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CARD15/NOD2 Mutational Analysis and Genotype-Phenotype Correlation in 612 Patients with Inflammatory Bowel Disease
Abstract
CARD15/NOD2 encodes a protein involved in bacterial recognition by monocytes. Mutations in CARD15 have recently been found in patients with Crohn disease (CD), a chronic inflammatory condition of the digestive tract. Here, we report the mutational analyses of CARD15 in 453 patients with CD, including 166 sporadic and 287 familial cases, 159 patients with ulcerative colitis (UC), and 103 healthy control subjects. Of 67 sequence variations identified, 9 had an allele frequency >5% in patients with CD. Six of them were considered to be polymorphisms, and three (R702W, G908R, and 1007fs) were confirmed to be independently associated with susceptibility to CD. Also considered as potential disease-causing mutations (DCMs) were 27 rare additional mutations. The three main variants (R702W, G908R, and 1007fs) represented 32%, 18%, and 31%, respectively, of the total CD mutations, whereas the total of the 27 rare mutations represented 19% of DCMs. Altogether, 93% of the mutations were located in the distal third of the gene. No mutations were found to be associated with UC. In contrast, 50% of patients with CD carried at least one DCM, including 17% who had a double mutation. This observation confirmed the gene-dosage effect in CD. The patients with double-dose mutations were characterized by a younger age at onset (16.9 years vs. 19.8 years; P=.01), a more frequent stricturing phenotype (53% vs. 28%; P=.00003; odds ratio 2.92), and a less frequent colonic involvement (43% vs. 62%; P=.003; odds ratio 0.44) than were seen in those patients who had no mutation. The severity of the disease and extraintestinal manifestations were not different for any of the CARD15 genotypes. The proportion of familial and sporadic cases and the proportion of patients with smoking habits were similar in the groups of patients with CD with or without mutation. These findings provide tools for a DNA-based test of susceptibility and for genetic counseling in inflammatory bowel disease.
... In the European population, the most studied worldwide, approximately 35% to 40% of CD patients are carriers of at least 1 NOD2 SV, supporting the role as a genetic modifier of disease development. 39,40 This strong association has also been described in Australia, 41 Canada, 42 and the United States, 43 countries with a predominantly Anglo-Saxon population. A relevant metaanalysis showed the tight association between NOD2 and CD in Caucasians. ...
... While the analysis of Economou et al 44 indicates that this relationship exists regardless of colonic involvement, other studies, such as Ahmad et al, 24 reported a negative association between NOD2 SVs and colic disease. 39,42 A widely accepted explanation is based on the difference between immune tolerance mechanisms in the colonic and ileal gut mucosa. The higher concentration of bacteria encountered in the former may trigger the activation of NOD2-independent surveillance mechanisms. ...
Crohn’s disease (CD) is a chronic inflammatory bowel disease with a multifactorial pathogenesis involving environmental and genetic factors. Since the late 20th century, the discovery of the first susceptibility gene (NOD2, previously referred to as CARD15) for CD has paved the way for further investigations into the correlations between clinical features and genetics, and its potential impact on clinical practice has fueled the research in the last 2 decades. Recent therapeutic advancements involving novel biologic drugs and small molecules have shifted inflammatory bowel disease management from a disease-centered to a patient-centric approach. To date, the role of NOD2 has not been fully understood yet. Recent data suggest that its clinical impact may be greater than currently recognized. This review overviews the most common NOD2 variants’ role in real-life clinical practice. These genetic variants increase the risk of developing the disease and can aid in tailoring diagnosis and treatment. They are associated with the stricturing phenotype and ileal involvement and increase the risk of steroid refractoriness. In the meantime, limited and inconclusive evidence exists regarding their predictive role in response to azathioprine, biologic drugs, and small molecules. Eventually, their role in increasing the risk for surgery is evident, especially in those with the L1007fs variant. If further trials will support the initial evidence reported so far, NOD2 genetic variants will emerge as possible candidates for developing precision medicine in CD.
... NOD2 was also the first gene identified as a risk factor for ileal CD (Lesage et al. 2002). Several NOD2 variants increase the risk of CD (Kufer et al. 2006;Nelson et al. 2021;Rescigno and Nieuwenhuis 2007;Vignal et al. 2007) and the disease course of ulcerative colitis (UC) (Freire et al. 2014). ...
... ; https://doi.org/10. 1101/2022 We next asked if/how binding of GIV maybe impacted by 3 NOD2 variants (R702W, G908R and 1007fs) that represent ~80% of the mutations independently associated with susceptibility to CD (Lesage et al. 2002). ...
Sensing of pathogens by Nucleotide oligomerization domain (NOD)-like 2 receptor (NOD2) induces a protective inflammatory response that coordinates bacterial clearance. Polymorphisms in NOD2 impair bacterial clearance, leading to chronic gut inflammation in Crohn’s disease (CD) via mechanisms that remain incompletely understood. We identify GIV/Girdin (CCDC88A) as a NOD2-interactor that shapes bacterial sensing-and-signaling in macrophages. Myeloid-specific GIV depletion exacerbated and protracted infectious colitis and abolished the protective effect of muramyl dipeptide (MDP) in both chemical colitis and severe sepsis. In the presence of GIV, macrophages enhance anti-bacterial pathways downstream of NOD2, clear microbes rapidly and concomitantly suppress inflammation. GIV’s actions are mediated via its C-terminus, which directly binds the terminal leucine-rich repeat (LRR#10) of NOD2; binding is augmented by MDP and ATP, precedes receptor oligomerization, and is abolished by the 1007fs CD-risk variant which lacks LRR#10. Findings illuminate mechanisms that underlie protective NOD2 signaling and loss of function in the major 1007fs variant.
GRAPHIC ABSTRACT
In brief
This work reveals a mechanism by which macrophages use their innate immune sensor, NOD2, to protect the host against overzealous inflammation during bacterial infections, and the consequences of its loss, as occurs in the most important Crohn’s disease-risk variant.
HIGHLIGHTS
GIV is a functional and direct interactor of the terminal LRR repeat of NOD2
Mice lacking MФ GIV develop dysbiosis, protracted ileocolitis and sepsis
MDP/NOD2-dependent protective host responses require GIV
CD-risk NOD2 1007fs variant lacking the terminal LRR#10 cannot bind GIV
... In 2001, Dr. Colombel and his colleagues published a study in the journal Nature Genetics that identified a gene called NOD2 as a major susceptibility gene for Crohn's disease 22 . This discovery was a significant step forward in understanding the genetic foundation of IBD, and it provided the way for more study into the complex genetic components that contribute to the disease's genesis and progression. ...
... In 2001, Dr. Colombel and his colleagues published a study in the journal Nature Genetics that identified a gene called NOD2 as a major susceptibility gene for Crohn's disease 22 . This discovery was a significant step forward in understanding the genetic foundation of IBD, and it provided the way for more study into the complex genetic components that contribute to the disease's genesis and progression. ...
Inflammatory bowel disease (IBD) is a chronic inflammatory illness of the gastrointestinal tract that consists of Crohn's disease and ulcerative colitis. The pathogenesis of IBD is multifactorial and involves a complex interplay between genetic, environmental, and immunological factors. A comprehensive literature search was conducted using relevant databases, such as PubMed, Scopus, and Web of Science. A tool such as PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) ensures that the review is conducted systematically and transparently. Alterations in gut microbial ecology are a hallmark of IBD and include decreased microbial diversity, an expansion of potentially harmful bacteria, and a loss of beneficial bacteria. These alterations are thought to contribute to the dysregulated immune response observed in IBD. The current understanding of the pathogenesis of IBD suggests that a dysregulated immune response to commensal bacteria in the gut plays a central role in the development and progression of the disease. Alterations in gut microbial ecology are a hallmark of IBD and include decreased microbial diversity, an expansion of potentially harmful bacteria, and a loss of beneficial bacteria. These alterations in the gut microbiota are thought to contribute to the dysregulated immune response observed in IBD.
... Esta variante tiene una frecuencia alélica en gnomAD (Genome Aggregation Database) de 0,00283. Esta variante se ha descrito en la literatura en individuos con enfermedad autoinflamatoria (34,35) , y también se ha descrito con mayor frecuencia en individuos con EC y CU (36,37) . En conclusión, esta variante en el gen NOD2 pareciese que pudiese contribuir a la EC como factor de riesgo, posiblemente junto con otros factores genéticos y / o ambientales, pero son necesarios más estudios, porque hasta la fecha no hay evidencia suficiente para otorgar patogenicidad. ...
Introducción:
Se ha tratado de identificar los factores genéticos relacionados con susceptibilidad para enfermedad inflamatoria intestinal (EII), y los hallazgos actuales se inclinan por un modelo de patología complejo, sin un patrón hereditario claro.
Objetivo:
Realizar caracterización fenotípica y genotípica de pacientes con EII en población colombiana y describir su posible asociación con predisposición.
Materiales y métodos:
Serie de casos, 16 pacientes con EII por criterios clínicos y anatomopatológicos, inicio de síntomas gastrointestinales después de los 18 años. Todos tuvieron asesoramiento genético pre-test y se realizaron árboles genealógicos de mínimo tres generaciones. También, genotipificación, por medio de un panel de genes múltiples que incluía genes relacionados con EII y algunos trastornos autoinmunitarios. Finalmente, se realizó análisis genómico de variantes.
