NOD2/CARD15 , ATG16L1 and IL23R gene polymorphisms and childhood-onset of Crohn’s disease

AIM: To assess whether the polymorphisms of NOD2/ CARD15 , autophagy-related 16-like 1 (ATG16L1 ), and interleukin-23 receptor (IL23R ) genes play a more critical role in the susceptibility of childhood-onset than in adult-onset Crohn’s disease (CD). METHODS: Polymorphisms R702W, G908R, and 3020insC of NOD2/CARD15 ; rs2241880 A/G of ATG16L1 , and rs11209026 (R381Q) of IL23R gene were assessed in 110 childhood-onset CD, 364 adult-onset CD, and 539 healthy individuals. Analysis of polymorphisms R702W, G908R, ...     

NOD2,IL23R and ATG16L1 polymorphisms in Lithuanian patients with inflammatory bowel disease

AIM:To investigate the frequency of NOD2, IL23R and ATG16L1 genetic variants in a case-control panel for inflammatory bowel disease (IBD) from Lithuania.METHODS: One hundred and eighty unrelated IBD pa- tients [57 Crohn‘s disease (CD) and 123 ulcerative colitis (UC)] and 186 healthy controls were genotyped for the following known genetic susceptibility variants:NOD2-Arg702Trp (rs2066844), Gly908Arg (rs2066845) and Leu1007insC (rs2066847), as well as IL23R-Arg381Gln (rs11209026) and ATG16L1-Thr300Ala (rs2241...     

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 and ATG16L1 polymorphisms affect monocyte responses in Crohn's disease

AIM:To assess whether polymorphisms in NOD2 and ATG16L1 affect cytokine responses and mycobacterium avium subspecies paratuberculosis (MAP) survival in monocytes from Crohn's disease (CD) patients.METHODS:Monocytes were isolated from peripheral blood of CD patients of known genotype for common single nucleotide polymorphisms of NOD2 and ATG16L1.Monocytes were challenged with MAP and bacterial persistence assessed at subsequent time-points.Cytokine responses were assayed using a Milliplex multi-analyte profi...     

Role of Salmonella enterica exposure in Chilean Crohn’s disease patients

AIM:To study the association between exposure toSalmonella enterica(SE)and Crohn’s disease(CD)and its clinical implications in Chilean patients.METHODS:Ninety-four unrelated Chilean CD patients from CAREI(Active Cohort Registry of Inflammatory Bowel Disease)presenting to a single inflammatory bowel disease(IBD)unit of a University Hospital were prospectively included in this study.A complete clinical evaluation,including smoking history,was performed at the initial visit,and all the important data of clinical evolution of CD were obtained.Blood samples from these CD patients and 88 healthy sex-and agematched control subjects were analyzed for exposure to SE and for their NOD2/CARD15 gene status using the presence of anti-Salmonella lipopolysaccharide antibodies[immunoglobulin-G type(IgG)]and polymerase chain reaction(PCR),respectively.We also evaluated exposure to SE in 90 sex-and age-matched patients without CD,but with known smoking status(30 smokers,30 non-smokers,and 30 former smokers).RESULTS:CD patients comprised 54 females and 40males,aged 35.5±15.2 years at diagnosis with a mean follow-up of 9.0±6.8 years.CD was inflammatory in 59 patients(62.7%),stricturing in 24(25.5%)and penetrating in 15(15.5%).Thirty cases(31.9%)had lesions in the ileum,29(30.8%)had ileocolonic lesions,32(34.0%)had colonic lesions and 23(24.4%)had perianal disease.Sixteen CD patients(17%)were exposed to SE compared to 15(17%)of 88 healthy control subjects(P=0.8).Thirty-one CD patients(32.9%)were smokers,and 7(7.4%)were former smokers at diagnosis.In the group exposed to SE,10 of 16 patients(62.5%)were active smokers compared to 21 of 78 patients(26.9%)in the unexposed group(P=0.01).On the other hand,10 of 31 smoking patients(32%)were exposed to SE compared to 5 of 56 nonsmoking patients(9%),and one of the seven former smokers(14%)(P=0.01).In the group of 90 patients without CD,but whose smoking status was known,there was no differ-ence in exposure to SE[3 of 30 smokers(10%),5 of30 non-smokers(16%),and 5 of 30 former smokers(16%)     

