoking was relatively protective against colonic, rather than ileal involvement at diagnosis (p < 0.02). At 20 years, 92% had progressed to a more severe disease type. Patients who progress to a more severe disease type require more frequent surgery (p < 0.00001). Multivariate analysis found disease progression to be associated with ileal disease location (p = 0.001) and positive ASCA (p = 0.003). Variant NOD2/CARD15 alleles were protective against rapid progression of disease phenotype (p = 0.04). The presence of perianal disease was independent of intestinal penetrating disease. CONCLUSIONS: The progression of disease type in CD is associated with the need for more frequent surgery. Rapid progression is associated with ileal disease and positive ASCA, and delayed progression is associated with variant NOD2/CARD15 alleles. Consideration should be given to a separate Vienna classification for perianal disease.     

A nonsynonymous SNP in ATG16L1 predisposes to ileal Crohn's disease and is independent of CARD15 and IBD5

BACKGROUND & AIMS: A genome-wide association scan of nonsynonymous DNA polymorphisms identified association of a threonine-to-alanine substitution (T300A) in the autophagy-related 16-like gene ATG16L1 with Crohn's disease. We investigated this association in independent U.K. cohorts of Crohn's disease and ulcerative colitis. METHODS: The T300A variant (rs2241880) was genotyped in an independent sample of 727 Crohn's disease and 877 ulcerative colitis cases, and in 579 controls. We then performed an extension analysis combining these data with the U.K. data from the initial study to give a total of 1236 U.K. Crohn's disease cases and 1235 controls to estimate disease risk and test for interaction with the CARD15 and IBD5 risk loci and for association with disease subtypes. RESULTS: The association of T300A was replicated in the independent sample of 727 Crohn's disease cases (P = .001), and was strongly associated in the extended analysis of 1236 Crohn's cases (P = 2.4 x 10(-6)). The 300A/A genotype conferred a 1.65-fold risk of Crohn's disease, with a 2.2-fold risk of ileal disease. Analysis of the interaction of ATG16L1 with CARD15 and IBD5 indicated that all 3 loci contribute independently to disease risk. Homozygosity for the risk allele at all 3 loci conferred a combined risk of 20.4 (95% confidence interval: 8.71, 47.7) for Crohn's disease. The ATG16L1 risk genotype showed a modest but significant association with ulcerative colitis (P = .026). CONCLUSIONS: The association of ATG16L1 with Crohn's disease and possibly with ulcerative colitis supports a role for autophagy in the pathogenesis of inflammatory bowel disease.     

Genetic and serological markers to identify phenotypic subgroups in a Dutch Crohn’s disease population

BACKGROUND AND AIMS: Both genetic and microbial factors seem to play a pivotal role in the aetiopathogenesis of Crohn's disease. The CARD15 frameshift mutation might link host genetic factors and the indigenous microbial flora, since CARD15 expression is stimulated by peptidoglycan, thereby activating NF-kappaB. It is hypothesised that CARD15 mutation carriers have defective anti-microbial reactions, resulting in more penetrating lesions and antibody responses, which are now being used as highly specific markers for Crohn's disease. The serological marker anti-Saccharomyces cerevisiae antibody directed against cell wall oligomannosidic epitopes has high specificity for Crohn's disease. Perinuclear anti-neutrophil cytoplasmic antibodies have been found in a subgroup of Crohn's disease patients, mostly with colonic involvement. METHODS: We investigated the incidence of two CARD15 mutations (3020insC and 2722G>C), anti-S. cerevisiae antibody, and perinuclear anti-neutrophil cytoplasmic antibody in 108 (73F/35M) patients with Crohn's disease with a mean duration of disease since diagnosis of 16 (1-41) years in relation to their phenotype, according to the Vienna classification. RESULTS: The prevalence of CARD15 frameshift mutation was 21%. Of all patients, 62% were anti-S. cerevisiae antibody positive, and 9% had perinuclear anti-neutrophil cytoplasmic antibodies. The prevalence of both anti-S. cerevisiae antibodies and perinuclear anti-neutrophil cytoplasmic antibodies was higher in the mutation carriers compared to non-carriers. Remarkably, all patients with a CARD15 mutation and positive anti-S. cerevisiae antibody had ileal disease. Carriership of the mutation was significantly associated with penetrating behaviour of the disease and weakly associated with stricturing behaviour. Furthermore, anti-S. cerevisiae antibody was associated with ileal disease involvement. Finally, most perinuclear anti-neutrophil cytoplasmic antibody positive patients showed ulcerative-like behaviour of disease (by means of colonic localisation). CONCLUSIONS: Genetic and serologic markers might be useful in defining patient subgroups. This may result in a more accurate prediction of disease behaviour, prognosis and therapeutic approach.