Resultados:
9 mujeres y 7 hombres, con edad media de diagnóstico de EII 35 años, y 32 años para aparición de síntomas gastrointestinales. 11/16(68,75%) requirieron terapia biológica. 10/16 (62,5%) presentaron refractariedad a terapia estándar. 3/16 (18,75%) tenían antecedentes familiares positivos de EII. 100% casos presentaron al menos un single nucleotide polymorphism relacionado con riesgo de EII en más de un gen. Los genes más relacionados con colitis ulcerosa (CU), fueron CD48, CD6, y TYK2 para CU, y CD6 e ITGAM para la enfermedad de Crohn. El gen más frecuente fue CD6. Se observó en 3/16 (18,75%) presencia de hasta 5 genes, 4 en 3/16 (18,75%), y tres en 5/16 (31,25%).
Conclusión:
En EII hay presencia de variantes genéticas con predisposición asociada, pero sin patogenicidad confirmada, y cuya sumatoria parece contribuir en su fisiopatología.
... IL-10 is crucial for mucosal homeostasis and regulation of the immune response in the colon [178,179]. Indeed, a mutation in IL-10 signaling, as seen in nucleotide-binding oligomerization domain 2 (NOD2) mutations, is linked to an increased susceptibility to IBD [175,180]. H 2 S thereby decreases inflammation by stimulation of IL-10. This is also observed in other organs, such as the brain, liver or lung [181][182][183]. ...
Hydrogen sulfide (H2S), originally known as toxic gas, has now attracted attention as one of the gasotransmitters involved in many reactions in the human body. H2S has been assumed to play a role in the pathogenesis of many chronic diseases, of which the exact pathogenesis remains unknown. One of them is inflammatory bowel disease (IBD), a chronic intestinal disease subclassified as Crohn’s disease (CD) and ulcerative colitis (UC). Any change in the amount of H2S seems to be linked to inflammation in this illness. These changes can be brought about by alterations in the microbiota, in the endogenous metabolism of H2S and in the diet. As both too little and too much H2S drive inflammation, a balanced level is needed for intestinal health. The aim of this review is to summarize the available literature published until June 2023 in order to provide an overview of the current knowledge of the connection between H2S and IBD.
... This has been shown to be important in intestinal epithelial cells at the interface with the gut microbiome [38,49]. Genetic variation in NOD2 accounts for 20% of CD genetic risk, with three common variants, R702W, G908R, and L1007fs, particularly associated with ileal CD [50]. NOD2 mutations impair NFκB and autophagic responses to intracellular bacteria, leading to the accumulation of bacteria in intestinal epithelial cells [51]. ...
Crohn’s disease (CD) is a chronic inflammatory bowel disease marked by relapsing, transmural intestinal inflammation driven by innate and adaptive immune responses. Autophagy is a multi-step process that plays a critical role in maintaining cellular homeostasis by degrading intracellular components, such as damaged organelles and invading bacteria. Dysregulation of autophagy in CD is revealed by the identification of several susceptibility genes, including ATG16L1, IRGM, NOD2, LRRK2, ULK1, ATG4, and TCF4, that are involved in autophagy. In this review, the role of altered autophagy in the mucosal innate immune response in the context of CD is discussed, with a specific focus on dendritic cells, macrophages, Paneth cells, and goblet cells. Selective autophagy, such as xenophagy, ERphagy, and mitophagy, that play crucial roles in maintaining intestinal homeostasis in these innate immune cells, are discussed. As our understanding of autophagy in CD pathogenesis evolves, the development of autophagy-targeted therapeutics may benefit subsets of patients harboring impaired autophagy.
... Limitation of the study: It is estimated that homozygous or heterozygous NOD2 mutations are prevalent in up to 40% of patients with Crohn's disease [46,47]. In the experimental setting, there is a frameshift model resulting in a gain of function with overexpression of NFkB [48] and a loss of function model using Nod2 knockout mice with complete Nod2 deficiency [49]. ...
Ileocecal resection (ICR) is frequently performed in Crohn’s disease (CD). NOD2 mutations are risk factors for CD. Nod2 knockout (ko) mice show impaired anastomotic healing after extended ICR. We further investigated the role of NOD2 after limited ICR. C57B16/J (wt) and Nod2 ko littermates underwent limited ICR including 1–2 cm terminal ileum and were randomly assigned to vehicle or MDP treatment. Bursting pressure was measured on POD 5, and the anastomosis was analyzed for matrix turn-over and granulation tissue. Wound fibroblasts from subcutaneously implanted sponges were used for comparison. The M1/M2 macrophage plasma cytokines were analyzed. Mortality was not different between groups. Bursting pressure was significantly decreased in ko mice. This was associated with less granulation tissue but was not affected by MDP. However, anastomotic leak (AL) rate tended to be lower in MDP-treated ko mice (29% vs. 11%, p = 0.07). mRNA expression of collagen-1α (col1 α), collagen-3α (col3 α), matrix metalloproteinase (mmp)2 and mmp9 was increased in ko mice, indicating increased matrix turn-over, specifically in the anastomosis. Systemic TNF-α expression was significantly lower in ko mice. Ileocolonic healing is impaired in Nod2 ko mice after limited ICR by local mechanisms maybe including local dysbiosis.
... Genetic polymorphism in NOD2 was the first found to be associated with the risk of CD development and confers the strongest genetic risk (62)(63)(64). Around 40-50% of CD patients carry at least one mutation in the NOD2/CARD15 gene (65,66). However it has heterogeneity and many studies have shown that NOD2 variants play no role in CD in East Asian populations (67)(68)(69). ...
Inflammatory bowel disease (IBD) is a group of disorders that cause chronic inflammation in the intestines, with the primary types including ulcerative colitis and Crohn's disease. The link between autophagy, a catabolic mechanism in which cells clear protein aggregates and damaged organelles, and intestinal health has been widely studied. Experimental animal studies and human clinical studies have revealed that autophagy is pivotal for intestinal homeostasis maintenance, gut ecology regulation and other aspects. However, few articles have summarized and discussed the pathways by which autophagy improves or exacerbates IBD. Here, we review how autophagy alleviates IBD through the specific genes (e.g., ATG16L1, IRGM, NOD2 and LRRK2), crosstalk of multiple phenotypes with autophagy (e.g., Interaction of autophagy with endoplasmic reticulum stress, intestinal antimicrobial defense and apoptosis) and autophagy-associated signaling pathways. Moreover, we briefly discuss the role of autophagy in colorectal cancer and current status of autophagy-based drug research for IBD. It should be emphasized that autophagy has cell-specific and environment-specific effects on the gut. One of the problems of IBD research is to understand how autophagy plays a role in intestinal tract under specific environmental factors. A better understanding of the mechanism of autophagy in the occurrence and progression of IBD will provide references for the development of therapeutic drugs and disease management for IBD in the future.
... The component of CARD15 associated with Blau syndrome susceptibility is found within the nucleotide binding site domain [102], whereas the Crohn's susceptibility is in the N-terminal leucine-rich repeat domain [103]. In a small series looking for MAP in six individual Blau tissues, including skin, synovium, and granulomas of the liver and kidney, all six tissues had the DNA of MAP [104]. ...
Clinical and histological similarities between sarcoidosis and tuberculosis have driven repeated investigations looking for a mycobacterial cause of sarcoidosis. Over 50 years ago, “anonymous mycobacteria” were suggested to have a role in the etiology of sarcoidosis. Both tuberculosis and sarcoidosis have a predilection for lung involvement, though each can be found in any area of the body. A key histopathologic feature of both sarcoidosis and tuberculosis is the granuloma—while the tuberculous caseating granuloma has an area of caseous necrosis with a cheesy consistency; the non-caseating granuloma of sarcoidosis does not have this feature. This article reviews and reiterates the complicity of the infectious agent, Mycobacterium avium subsp. paratuberculosis (MAP) as a cause of sarcoidosis. MAP is involved in a parallel story as the putative cause of Crohn’s disease, another disease featuring noncaseating granulomas. MAP is a zoonotic agent infecting ruminant animals and is found in dairy products and in environmental contamination of water and air. Despite increasing evidence tying MAP to several human diseases, there is a continued resistance to embracing its pleiotropic roles. “Who Moved My Cheese” is a simple yet powerful book that explores the ways in which individuals react to change. Extending the metaphor, the “non-cheesy” granuloma of sarcoidosis actually contains the difficult-to-detect “cheese”, MAP; MAP did not move, it was there all along.
... Blau syndrome has generated interest in medical literature because of the discovery that places its genetic defect with the same Crohn's susceptibility CARD15 gene (88,89). The Blau syndrome susceptibility component of CARD15 is found within the nucleotide binding site domain (90) while the Crohn's susceptibility is in the N-terminal leucine-rich repeat domain (91). In a small series looking for MAP in six individual Blau tissues, including skin, synovium and granulomas of the liver and kidney, all six tissues had the DNA of MAP (92). ...
Clinical and histological similarities between sarcoidosis and tuberculosis have driven repeated investigations looking for a mycobacterial cause of sarcoidosis. Over 50 years ago “anonymous mycobacteria” were suggested to have a role in the etiology of sarcoidosis. Both tuberculosis and sarcoidosis have a predilection for lung involvement though each can be found in any area of the body. A key histopathologic feature of both sarcoidosis and tuberculosis is the granuloma – while the tuberculous caseating granuloma has an area of caseous necrosis with a cheesy consistency; the non-caseating granuloma of sarcoidosis does not have this feature. This article reviews and reiterates the complicity of the infectious agent, Mycobacterium avium ss. paratuberculosis (MAP) as a cause of sarcoidosis. MAP is involved in a parallel story as the putative cause of Crohn’s disease, another disease featuring noncaseating granulomas. MAP is a zoonotic agent infecting ruminant animals and is found in dairy products and in environmental contamination of water and air. Despite increasing evidence tying MAP to several human diseases, there is a continued resistance to embracing its pleiotropic roles. "Who Moved My Cheese" is a simple yet powerful book that explores the ways in which individuals react to change. Extending the metaphor, the “non-cheesy” granuloma of sarcoidosis actually contains the difficult-to-detect “cheese”, MAP; MAP did not move, it was there all along.