中国汉族人群NOD2、IRGM、ATG16L1和STAT4基因多态性与克罗恩病的相关性研究

背景：克罗恩病（CD）的病因和发病机制尚未完全阐明,近年国外研究发现NOD2、IRGM、ATG16L1、STAT4基因突变与CD相关。目的：分析NOD2、IRGM、ATG16L1、STAT4基因多态性与中国汉族人群CD发病的相关性。方法：连续纳入2007年1月～2010年1月苏州市立医院中国汉族CD患者66例,66名健康体检者作为正常对照,以PCR联合基因测序检测4种基因相应单核苷酸多态性（SNP）位点的基因型,分析各基因型和等位基因频率。结果：CD组和正常对照组NOD2基因rs2066842位点、IRGM基因rs13361189位点、ATG16L1基因rs2241880位点和STAT4基因rs7574865位点基因型和等位基因频率分布均符合Hardy-Weinberg遗传平衡定律,两组间4种基因相应SNP位点的基因型和等位基因频率差异均无统计学意义。结论：NOD2、IRGM、ATG16L1和STAT4基因多态性与中国汉族人群CD发病不相关。     

Viruses, autophagy genes, and Crohn's disease

The etiology of the intestinal disease Crohn's disease involves genetic factors as well as ill-defined environmental agents. Several genetic variants linked to this disease are associated with autophagy, a process that is critical for proper responses to viral infections. While a role for viruses in this disease remains speculative, accumulating evidence indicate that this possibility requires serious consideration. In this review, we will examine the three-way relationship between viruses, autophagy genes, and Crohn's disease and discuss how host-pathogen interactions can mediate complex inflammatory disorders.     

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.     

Toll-like receptor 4 and NOD2／CARD15 mutations in Hungarian patients with Crohn＇s disease： Phenotype-genotype correlations

AIM: To determine common NOD2/CARD15 mutations and TLR4 D299G polymorphism in Hungarian patients with CD. METHODS: A total of 527 unrelated patients with CD (male/female: 265/262, age: 37.1 (SD 7.6) years) and 200 healthy subjects were included. DNA was screened for possible NOD2/CARD15 mutations by denaturing high-performance liquid chromatography (confirmed by direct sequencing). TLR4 D299G was tested by PCR-RFLP. RESULTS: NOD2/CARD15 mutations were found in 185 patients (35.1%) and in 33 controls (16.5%,P<0.0001). SNP8/R702W (10.8% vs 6%, P= 0.02), SNP13/3020insC (19.4% vs 5%, P<0.0001) and exon4 R703C (2.1% vs 0%, P= 0.02) mutations were more frequent in CD, while the frequency of SNP12/G908R was not increased. The frequency of TLR4 D299G was not different (CD: 9.9% vs controls: 12.0%). Variant NOD2/CARD15 allele was associated with an increased risk for CD (ORhet=1.71, 95%CI=1.12-2.6, P= 0.0001, ORtwo-risk alleles = 25.2, 95%CI =4.37- ,P<0.0001), early disease onset (carrier: 26.4 years vs non-carrier: 29.8 years, P=0.0006), ileal disease (81.9% (?) 69.5%, OR = 1.99, 95%CI = 1.29-3.08, P= 0.02, presence of NOD2/CARD15 and TLR4: 86.7% vs 64.8%), stricturing behavior (OR = 1.69,95%CI = 1.13-2.55, P= 0.026) and increased need for resection (OR=1.71, 95%CI: 1.13-2.62, P= 0.01), but not with duration, extra-intestinal manifestations, familial disease or smoking. TLR4 exhibited a modifier effect: age of onset in wt/TLR4 D299G carriers: 27.4 years vs NOD2mut/TLR D299G: 23 years (P = 0.06), in NOD2mut/wt: 26.7 years. CONCLUSION: These results confirm that variant NOD2/ CARD15 (R702W, R703C and 3020insC) alleles are associated with earlier disease onset, ileal disease, stricturing disease behavior in Hungarian CD patients. In contrast, although the frequency of TLR4 D299G polymorphism was not different from controls, NOD2/TLR4 mutation carriers tended to present at earlier age.     

Polymorphisms in interleukin-10 gene according to mutations of NOD2/CARD15 gene and relation to phenotype in Spanish patients with Crohn＇s disease