... NOD2 was the first gene to be associated with CD and therefore several genotype-phenotype studies with a few hundred individuals tried to associate NOD2 variants with CD sub-phenotypes (Cuthbert et al. 2002;Lesage et al. 2002;Hampe et al. 2002;Vermeire et al. 2002;Abreu et al. 2002;Louis et al. 2003;Brant et al. 2003;Seiderer et al. 2006). Similar to the HLA region studies, results between the studies varied. ...
Inflammatory bowel disease (IBD), with Crohn’s disease and ulcerative colitis as main subtypes, is a prototypical multifactorial disease with both genetic and environmental factors involved. Genetically, IBD covers a wide spectrum from monogenic to polygenic forms. In polygenic disease, many genetic variants each contribute a small amount to disease risk. With the advent of genome-wide association studies (GWAS), it became possible to find these variants and corresponding genes, leading so far to the discovery of ca 240 loci associated with IBD. Together, these however explain only 20–25% of the heritability of IBD, leaving a large portion unaccounted for. This missing heritability might be hidden in common variants with even lower effect than the ones currently found through GWAS, but also in rare variants which can be found through large-scale sequencing studies or potentially in multiplex families. In this review, we will give an overview of the current knowledge about the genetics of non-monogenic IBD and how it differs from the monogenic form(s), and future perspectives. The history of IBD genetic studies from twin studies over linkage studies to GWAS, and finally large-scale sequencing studies and the revisiting of multiplex families will be discussed.
... CD patients carrying the most prevalent of these NOD2 variants (Arg702Trp and Gly908Arg) are also at increased risk for complications such as bowel obstruction and the need for surgery [158]. Patients carrying two SNPs have a higher incidence of stenosis compared to patients with one or two wild-type alleles [159]. The V249I polymorphism of the CX3CR1 gene is also associated with intestinal stricture, especially in smokers, which is independent of NOD2 (CARD2) mutations [160]. ...
Introduction
Intestinal fibrosis is a common complication of Inflammatory Bowel Disease (IBD) with no available drugs. The current therapeutic principle is surgical intervention as the core. Intestinal macrophages contribute to both the progression of inflammation and fibrosis. Understanding the role of macrophages in the intestinal microenvironment could bring new hope for fibrosis prevention or even reversal.
Areas covered
This article reviewed the most relevant reports on macrophage in the field of intestinal fibrosis. The authors discussed current opinions about how intestinal macrophages function and interact with surrounding mediators during inflammation resolution and fibrostenotic IBD. Based on biological mechanisms findings, authors summarized related clinical trial outcomes.
Expert opinion
The plasticity of intestinal macrophages allows them to undergo dramatic alterations in their phenotypes or functions when exposed to gastrointestinal environmental stimuli. They exhibit distinct metabolic characteristics, secrete various cytokines, express unique surface markers, and transmit different signals. Nevertheless, the specific mechanism through which the intestinal macrophages contribute to intestinal fibrosis remains unclear. It should further elucidate a novel therapeutic approach by targeting macrophages, especially distinct mechanisms in specific subgroups of macrophages involved in the progression of fibrogenesis in IBD.
... This is accompanied by SNP8 located in exon 4 (c.2104C>T, p.Arg702Trp), SNP12 in exon 8 (c.2722G>C, p.Gly908Arg), and SNP13 in exon 11 (c.3019_3020insC, p.Leu1007fs) [28]. By conducting further investigation, a recent research group found that different haplotypes of risk alleles are associated with the other onset of IBD. ...
Inflammatory bowel disease (IBD) is a chronic inflammatory gastrointestinal disease characterized by weight loss, abdominal pain, and bloody diarrhea. The number of affected patients has increased in recent years. Despite the fact that scientists have been studying the pathogenesis of IBD for many years, the specific pathogenesis pathway remains unclear. As a result, none of the therapeutic approaches can cure IBD patients completely. However, the increasing research factors associated with the incidence of IBD are reasonable. These variables can be divided into two categories: microbiome-related factors (bacteria, fungi, and viruses) and nonmicrobiome-related factors (diet, gene, host immune system, gender, and ethnicity). Surprisingly, we found that all the variables impact the gut flora in IBD patients, either directly or indirectly. Dysbiosis of the gut microbiota eventually leads to an increase in the incidence of IBD. As a result, therapeutic targets focusing on correcting dysbiosis in the gut microbiome, including using probiotics and postbiotics, could become one of the most promising IBD treatments in the future. We went through each linked factor and explained how they contribute to an increased risk of IBD. We will review some existing conventional therapies for IBD before moving on to a revolutionary therapy strategy that employs prebiotics, probiotics, and postbiotics to treat IBD based on the criteria stated. Furthermore, different persons have varying reactions to the same probiotic strain. As a result, we also provide the option of having individualized probiotic medication tailored to each IBD patient.
... Certaines mutation génétiques -sur le gène NOD2 [294,295] (gène d'expression d'un PRR) ou le gène XBP1 [296] (facteur de transcription impliqué dans la réponse au stress du réticulum endoplasmique) par exemple-sont liés à une déficience des cellules de Paneth et une forte réduction de la production de HD5 associée à une plus grande susceptibilité à des formes sévères de la MC [297]. Chez la souris, une baisse en Autophagy Related 16 like 1 (ATG16L1) conduit à la baisse de la sécrétion des défensines dans le lumen par les cellules de Paneth et le développement d'une iléite [298]. ...
Le microbiote intestinal est un écosystème de microorganismes dont les nombreuses fonctions digestives et immunitaires le rendent indispensable à la bonne santé de son hôte. Une susceptibilité génétique associée à des perturbations environnementales peut rompre cet équilibre et entrainer une dysbiose. L’inflammation chronique en résultant se traduit en différents troubles locaux et systémiques. Ainsi la dysbiose a été mise en cause dans la physiopathologie des maladies inflammatoires chroniques de l’intestin et également au développement de troubles métaboliques associés à l’obésité.Les peptides antimicrobiens sont des molécules du système immunitaire innée ayant des fonctions de contrôle de la population bactérienne au niveau de la barrière intestinale et empêchant le contact direct entre celles-ci et les cellules épithéliales.L’objectif de cette thèse est d’étudier le potentiel thérapeutique des peptides antimicrobiens dans le traitement de maladies liées à la dysbiose comme les MICI et le syndrome métabolique. Pour cela Lactococcus lactis, une bactérie lactique interagissant de manière transitoire avec la barrière épithéliale intestinale, a été utilisée comme vecteur de la molécule d’intérêt. Durant cette thèse j’ai pu établir que la cathélicidine humaine (hCAP18) et REG3A avaient un impact sur le microbiote et amélioraient les symptômes de la colite induite et de l’obésité chez le rongeur.
... Among these, three major loss-of-function mutations in NOD2 [eg. R702W, G908R, and 1007fs] have been associated with enhanced susceptibility to CD. 15 Whereas a link between NOD2 mutations and susceptibility to UC remains controversial, [16][17][18][19] the nucleotide-binding oligomerisation domain protein 2 [NOD2] is thought to play a decisive role in maintaining tolerance at the intestinal barrier. 20 NOD2 is a prototypic member of the family of intracellular Nods-like receptors that share similarities with the resistance proteins in plants. ...
Background and Aims
NOD2 has emerged as a critical player in the induction of both Th1- and Th2-responses for potentiation and polarization of antigen-dependent immunity. Loss-of-function mutations in the NOD2-encoding gene and deregulation of its downstream signaling pathway have been linked to Crohn’s disease. While it is well documented that NOD2 is capable of sensing bacterial muramyl dipeptide, it remains counterintuitive to link development of overt intestinal inflammation to a loss of bacterial-induced inflammatory response. We hypothesized that a T-helper bias could also contribute to an auto-immune like colitis different than inflammation that is fully fledged by Th1-type cells.
Methods
An edematous bowel wall with a mixed Th1/Th2 response was induced in mice by intrarectal instillation of the haptenating agent oxazolone. Survival and clinical scoring were evaluated. At several time-points after instillation, colonic damage was assessed by macroscopic and microscopic observations. To evaluate the involvement of NOD2 in immunochemical phenomena, quantitative PCR and flow cytometry analysis were performed. Bone-marrow chimera experiment allowed us to evaluate the role of hematopoietic/non-hematopoietic NOD2-expressing cells.
Results
Herein, we identified a key regulatory circuit whereby NOD2-mediated sensing of MDP by radio-resistant cells improves colitis with a mixed Th1/Th2 response that is induced by oxazolone. Genetic ablation of either Nod2 or Ripk2 precipitated oxazolone colitis that is predominantly linked to a lack of Interferon-gamma. Bone-marrow chimera experiments revealed that inactivation of Nod2 signaling in non-hematopoietic cells is causing a biased M1-M2 polarization of macrophages and a decreased frequency of splenic regulatory T cells that correlates with an impaired activation of CD4 + T cells within mesenteric lymph nodes. Mechanistically, mice were protected from oxazolone-induced colitis upon administration of MDP in an interleukin-1- and interleukin-23-dependent manner.
Conclusions
these findings indicate that Nod2 signaling may prevent pathologic conversion of T helper cells for maintenance of tissue homeostasis.
... NOD2 (HGNC:5331) has been repeatedly associated with IBD [83][84][85]. A recent study suggested that compound heterozygosity of known NOD2 risk alleles explains up to 10% of pediatric IBD in European-ancestry cases [86]. ...
Despite increased use of whole exome sequencing (WES) for the clinical analysis of rare disease, overall diagnostic yield for most disorders hovers around 30%. Previous studies of mRNA have succeeded in increasing diagnoses for clearly defined disorders of monogenic inheritance. We asked if targeted RNA sequencing could provide similar benefits for primary immunodeficiencies (PIDs) and very early-onset inflammatory bowel disease (VEOIBD), both of which are difficult to diagnose due to high heterogeneity and variable severity. We performed targeted RNA sequencing of a panel of 260 immune-related genes for a cohort of 13 patients (seven suspected PID cases and six VEOIBD) and analyzed variants, splicing, and exon usage. Exonic variants were identified in seven cases, some of which had been previously prioritized by exome sequencing. For four cases, allele specific expression or lack thereof provided additional insights into possible disease mechanisms. In addition, we identified five instances of aberrant splicing associated with four variants. Three of these variants had been previously classified as benign in ClinVar based on population frequency. Digenic or oligogenic inheritance is suggested for at least two patients. In addition to validating the use of targeted RNA sequencing, our results show that rare disease research will benefit from incorporating contributing genetic factors into the diagnostic approach.
... NOD2 is of particular interest in relation to inflammatory bowel diseases, especially Crohn's disease (CD). Although the specific etiology of CD is still unknown and likely multifactorial, meta-analyses of CD cohorts have revealed that specific mutations in NOD2 are strongly associated with CD onset, with the majority being ileal CD, as well as other inflammatory bowel diseases [32][33][34][35][36] . One characteristic of CD is a shift in the composition of the microbiome, usually with an increase in pro-inflammatory Proteobacteria species 37 . ...
Nucleotide-binding oligomerization domain containing 2 (NOD2) is a critical regulator of immune responses within the gastrointestinal tract. This innate immune receptor is expressed by several cell types, including both hematopoietic and nonhematopoietic cells within the gastrointestinal tract. Vaccination targeting the gastrointestinal mucosal immune system is especially difficult due to both physical and mechanistic barriers to reaching inductive sites. The use of lactic acid bacteria is appealing due to their ability to persist within harsh conditions, expression of selected adjuvants, and manufacturing advantages. Recombinant Lactobacillus acidophilus (rLA) has shown great promise in activating the mucosal immune response with minimal impacts on the resident microbiome. To better classify the kinetics of mucosal vaccination with rLA, we utilized mice harboring knockouts of NOD2 expression specifically within CD11c + cells. The results presented here show that NOD2 signaling in CD11c + cells is necessary for mounting a humoral immune response against exogenous antigens expressed by rLA. Additionally, disruption of NOD2 signaling in these cells results in an altered bacterial microbiome profile in both control mice and mice receiving L. acidophilus strain NCK1895 and vaccine strain LaOVA.
... Numerous mutations in the CARD15 gene are thought to modify the protective role at the ileal level by altering the production of defensins [62]. Among these mutations associated with susceptibility to Crohn's disease are R702W (arginine [R] to tryptophan [W] at position 702 of the protein), Leu1007fsInsC (insertion of the cytosine base at position 1007 of the coding DNA, responsible for a reading shift), as well as G908R (glycine [G] to arginine [R] at position 908 of the protein) [63]. Numerous other genes and genetic loci are also involved in Crohn's disease [64,65]. ...
Autoimmune diseases are caused by the overactivity of the immune system towards self-constituents. Risk factors of autoimmune diseases are multiple and include genetic, epigenetic, environmental, and psychological. Autoimmune chronic inflammatory bowel diseases, including celiac and inflammatory diseases (Crohn's disease and ulcerative colitis), constitute a significant health problem worldwide. Besides the complexity of the symptoms of these diseases, their treatments have only been palliative. Numerous investigations showed that natural phytochemicals could be promising strategies to fight against these autoimmune diseases. In this respect, plant-derived natural compounds such as flavonoids, phenolic acids, and terpenoids exhibited significant effects against three autoimmune diseases affecting the intestine, particularly bowel diseases. This review focuses on the role of natural compounds obtained from medicinal plants in modulating inflammatory auto-immune diseases of the intestine. It covers the most recent literature related to the effect of these natural compounds in the treatment and prevention of auto-immune diseases of the intestine.
... NOD2 mutations were previously linked to CD as risk factors [95]. Recently, the presence of NOD2 mutations in CD patients was described as affecting the crosstalk between fibroblasts and macrophages [46]. ...
The gastrointestinal tract is the largest mucosal surface in our body and accommodates the majority of the total lymphocyte population. Being continuously exposed to both harmless antigens and potentially threatening pathogens, the intestinal mucosa requires the integration of multiple signals for balancing immune responses. This integration is certainly supported by tissue-resident intestinal mesenchymal cells (IMCs), yet the molecular mechanisms whereby IMCs contribute to these events remain largely undefined. Recent studies using single-cell profiling technologies indicated a previously unappreciated heterogeneity of IMCs and provided further knowledge which will help to understand dynamic interactions between IMCs and hematopoietic cells of the intestinal mucosa. In this review, we focus on recent findings on the immunological functions of IMCs: On one hand, we discuss the steady-state interactions of IMCs with epithelial cells and hematopoietic cells. On the other hand, we summarize our current knowledge about the contribution of IMCs to the development of intestinal inflammatory conditions, such as infections, inflammatory bowel disease, and fibrosis. By providing a comprehensive list of cytokines and chemokines produced by IMCs under homeostatic and inflammatory conditions, we highlight the significant immunomodulatory and tissue niche forming capacities of IMCs.
... NOD2 is critical in mediating inflammatory signaling pathways in response to invading pathogens via the interaction of RIPK2 with its CARD domain 6 . Previously reported NOD2 polymorphisms associated with CD are mainly localized in the LRR domain 14,[18][19][20][21][22][23][24][25] , however two heterozygous variants (p.R38M and p.R138Q) affecting the first and second CARD domain have been suggested to alter RIPK2 recruitment and NF-κB signaling 26 . To assess whether the NOD2 variant p.E54K affects interaction with RIPK2, we conducted co-immunoprecipitation experiments using anti-FLAG beads in HEK293T cells ectopically expressing FLAG-tagged NOD2 WT or mutants along with WT RIPK2. ...
NOD2 polymorphisms may affect sensing of the bacterial muramyl dipeptide (MDP) and trigger perturbed inflammatory responses. Genetic screening of a patient with immunodeficiency and enteropathy revealed a rare homozygous missense mutation in the first CARD domain of NOD2 (ENST00000300589; c.160G > A, p.E54K). Biochemical assays confirmed impaired NOD2-dependent signaling and proinflammatory cytokine production in patient's cells and heterologous cellular models with overexpression of the NOD2 mutant. Immunoprecipitation-coupled mass spectrometry unveiled the ATPase valosin-containing protein (VCP) as novel interaction partner of wildtype NOD2, while the binding to the NOD2 variant p.E54K was abrogated. Knockdown of VCP in coloncarcinoma cells led to impaired NF-κB activity and IL8 expression upon MDP stimulation. In contrast, tunicamycin-induced ER stress resulted in increased IL8, CXCL1, and CXCL2 production in cells with knockdown of VCP, while enhanced expression of these proinflammatory molecules was abolished upon knockout of NOD2. Taken together, these data suggest that VCP-mediated inflammatory responses upon ER stress are NOD2-dependent.
... Approximately 30% of patients with CD harbor NOD2 variants. [17] There have been studies investigating the association between FMF and CD. Some reported a higher prevalence of CD among patients with FMF, and others found an increased rate of FMF among CD patients. ...
Objectives
Yao syndrome (YAOS, OMIM 617321) was formerly designated as nucleotide-binding oligomerization domain-containing protein-2 (NOD2)-associated autoinflammatory disease (NAID). This disorder shares similar clinical phenotypes with hereditary periodic fever syndromes (HPFS). This study aimed to compare YAOS with familial Mediterranean fever (FMF).
Methods
In this retrospective study, electronic medical records of a case series of YAOS were reviewed and data were analyzed. All patients underwent genetic testing for periodic fever syndrome 6-gene panel.
Results
A total of 6 cases were presented. These patients were initially thought to have MEditerranean FeVer (MEFV)-negative FMF and received treatment with colchicine. They were eventually diagnosed with YAOS. The differences between these diseases were illustrated. In addition, both MEFV and NOD2 mutations were detected in some patients and family members. Patients with carriage of both gene mutations may present with heterogeneous disease expression. A close correlation between phenotypes and genotypes is needed to make a diagnosis.
Conclusions
YAOS may mimic FMF. Molecular analysis should cover NOD2 whole gene sequencing to help distinguish these diseases. Both NOD2 and MEFV mutations may contribute to disease expression in an individual.
... At least one of the three of the NOD2 gene polymorphisms was expressed in nearly 30% of European ancestry patients. NOD2 carriers are more probable to have an ileal complications and involvement related to fibrostenosis and require intestinal trajection than non-carriers (Lesage et al. 2002). A higher risk of Crohn's disease is conferred by heterozygosity for a polymorphism (by a factor of 1. . ...
Background
Perianal fistulas are painful ulcers or sinus tracts that disproportionately affect German shepherd dogs and are proposed as a spontaneous animal model of fistulising Crohn's disease.
Objectives
To characterise the rectal and cutaneous microbiota in German shepherd dogs with perianal fistulas and to investigate longitudinal shifts with lesion resolution during immunomodulatory therapy.
Animals
Eleven German shepherd dogs with perianal fistulas and 15 healthy German shepherd dogs.
Materials and Methods
Affected dogs were evaluated and swabbed at three visits, 30 days apart, while undergoing treatment with ciclosporin and ketoconazole. Healthy German shepherd dogs were contemporaneously sampled. Sites included the rectum, perianal skin and axilla. The microbiome was evaluated following sequencing of the V4 hypervariable region of the 16S ribosomal RNA (rRNA) gene.