AIM:To examine the contribution of interleukin-10(IL-10)gene polymorphisms to Crohn's disease(CD)phenotype,and the possible genetic epistasis betweenIL-10 gene polymorphisms and CARD15/NOD2 genemutations.METHODS:A cohort of 205 Spanish unrelated patientswith Crohn's disease recruited from a single centerwas studied.All patients were rigorously phenotypedand followed-up for at least 3 years(mean time,12.5years).The clinical phenotype was established prior togenotyping.RESULTS:The correlat on of genotype-Viennaclassification groups showed that the ileocolonic locationwas significantly associated with the-1082G allele in theNOD2/CARD15 mutation-positive patients(RR=1.52,95%CI,1.21 to 1.91,P=0.008).The multivariate analysisdemonstrated that the IL-10 G14 microsatellite allelein the NOD2/CARD15 mutation positive patients wasassociated with two risk factors,history of appendectomy(RR=2.15,95%CI=1.1-4.30,P=0.001)and smokinghabit at diagnosis(RR=1.29,95%CI=1.04-4.3,P=0.04).CONCLUSION:In Spanish population from Madrid,inCD patients carrying at least one NOD2/CARD15 mutation,the-1082G allele is associated with ileocolonic disease and the IL-10G14 microsatellite allele is associated withprevious history of appendectomy and smoking habit atdiagnosis.These data provide further molecular evidencefor a genetic basis of the clinical heterogeneity of CD.     

Mycobacterium avium paratuberculosis and the etiology of Crohn's disease: a review of the controversy from the clinician's perspective

Mycobacterium avium paratuberculosis (MAP) is an obligate intracellular organism that has frequently been associated with Crohn's disease (CD). Because CD is a chronic inflammatory condition, many researchers have speculated that an infectious agent must be the cause of CD. MAP has often been proposed to be one such agent; however, despite considerable research, the evidence remains inconclusive. Higher levels of MAP have been found in the tissues and blood of CD patients than in controls, forming the foundation for much of the research into the role of MAP in CD and the primary argument in support of a causative role for MAP in CD. MAP is a slow-growing and fastidious organism that is difficult to grow in culture and, therefore, challenging to detect in patients. As a result, there has been variability in the results of studies attempting to detect the presence of MAP in CD patients, and considerable controversy over whether this organism has a causative role in the etiology of CD. Two main hypotheses exist with respect to the role of MAP in CD. The first is that MAP is a principal cause of CD, while the second is that MAP is more prevalent because of the immune dysfunction seen in CD but does not play a causative role. Clinicians are often faced with questions regarding the role of this organism and the need to treat it. The present article attempts to provide an overview of the controversy including the nature of the mycobacterium, the difficulty in detecting it, the use of antimycobacterial agents to treat it and the effect of immunosuppressive agents - all from a clinician's perspective. Although the role of MAP in CD remains controversial and an area of considerable research, it is currently only of academic interest because there is no clinically useful test to identify the presence of the organism, and no evidence to support the use of antibiotics to eradicate it for the treatment of CD.     

Why does Crohn's disease usually occur in terminal ileum?

Crohn's disease can affect any part of the gastrointestinal tract, but terminal ileum is the most frequent localization. The reason why Crohn's disease is primarily located in the distal part of the ileum remains unexplained.In this article it has been attempted to provide a compelling explanation why Crohn's disease usually occurs in terminal ileum. Recent data indicate that some individuals are genetically predisposed to develop ileal Crohn's disease. Two genetic alterations, the polymorphism of Caspase Associated Recruitment Domain (CARD15) and Carcinoembryonic Antigen-related Cell Adhesion Molecule 6 (CEACM6), favour the colonization of terminal ileum by entero adherent-invasive Escherichia coli (AIEC). The adhesion of these bacteria to epithelial intestinal cells depends on Carcinoembryonic Antigen-related Cell Adhesion Molecule 6 expression in ileal epithelial cells and on the reduced ileal defensins expressed in a CARD15 dependent manner. Genetic defects in Authophagy-related 16-like gene (ATG16L1) and Immunity-related Guanosine Triphospatase (IRGM) recently found in ileal CD patients lead to a reduction of bacterial killing by macrophages and consequent continuous immunological upstimulation, cytokine secretion, chronic inflammation of the ileum and tissue injury. On the basis of all these data Crohn's disease of the ileum seems to be a subset of the disease mainly genetically determined.     

Association between inflammatory bowel disease gene 5 (IBD5) and interleukin-23 receptor (IL23R) genetic polymorphisms in Malaysian patients with Crohn's disease

OBJECTIVE: This study was aimed to investigate the possible association of Crohn's disease (CD) with inflammatory bowel disease gene 5 (IBD5) IGR2198a_1 (rs11739135), IGR2096a_1 (rs12521868) and interleukin‐23 receptor (IL23R) genetic variant (rs1004819) in the Malaysian population. METHODS: Blood samples from 80 CD patients and 100 healthy controls were recruited. Genomic DNA was extracted and analyzed by polymerase chain reaction‐restriction fragment length polymorphism. RESULTS: The results revealed that there was an increased frequency of IGR2198a_1 C allele (8.8% in CD, 1.5% in controls, P < 0.05, OR 6.30, 95% CI 1.77–22.31) and IGR2096a_1 T allele (6.9% in CD, 1.5% in controls, P < 0.05, OR 4.85, 95% CI 1.33–17.69) in the CD patients as compared to the controls, suggesting the two variants were potential risk factors of CD. Both risk alleles (C and T) were highest in Indians. In contrast, no significant difference was observed for the IL23R gene variant (rs1004819) between these two groups (P = 0.941). All genotypes and alleles of this gene variant were present in equal ratios in the CD and control groups (OR 1.02, 95% CI 0.66–1.57 for T allele and OR 0.98, 95% CI 0.64–1.52 for C allele). CONCLUSIONS: There is a strong association between both IBD5 locus variants but not the IL23R gene variant with CD in the Malaysian population. The IBD5 locus variants were highest in Indians, which may explain the increased susceptibility of this particular ethnic group to the disease.     