Results
Alpha diversity was not significantly different between healthy and affected dogs at each of the three body sites ( p > 0.5), yet rectal and perianal beta diversities from affected dogs differed significantly from those of healthy dogs at Day 0 ( p = 0.004). Rectal and perianal relative abundance of Prevotella spp. increased and perianal Staphylococcus spp. relative abundance decreased in affected dogs over time, coincident with lesion resolution.
Conclusions and Clinical Relevance
Changes in lesional cutaneous and rectal microbiota occur in German shepherd dogs with perianal fistulas and shift over time with lesion resolution during immunomodulatory therapy. Further investigations of the role of cutaneous and enteric microbiota in the pathogenesis of perianal fistulas, and whether manipulation of microbial populations may ameliorate disease, are needed.
Rare and complex rheumatic diseases (RDs) present with immense clinical variability inherent to all immunological diseases. In addition to systemic and organ‐specific inflammation, patients may display features of immunodeficiency or allergy, and as such, they may represent major diagnostic and therapeutic challenges. The person's genetic architecture has been a well‐established risk factor for RDs, albeit to variable degrees. Patients with early‐onset diseases and/or positive family history have a strong genetic component, while patients with late‐onset RDs demonstrate a more complex interplay of genetic and environmental risk factors. Overall, the genetic studies in RDs have been instrumental to our understanding of innate and adaptive immunity in human health and disease. The elucidation of the molecular causes underlying rare diseases has played a major role in the identification of genes that are critical in the regulation of inflammatory responses. In addition, studies of patients with rare disorders may help determine the mechanisms of more complex autoimmune diseases by identifying variants with small effect sizes in the same genes. In contrast, studies of common RDs are carried out in cohorts of patients with well‐established phenotypes and ancestry‐matched controls, and they aim to discover disease‐related pathways that can inform the development of novel targeted therapies. Knowing the genetic cause of a disease has helped patients and families understand the disease progression and outcome. Here, we discuss the current understanding of genetic heritability and challenges in the diagnosis of RDs and how this field may develop in the future.
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Objectives
Crohn disease is a chronic inflammatory bowel disorder that is caused by environmental and genetic factors. Mutations in the CARD15 gene have been recently identified to be associated with the disease. Until now no genetic study has focused directly on a pediatric population.
Methods
The authors sequenced all 12 exons of the CARD15 gene in 55 pediatric patients with Crohn disease from Saxony. Their average age at onset was 11.2 years (1–17.5 years). The authors also evaluated the genotype‐phenotype relationship in the patients.
Results
Fourteen different polymorphic and/or disease‐related nucleotide alterations have been identified in the patients. Sixty‐five percent of their genomic DNA samples harbored at least one of six mutations within the CARD15 gene, which previously has been identified as being associated with Crohn disease. The authors found that the cytosine insertion mutation 3020insC was significantly more common in their pediatric population than in patients with Crohn disease (26% versus 11% of the alleles) whose results were reported in the literature. The genotype–phenotype analysis showed that the authors' patients with at least one of the six CARD15 disease‐associated mutations had a high risk of inflammation located in the terminal ileum and ascending colon. In 10 of 19 patients with two mutations, intestinal resection surgery was necessary because of stricturing.
Conclusions
In the authors' pediatric patients, the genetic influence on Crohn disease was more pronounced than that reported in any other study, and it strongly affected the clinical phenotype.
The intestine is an important immune organ consisting of a complex cellular network, secreted peptides and proteins and other host defenses. Innate immunity plays a central role in intestinal immune defense against invading pathogens. It also serves as a bridge to the activation of the adaptive immune system. Pattern recognition molecules of microorganisms are an essential component for identifying invading pathogens. Toll‐like receptors (TLRs), CARD15/NOD2 and scavenger receptors all serve as the pattern recognition receptors in the innate immune defense system. Secreted bactericidal peptides or defensins produced by the intestinal epithelia represent another crucial element of innate mucosal immune defense. Mutations in pattern recognition receptors and dysfunction of secretory bactericidal peptides may impair host immune defenses leading to an invasion of pathogens resulting in chronic inflammation of the gut. This review updates our current understanding of innate immunity of the gastrointestinal tract.
Inflammatory bowel disease (IBD) is a collective term for two diseases: ulcerative colitis (UC) and Crohn’s disease (CD). There are many factors, e.g., genetic, environmental and immunological, that increase the likelihood of these diseases. Indicators of IBDs include extracellular matrix metalloproteinases (MMPs). The aim of this review is to present data on the role of selected cytokines and metalloproteinases in IBD. In recent years, more and more transcriptomic studies are emerging. These studies are improving the characterization of the cytokine microenvironment inside inflamed tissue. It is observed that the levels of several cytokines are consistently increased in inflamed tissue in IBD, both in UC and CD. This review shows that MMPs play a major role in the pathology of inflammatory processes, cancer, and IBD. IBD-associated inflammation is associated with increased expression of MMPs and reduced ability of tissue inhibitors of metalloproteinases (TIMPs) to inhibit their action. In IBD patients in tissues that are inflamed, MMPs are produced in excess and TIMP activity is not sufficient to block MMPs. This review is based on our personal selection of the literature that was retrieved by a selective search in PubMed using the terms “Inflammatory bowel disease” and “pathogenesis of Inflammatory bowel diseases” that includes systematic reviews, meta-analyses, and clinical trials. The involvement of the immune system in the pathophysiology of IBD is reviewed in terms of the role of the cytokines and metalloproteinases involved.
Background
Crohn’s disease (CD) is a complex and poorly understood myeloid-mediated disorder. Genetic variants with loss of function in the NOD2 gene confer an increased susceptibility to ileal CD. While Nod2 in myeloid cells may confer protection against T-cell mediated ileopathy, it remains unclear whether it may promote resolution of the inflamed colon. In this study, we evaluated the function of Nod2 in myeloid cells in a model of acute colitis and colitis-associated colon cancer (CAC).
Methods
To ablate Nod2 specifically within the myeloid compartment, we generated LysM Cre/+ ;Nod2 fl/fl mice. The role of NOD2 was studied in a setting of Dextran Sodium Sulfate (DSS)-induced colitis and in azoxymethane (AOM)/DSS model. Clinical parameters were quantified by colonoscopy, histological, flow cytometry, and qRT-PCR analysis.
Results
Upon DSS colitis model, LysM Cre/+ ;Nod2 fl/fl mice lost less weight than control littermates and had less severe damage to the colonic epithelium. In the AOM/DSS model, endoscopic monitoring of tumor progression revealed a lowered number of adenomas within the colon of LysM Cre/+ ;Nod2 fl/fl mice, associated with less expression of Tgfb . Mechanistically, lysozyme M was required for the improved disease severity in mice with a defect of NOD2 in myeloid cells.
Conclusion
Our results indicate that loss of Nod2 signaling in myeloid cells aids in the tissue repair of the inflamed large intestine through lysozyme secretion by myeloid cells. These results may pave the way to design new therapeutics to limit the inflammatory and tumorigenic functions of NOD2.
Mycobacteria are responsible for several human and animal diseases. NOD2 is a pattern recognition receptor that has an important role in mycobacterial recognition. However, the mechanisms by which mutations in NOD2 alter the course of mycobacterial infection remain unclear. Herein, we aimed to review the totality of studies directly addressing the relationship between NOD2 and mycobacteria as a foundation for moving the field forward. NOD2 was linked to mycobacterial infection at 3 levels: (1) genetic, through association with mycobacterial diseases of humans; (2) chemical, through the distinct NOD2 ligand in the mycobacterial cell wall; and (3) immunologic, through heightened NOD2 signaling caused by the unique modification of the NOD2 ligand. The immune response to mycobacteria is shaped by NOD2 signaling, responsible for NF-κB and MAPK activation, and the production of various immune effectors like cytokines and nitric oxide, with some evidence linking this to bacteriologic control. Absence of NOD2 during mycobacterial infection of mice can be detrimental, but the mechanism remains unknown. Conversely, the success of immunization with mycobacteria has been linked to NOD2 signaling and NOD2 has been targeted as an avenue of immunotherapy for diseases even beyond mycobacteria. The mycobacteria-NOD2 interaction remains an important area of study, which may shed light on immune mechanisms in disease.
NOD2/CARD15 was the first susceptibility gene recognized for adult-onset Crohn's (or Crohn) disease (CD). Recessive inheritance of NOD2 polymorphisms has been implicated as a mechanistic driver of pediatric-onset CD. In patients with Very Early-Onset Inflammatory Bowel Disease (VEO-IBD), however, the clinical relevance of NOD2 polymorphisms has not been fully established. Ten VEO-IBD patients with NOD2 polymorphisms (NOD2+) were compared to 16 VEO-IBD patients without genetic variants in NOD2 or any other VEO-IBD susceptibility genes (NOD2-). The majority of NOD2+ patients exhibited a CD-like phenotype (90%), linear growth impairment (90%), and arthropathy (60%), all of which were significantly more common than in the NOD2- group (p=0.037, p=0.004, p=0.026, respectively). We propose that the presence of NOD2 polymorphisms in patients with VEO-IBD might confer a CD-like phenotype, linear growth impairment, and arthropathy. These findings should be validated in larger cohorts and may guide precision medicine for patients with VEO-IBD in the future.
Macrophages are essential for the maintenance of intestinal homeostasis, yet appear to be drivers of inflammation in the context of inflammatory bowel disease (IBD). How these peacekeepers become powerful aggressors in IBD is still unclear, but technological advances have revolutionized our understanding of many facets of their biology. In this Review, we discuss the progress made in understanding the heterogeneity of intestinal macrophages, the functions they perform in gut health and how the environment and origin can control the differentiation and longevity of these cells. We describe how these processes might change in the context of chronic inflammation and how aberrant macrophage behaviour contributes to IBD pathology, and discuss how therapeutic approaches might target dysregulated macrophages to dampen inflammation and promote mucosal healing. Finally, we set out key areas in the field of intestinal macrophage biology for which further investigation is warranted.