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.     

Role of genetics in the diagnosis and prognosis of Crohn's disease

Considering the epidemiological, genetic and immunological data, we can conclude that the inflammatory bowel diseases are heterogeneous disorders of multifactorial etiology in which hereditability and environment interact to produce the disease. It is probable that patients have a genetic predisposition for the development of the disease coupled with disturbances in immunoregulation. Several genes have so far been related to the diagnosis of Crohn's disease. These genes are related to innate pattern recogni...     

Prevalence of mutations of the NOD2/CARD15 gene and relation to phenotype in Spanish patients with Crohn disease

Background: We assessed the prevalence of R702W, G908R, and L1007fs coding mutations in the NOD2/CARD15 gene and the genotype–phenotype relation in Spanish patients with Crohn disease. Methods: A cohort of 204 unrelated patients with Crohn disease and 140 healthy controls were studied. The phenotype was established before commencement of genotyping. Genotyping of the R702W, G908R, and L1007fs gene polymorphisms of NOD2/CARD15 was performed by two independent laboratories using different techniques. In the case of discordant results, specific sequencing of DNA strands was performed. Results: At least one mutation was present in 32.8% of patients compared to 10.7% in controls (OR?=?4.08, 95% CI 2.21 to 7.50). In patients with Crohn disease, the frequency of R702W, G908R, and L1007fs carriers was 13.7%, 8.3%, and 14.2%, respectively. Compound heterozygotes and homozygotes occurred in 3.4% and 2.9% of patients and in none of the controls. The correlation of genotype–Vienna classification showed a significant association with ileal disease (RR?=?1.61, 95% CI 1.21–2.15, P?=?0.001) and an inverse association with colonic localization (RR?= 0.29, 95% CI 0.11–0.80, P?=?0.007). There was a significant association between G908R carriership and previous appendectomy, surgical interventions, and stricturing behavior. A gene‐dosage effect on phenotypic characteristics was not observed. Conclusions: In a Spanish population from Madrid, mutations of the NOD2/CARD15 gene were a marker of susceptibility to Crohn disease and were associated with ileal disease. Carriers of the G908R mutation showed a stricturing disease behavior, history of appendectomy, and surgical interventions over the course of the disease.     

Mycobacterium avium subspecies paratuberculosis causes Crohn's disease in some inflammatory bowel disease patients

Crohn’s disease (CD) is a chronic inflammatory condition that plagues millions all over the world. This debilitating bowel disease can start in early childhood and continue into late adulthood. Signs and symptoms are usually many and multiple tests are often required for the diagnosis and confirmation of this disease. However, little is still understood about the cause(s) of CD. As a result, several theories have been proposed over the years. One theory in particular is that Mycobacterium avium subspecies paratuberculosis (MAP) is intimately linked to the etiology of CD. This fastidious bacterium also known to cause Johne’s disease in cattle has infected the intestines of animals for years. It is believed that due to the thick, waxy cell wall of MAP it is able to survive the process of pasteurization as well as chemical processes seen in irrigation purification systems. Subsequently meat, dairy products and water serve as key vehicles in the transmission of MAP infection to humans (from farm to fork) who have a genetic predisposition, thus leading to the development of CD. The challenges faced in culturing this bacterium from CD are many. Examples include its extreme slow growth, lack of cell wall, low abundance, and its mycobactin dependency. In this review article, data from 60 studies showing the detection and isolation of MAP by PCR and culture techniques have been reviewed. Although this review may not be 100% comprehensive of all studies, clearly the majority of the studies overwhelmingly and definitively support the role of MAP in at least 30%-50% of CD patients. It is very possible that lack of detection of MAP from some CD patients may be due to the absence of MAP role in these patients. The latter statement is conditional on utilization of methodology appropriate for detection of human MAP strains. Ultimately, stratification of CD and inflammatory bowel disease patients for the presence or absence of MAP is necessary for appropriate and effective treatment which may lead to a cure.