Recently performed genome-wide association studies (GWAS) contributed to a better understanding of the etiology of inflammatory bowel disease (IBD). While over 240 genetic associations with IBD have since been identified, functional follow-up studies are still in their infancy with the overall pathogenesis of IBD remaining unsolved. For example, a functional understanding of the genetic association between the human leukocyte antigen (HLA) region and ulcerative colitis (UC) — one of the main subtypes of IBD — is still lacking. Here, we analyzed whether an autoimmune reaction involving the HLA class II proteins HLA-DQ and DR, both being strongly associated with UC, could be a disease trigger or driver. To this end, genotype data derived from whole exome sequencing and genome-wide SNP array data of 863 German UC patients as well as 4,185 healthy controls were analyzed. Association analyses identified novel variants in the NOD2 and SNX20 genes to be linked with UC and confirmed known HLA allele associations. Employing the genetic data, we generated patient-specific self-immunopeptidomes and in silico predicted HLA-peptide binding. Peptidome-wide association analyses of peptide binding preferences across cases and controls in a set of candidate proteins, yielded significant associations with 234 specific peptides. Interestingly, none of those peptides showed a differential presence in case and control samples, indicating an interaction effect between peptide presence and individual HLA genotype. The disease-associated candidate peptides predicted to be presented by risk HLA proteins contained predominantly aromatic amino acids. In contrast, protective HLA proteins were predicted to bind peptides enriched in acidic amino acids. In summary, we present a proof-of-concept immunogenetic analysis workflow that contributes to a better understanding of the role of the HLA in UC.
Receptor-interacting serine/threonine kinase 2 (RIPK2) is a vital immunomodulator that plays critical roles in nucleotide-binding oligomerization domain 1 (NOD1), NOD2, and Toll-like receptors (TLRs) signaling. Stimulated NOD1 and NOD2 interact with RIPK2 and lead to the activation of nuclear factor kappa B (NF-κB) and mitogen-activated protein kinases (MAPK), followed by the production of pro-inflammatory cytokines such as TNF-α, IL-6, and IL-12/23. Defects in NOD/RIPK2 signaling are associated with numerous inflammatory diseases, including asthma, sarcoidosis, inflammatory bowel disease (Crohn's disease and ulcerative colitis), multiple sclerosis, and Blau syndrome. As RIPK2 is a crucial element of innate immunity, small molecules regulating RIPK2 functions are attractive to establish novel immunotherapies. The increased interest in developing RIPK2 inhibitors has led to the clinical investigations of novel drug candidates. In this review, we attempt to summarize recent advances in the development of RIPK2 inhibitors and degraders.
The inflammatory bowel diseases, Crohn disease and ulcerative colitis, are caused by the immune system’s dysregulated response to gut flora and environmental exposures in genetically susceptible individuals. The last 20 years have shown great progress in understanding the basis of this genetic susceptibility. The first efforts involved epidemiology and family studies to assess the hereditary contribution to IBD. Later, studies utilizing sib-pair linkage analysis revealed one of the first replicable associations in IBD, indeed, in complex mode-of-inheritance diseases generally, the NOD2 polymorphisms. In 2006, the introduction of genome-wide association studies (GWAS) brought a new model for identifying genomic loci conferring a more modest risk of IBD. Through the aggregation of several GWAS datasets in meta-analysis, at least 240 loci have been identified. Next-generation sequencing (NGS) technology was introduced in the last decade and successfully identified multiple new risk variants, in previously identified GWAS genes. Sequencing has also been successful in identifying monogenic defects in very-early-onset (VEO) IBD cases as well as in some other specific pediatric populations with IBD or its genetic mimics. In this chapter, we will review some genetic epidemiology, specific genes identified, new approaches to identifying loci, and genotype-phenotype correlations.
Aims:
We propose a unified computational framework, PheGe-Net, to bridge phenotypes and genotypes.
Background:
Genotype is the genetic makeup of a cell, an organism, or an individual, usually regarding a specific characteristic under consideration. Phenotype can be regarded as the macroscopic description of an organism, while genotype is its microscopic expression.
Objective:
Identifying phenotype-genotype associations is the primary step in explaining the pathogenesis of complex human diseases. It is also of key importance for the development of genomic medicine, sometimes known as personalized medicine, which is a way to customize medical care to an individual's unique genetic makeup.
Methods:
PheGe-Net utilizes a phenotype similarity network, a genotype similarity network, and known phenotype-genotype associations to explore the potential associations among other unlinked phenotypes and genotypes. As by-products, PheGe-Net can also discover the phenotype and genotype groups, such that the phenotypes or genotypes within the same group are highly correlated with each other.
Results:
We validate the effectiveness of PheGe-Net on a real-world data set; our method outperformed the second-best one by around 3% on Accuracy and NMI when clustering the phenotype/genotype; it also successfully detected phenotype-genotype associations, for example, the association for obesity (OMIM id: 601665) was analyzed, and among the top ten scored genes, two known ones were assigned with scores more than 0.75, and other eight predicted ones are also explainable.
Conclusion:
Our method can reveal potential phenotype clusters and genotype clusters and their unknown associations through a variety of phenotype similarities, genotype similarities, and known phenotype-genotype associations.
Inflammatory bowel diseases (IBDs) result from dysregulated immune responses to environmental and microbial triggers in genetically susceptible hosts. Many clinical observations and animal studies support the role of the microbiome in the pathogenesis of IBD. Restoration of the fecal stream leads to postoperative Crohn's recurrence, while diversion can treat active inflammation. Antibiotics can be effective in prevention of postoperative Crohn's recurrence and in pouch inflammation. Several gene mutations associated with Crohn's risk lead to functional changes in microbial sensing and handling. However, the evidence linking the microbiome to the IBD is largely correlative, given the difficulty in studying the microbiome before disease occurs. Attempts to modify the microbial triggers of inflammation have had modest success to date. Exclusive enteral nutrition can treat Crohn's inflammation though no whole food diet to date has been shown to treat inflammation. Manipulation of the microbiome through fecal microbiota transplant and probiotics have had limited success. Further focus on early changes in the microbiome and functional consequences of microbial changes through the study of metabolomics are needed to help advance the field.
There is great heterogeneity among inflammatory bowel disease (IBD) patients in terms of pathogenesis, clinical manifestation, response to treatment, and prognosis, which requires the individualized and precision management of patients. Many studies have focused on prediction biomarkers and models for assessing IBD disease type, activity, severity, and prognosis. During the era of biologics, how to predict the response and side effects of patients to different treatments and how to quickly recognize the loss of response have also become important topics. Multiomics is a promising area for investigating the complex network of IBD pathogenesis. Integrating numerous amounts of data requires the use of artificial intelligence.
Background:
Family history increases the risk for inflammatory bowel diseases (IBDs). However, data on differences in phenotypic characteristics among patients with a strong family history of IBD are scarce and controversial. The aim of the study was to compare the phenotypic features of IBD patients with four or more affected first-degree relatives with sporadic cases of IBD.
Methods:
Patients with familial and sporadic IBD were identified from the institutional IBD database. IBD patients from families with at least four first-degree affected relatives were selected for analysis and were compared to non-matched sporadic cases with IBD chosen randomly. Comparison for type of IBD (Crohn's disease (CD) vs. ulcerative colitis (UC)), age at onset as well as for disease extent, behavior, extraintestinal manifestations and indicators of severe disease were analyzed.
Results:
Thirty-five patients with familial IBD (28 CD, seven UC) were compared to 88 sporadic IBD patients (61 CD, 24 UC and three IBDU). Disease duration was 10.3 ± 8.2 in the familial and 8.0 ± 7.2 years in the sporadic cases, p=.13. The familial cases were younger at diagnosis (19.3 ± 8.6 vs. 25.7 ± 11.8, p=.004). Patients with familial compared to sporadic IBD were significantly more likely to require steroid treatment (80% vs. 54.5%, p=.009), biological treatment (94.3%, vs. 63.6%, p<.001) or surgery (25.7%, vs. 11.4%, p=.048).
Conclusions:
IBD with a very strong positive family history is associated with younger age at onset and a more adverse IBD phenotype compared to sporadic IBD.
p>In this thesis I have undertaken two separate but related studies looking at the control and the source of MMP-3 in IBD tissues.
Thalidomide is an immunomodulatory drug that inhibits TNFα production by peripheral blood mononuclear cells (PBMC) and has been used in the treatment of resistant CD. However its mode of action is unknown and its clinical effectiveness remains unclear. Recently, experimental thalidomide derivatives have been developed that are reportedly more potent inhibitors of TNFα, though they have not been tested in IBD. In this study I investigated the effect of thalidomide and derivatives on lamina propria mononuclear cell (LPMC) TNFα and MMP-3 production. Though thalidomide inhibits PBMC TNFα production, it does not inhibit LPMC TNFα and MMP-3 production. In contrast, the thalidomide derivative CC-10004 inhibits production of both PBMC and LPMC TNFα, and downregulates LPMC MMP-3 production form subjects with IBD. Accordingly, CC-10004 may be a new and effective therapy for the treatment of IBD.
One of the most striking features in IBD is the vast influx of IgG plasma cells into the diseased mucosa. However these cells are difficult to isolate, and consequently little is known about the immune mediators they produce. In this study I have developed a novel method of isolating a pure population of functionally active plasma cell form the gut. I then showed that IgG plasma cells from patients with IBD are long-lived in-vitro and produce MMP-3. Depleting plasma cells may therefore represent a novel strategy to help treat IBD.</p
p>The aims of this study were to investigate MMP-3 production between individuals with different MMP-3 5A/6A genotypes: and to compare the association of the MMP-3 5A/6A genotype with susceptibility to CD and UC.
Myofibroblast cell lines were isolated from CD, UC patients and control subjects. The phenotype of these cells lines were characterised with immunohistochemistry. Cell lines were stimulated with TNF-α or IL-1β and the production of MMP-3 was measured by western blotting and ELISA.
Cell lines from CD, UC and control patients were myofibroblast-enriched populations of cells. Cell lines responded to stimulation with TNF-α by a dose-dependent rise in MMP-3 production. The 5A/6A polymorphism was analysed in 468 German sporadic inflammatory disease trios and 270 British and German multiplex IBD families using the transmission disequilibrium test (TDT). There was an overtransmission of the 5A allele to affected offspring (p=0.0012). There as an interaction between the MMP-3 gene 5A/6A polymorphism and the Caspase Activating Recruitment Domain (CARD) 15 gene, a well established gene for CD, such that overtransmission of the 5A allele was a significant in CARD15 carriers (p=0.0054) but not in non-carriers. In the CARD15 carriers, overtransmission of the 5A allele was associated with stenosis (p=0.0027), fistulising disease (p=0.007), previous surgical resection (p=0.0023), disease of the ileum (p=0.0001), and disease of the right colon (p=0.0015) in CD. The relationship of functional polymorphisms in promoters of the MMP-1,-3,-9 and -12 genes with IBD was also investigated in children and adults, and age-matched controls. No significant association was found between these polymorphisms and CD or UC, though the power of these studies was reduced by small sample numbers. In conclusion, the MMP-3 5A/6A polymorphism appears to be a genetic factor in CD.</p
Despite the increasing knowledge with regard to IBD (inflammatory bowel disease), including ulcerative colitis (UC) and Crohn’s disease (CD), the etiology of these conditions is still not fully understood. Apart from immunological, environmental and nutritional factors, which have already been well documented, it is worthwhile to look at the possible impact of genetic factors, as well as the composition of the microbiota in patients suffering from IBD. New technologies in biochemistry allow to obtain information that can add to the current state of knowledge in IBD etiology.
Enfermedad Inflamatoria Intestinal y tumores de piel
Les maladies inflammatoires chroniques de l’intestin (MICI) sont des pathologies multifactorielles complexes d’étiologie inconnue. Différentes mutations génétiques, l’exposition à des facteurs environnementaux ou une perte d’homéostasie du microbiote intestinal sont impliqués en proportions variables dans la perte de la fonction de barrière de la muqueuse, son invasion par les microorganismes intestinaux et finalement, le déclenchement d’une réponse inflammatoire excessive et chronique provoquant les lésions caractéristiques de ces pathologies. Différents composants du système immunitaire muqueux comme les cellules épithéliales intestinales, les cellules du système immunitaire inné et adaptatif et les médiateurs de l’inflammation sont impliqués dans la pathogenèse des MICI. D’autres mécanismes cellulaires comme des carences nutritionnelles, l’immuno-récepteur TREM-1 ainsi que l’autophagie amplifient l’inflammation intestinale et accentuent la sévérité de ces pathologies. Cette revue présente les différents mécanismes impliqués dans la physiopathologie des MICI en comparant les muqueuses intestinales saines et pathologiques.
Chronic inflammatory bowel disease (IBD) presents as two major clinical forms, Crohn's disease (CD) and ulcerative colitis (UC). Genetic epidemiological studies and animal models suggest that inherited factors play significant roles in the susceptibility to both forms of IBD. From four genome‐wide scans, putative susceptibility loci on chromosome 16 (IBD1 for CD), and on chromosomes 1, 3, 4, 6, 7, 10, and 12 for IBD, have been identified. Several other groups, including ours, have confirmed linkage to the loci on chromosomes 12 and 16. The aim of this study is to identify other potential susceptibility loci for CD with a genome‐wide search approach. In our sample of 222 individuals from 46 families (20 Jewish and 26 non‐Jewish), with a total of 65 sibpairs diagnosed with CD, we observed a novel locus with suggestive linkage [multipoint logarithm of the odds score (Mlod) >2] at chromosome 14q11.2 (Mlod = 2.8, p = 0.0002). In addition, suggestive linkage was observed in our Jewish families at chromosome 17q21‐q23 (Mlod = 2.1, p = 0.01) and chromosome 5q33‐q35 (Mlod = 2.2, p = 0.0003). The syntenic regions of the latter locus are mapped within two putative loci on mouse chromosomes 11 and 18, which were identified in a mouse IBD model induced by dextran sulfate sodium (29). Our preliminary results provide potential evidence for several susceptibility loci contributing to the risk of CD. The observation of man‐mouse synteny may accelerate the identification of CD susceptibility gene(s) on human chromosome 5.
The idiopathic inflammatory bowel diseases, Crohn’s disease (CD) and ulcerative colitis (UC), are chronic, frequently disabling
diseases of the intestines. Segregation analyses, twin concordance, and ethnic differences in familial risks have established
that CD and UC are complex, non-Mendelian, related genetic disorders. We performed a genome-wide screen using 377 autosomal
markers, on 297 CD, UC, or mixed relative pairs from 174 families, 37% Ashkenazim. We observed evidence for linkage at 3q
for all families (multipoint logarithm of the odds score (MLod) = 2.29, P = 5.7 × 10−4), with greatest significance for non-Ashkenazim Caucasians (MLod = 3.39, P = 3.92 × 10−5), and at chromosome 1p (MLod = 2.65, P = 2.4 × 10−4) for all families. In a limited subset of mixed families (containing one member with CD and another with UC), evidence for
linkage was observed on chromosome 4q (MLod = 2.76, P = 1.9 × 10−4), especially among Ashkenazim. There was confirmatory evidence for a CD locus, overlapping IBD1, in the pericentromeric region of chromosome 16 (MLod = 1.69, P = 2.6 × 10−3), particularly among Ashkenazim (MLod = 1.51, P = 7.8 × 10−3); however, positive MLod scores were observed over a very broad region of chromosome 16. Furthermore, evidence for epistasis
between IBD1 and chromosome 1p was observed. Thirteen additional loci demonstrated nominal (MLod > 1.0, P < 0.016) evidence for linkage. This screen provides strong evidence that there are several major susceptibility loci contributing
to the genetic risk for CD and UC.
Apaf-1 and Nod1 are members of a protein family, each of which contains a caspase recruitment domain (CARD) linked to a nucleotide-binding
domain, which regulate apoptosis and/or NF-κB activation. Nod2, a third member of the family, was identified. Nod2 is composed
of two N-terminal CARDs, a nucleotide-binding domain, and multiple C-terminal leucine-rich repeats. Although Nod1 and Apaf-1
were broadly expressed in tissues, the expression of Nod2 was highly restricted to monocytes. Nod2 induced nuclear factor
κB (NF-κB) activation, which required IKKγ and was inhibited by dominant negative mutants of IκBα, IKKα, IKKβ, and IKKγ. Nod2
interacted with the serine-threonine kinase RICK via a homophilic CARD-CARD interaction. Furthermore, NF-κB activity induced
by Nod2 correlated with its ability to interact with RICK and was specifically inhibited by a truncated mutant form of RICK
containing its CARD. The identification of Nod2 defines a subfamily of Apaf-1-like proteins that function through RICK to
activate a NF-κB signaling pathway.
Familial aggregation argues for genetic susceptibility to Crohn's disease. The aim of this study was to compare the age of onset and the clinical features of Crohn's disease between patients with familial disease and those with sporadic disease and investigate the concordance for disease location and type among relatives with Crohn's disease.
Seventy-two families with 2 (n = 55), 3 (n = 8), 4 (n = 6), and 5 or more (n = 3) affected first-degree relatives were selected for the study. A population of 1377 patients with sporadic nonfamilial Crohn's disease was used for comparison.
Clinical data were obtained from 176 patients with familial Crohn's disease (79 men and 97 women). Median age at onset was younger in familial Crohn's disease than in sporadic cases: 22 vs. 26.5 years (P < 0.01). In familial cases, fewer patients had exclusively colonic involvement and more patients had both small bowel and colonic involvement. Among relatives of families with 2 affected members, 56% were concordant for disease location and 49% for disease type. These percentages reached 83% and 76%, respectively, within families with more than 2 affected members.
Patients with familial Crohn's disease are characterized by an early age at onset with more extensive disease and may represent a homogeneous clinical subgroup with a particularly strong genetic influence.
Denaturing high performance liquid chromatography (DHPLC) has been described recently as a method for screening DNA samples
for single nucleotide polymorphisms and inherited mutations. Thirty-eight DNAs, 22 of which were heterozygous for previously
characterized rearranged transforming gene (RET) or cystic fibrosis transmembrane conductance regulator gene (CFTR) mutations or polymorphisms, were examined using DHPLC analysis to assess the accuracy of this scanning method. Ninety-one
per cent (20/22) of the PCR amplicons from specimens with heterozygous RET or CFTR sequence showed elution profiles distinct from corresponding homozygous normal patterns; whether the profiles for two amplicons
containing heterozygous RET sequence were distinct from homozygous cases was equivocal. To investigate the usefulness of this method for detecting mutations
in tumor DNAs, each of the phosphatase and tensin homologue deleted on chromosome ten gene (PTEN) exons were examined for mutations in 63 malignant gliomas. Seventeen PTEN PCR products from this series of brain tumors showed elution profiles indicating sample heterozygosity and in each instance
conventional sequencing confirmed the presence of a mutation. PTEN amplicons containing exons 1, 3 and 5 were sequenced for each of the 63 tumor DNAs to determine whether any mutations may
have escaped DHPLC detection, and this analysis identified one such alteration in addition to the eight mutations that DHPLC
had revealed. In total, DHPLC identified 37 of 40 (92.5%) PCR products containing defined sequence variation and no alterations
were indicated among 196 amplicons containing homozygous normal sequence.
Inflammatory bowel diseases (IBD) are complex disorders. While the exact etiology of these diseases remains unknown, recent progress in the epidemiology and genetics of IBD has clearly demonstrated both environmental and genetic factors to play a role in the development of the disease, and it is expected that some risk factors are common for both Crohn's disease (CD) and ulcerative colitis (UC). The environmental factor(s) are associated with the Western way of life in the second half of the twentieth century. Cigarette smoking is presently the best known environmental factor. However, the effect of tobacco is opposite in CD and UC. A familial history of IBD is the most important risk factor for developing the disease, suggesting a genetic predisposition to IBD. This hypothesis has recently been confirmed by the localization of at least two susceptibility loci on chromosomes 12 and 16. These genes seem to play a role in both CD and UC. They must now to be identified.
The nematode CED-4 protein and its human homolog Apaf-1 play a central role in apoptosis by functioning as direct activators of death-inducing caspases. A novel human CED-4/Apaf-1 family member called CARD4 was identified that has a domain structure strikingly similar to the cytoplasmic, receptor-like proteins that mediate disease resistance in plants. CARD4 interacted with the serine-threonine kinase RICK and potently induced NF-kappaB activity through TRAF-6 and NIK signaling molecules. In addition, coexpression of CARD4 augmented caspase-9-induced apoptosis. Thus, CARD4 coordinates downstream NF-kappaB and apoptotic signaling pathways and may be a component of the host innate immune response.
Crohn's disease is a chronic inflammatory disorder of the gastrointestinal tract, which is thought to result from the effect of environmental factors in a genetically predisposed host. A gene location in the pericentromeric region of chromosome 16, IBD1, that contributes to susceptibility to Crohn's disease has been established through multiple linkage studies, but the specific gene(s) has not been identified. NOD2, a gene that encodes a protein with homology to plant disease resistance gene products is located in the peak region of linkage on chromosome 16 (ref. 7). Here we show, by using the transmission disequilibium test and case-control analysis, that a frameshift mutation caused by a cytosine insertion, 3020insC, which is expected to encode a truncated NOD2 protein, is associated with Crohn's disease. Wild-type NOD2 activates nuclear factor NF-kappaB, making it responsive to bacterial lipopolysaccharides; however, this induction was deficient in mutant NOD2. These results implicate NOD2 in susceptibility to Crohn's disease, and suggest a link between an innate immune response to bacterial components and development of disease.
Crohn's disease and ulcerative colitis, the two main types of chronic inflammatory bowel disease, are multifactorial conditions of unknown aetiology. A susceptibility locus for Crohn's disease has been mapped to chromosome 16. Here we have used a positional-cloning strategy, based on linkage analysis followed by linkage disequilibrium mapping, to identify three independent associations for Crohn's disease: a frameshift variant and two missense variants of NOD2, encoding a member of the Apaf-1/Ced-4 superfamily of apoptosis regulators that is expressed in monocytes. These NOD2 variants alter the structure of either the leucine-rich repeat domain of the protein or the adjacent region. NOD2 activates nuclear factor NF-kB; this activating function is regulated by the carboxy-terminal leucine-rich repeat domain, which has an inhibitory role and also acts as an intracellular receptor for components of microbial pathogens. These observations suggest that the NOD2 gene product confers susceptibility to Crohn's disease by altering the recognition of these components and/or by over-activating NF-kB in monocytes, thus documenting a molecular model for the pathogenic mechanism of Crohn's disease that can now be further investigated.
Background Genetic predisposition to inflammatory bowel disease (IBD) has been shown by epidemiological and linkage studies. Genetic linkage of IBD to chromosome 16 has been previously observed and replicated in independent populations. The recently identified NOD2 gene is a good positional and functional candidate gene since it is located in the region of linkage on chromosome 16q12, and activates nuclear factor (NF) kappaB in response to bacterial lipopolysaccharides. Methods We sequenced the coding region of the NOD2 gene and genotyped an insertion polymorphism affecting the leucine-rich region of the protein product in 512 individuals with IBD from 309 German or British families, 369 German trios (ie, German patients with sporadic IBD and their unaffected parents), and 272 normal controls. We then tested for association with Crohn's disease and ulcerative colitis. Findings Family-based association analyses were consistently positive in 95 British and 99 German affected sibling pairs with Crohn's disease (combined p<0.0001); the association was confirmed in the 304 German trios with Crohn's disease. No association was seen in the 115 sibling pairs and 65 trios with ulcerative colitis. The genotype-specific disease risks conferred by heterozygous and homozygous mutant genotypes were 2.6 (95% CI 1.5-4.5) and 42.1 (4.3-infinity), respectively. Interpretation The insertion mutation in the NOD2 gene confers a substantially increased susceptibility to Crohn's disease but not to ulcerative colitis.
Background & aims:
In Crohn's disease, smoking increases the risk for surgical procedures. The influence of smoking on the overall severity of the disease remains controversial. The purpose of the study was to examine the effects of smoking on the long-term course of Crohn's disease and the relationship between smoking and therapy.
Methods:
The medical charts of 400 consecutive patients whose smoking habits were specified by direct interview were reviewed.
Results:
Frequency and extent of excisional surgery were not significantly different in smokers and nonsmokers, but smokers required more glucocorticoids and immunosuppressive drugs. The effect of smoking on the need for immunosuppressive drugs was dose-dependent and was significant in women but not in men. For female smokers, the 10-year risk of immunosuppressive therapy was 52% +/- 11% compared with 24% +/- 10% for nonsmokers (P < 0.001). The risk of surgery increased only in patients who smoked and did not take immunosuppressive drugs. The surgical rate increased significantly during smoking in 19 patients who started smoking after diagnosis and decreased significantly in 34 patients who stopped compared with matched controls.
Conclusions:
Patients who smoke, particularly women and heavy smokers, run a high risk of developing severe disease. Immunosuppressive therapy neutralizes the influence of smoking on surgical rates.
We propose to describe, in its pathologic and clinical details, a disease of the terminal ileum, affecting mainly young adults, characterized by a subacute or chronic necrotizing and cicatrizing inflammation. The ulceration of the mucosa is accompanied by a disproportionate connective tissue reaction of the remaining walls of the involved intestine, a process which frequently leads to stenosis of the lumen of the intestine, associated with the formation of multiple fistulas.The disease is clinically featured by symptoms that resemble those of ulcerative colitis, namely, fever, diarrhea and emaciation, leading eventually to an obstruction of the small intestine; the constant occurrence of a mass in the right iliac fossa usually requires surgical intervention (resection). The terminal ileum is alone involved. The process begins abruptly at and involves the ileocecal valve in its maximal intensity, tapering off gradually as it ascends the ileum orally for from 8 to 12 inches (20
Infection, ischaemia, physical damage, or specific immunologic sensitivity should be excluded as far as possible before a diagnosis of non-specific inflammatory bowel disease is made. Non-specific inflammations can be subdivided on the basis of macroscopic and microscopic anatomical criteria. Macroscopic structural abnormalities can be recognized by clinical examination, endoscopy, radiology, and inspection of an operation specimen. These complementary methods of data collection combine with microscopic examinations of tissue to separate disorders that differ in prognosis and possible response to treatment. Anatomic classifications do not necessarily imply differences in aetiology and may change with advances in knowledge.
In a case-control study 280 patients with inflammatory bowel disease and matched community controls taken from general practitioner lists were questioned about their smoking habits. Whether assessed by current or previous habits, patients with Crohn's disease were more likely to be or have been smokers than their matched controls and the association appeared strongest at disease onset (matched relative risk 4.1, p less than 0.001). In contrast, patients with ulcerative colitis assessed in the same way were less likely to be smokers compared with their matched controls, with the association being equally strong at disease onset and currently (matched relative risk 0.17, p less than 0.001). Among patients with ulcerative colitis who were exsmokers, 76% reported stopping smoking before disease onset compared with 13% of exsmokers with Crohn's disease. Contrary to earlier reports, there were only small and nonsignificant increases in the risk of Crohn's disease or ulcerative colitis in exsmokers. The strength, temporal precedence, and contrasting nature of the associations are evidence that smoking has an important etiologic role in inflammatory bowel disease. Smoking appears to be a factor in determining whether Crohn's disease or ulcerative colitis develops in predisposed individuals.
In Crohn's disease, smoking increases the risk for surgical procedures. The influence of smoking on the overall severity of the disease remains controversial. The purpose of the study was to examine the effects of smoking on the long-term course of Crohn's disease and the relationship between smoking and therapy.
The medical charts of 400 consecutive patients whose smoking habits were specified by direct interview were reviewed.
Frequency and extent of excisional surgery were not significantly different in smokers and nonsmokers, but smokers required more glucocorticoids and immunosuppressive drugs. The effect of smoking on the need for immunosuppressive drugs was dose-dependent and was significant in women but not in men. For female smokers, the 10-year risk of immunosuppressive therapy was 52% +/- 11% compared with 24% +/- 10% for nonsmokers (P < 0.001). The risk of surgery increased only in patients who smoked and did not take immunosuppressive drugs. The surgical rate increased significantly during smoking in 19 patients who started smoking after diagnosis and decreased significantly in 34 patients who stopped compared with matched controls.
Patients who smoke, particularly women and heavy smokers, run a high risk of developing severe disease. Immunosuppressive therapy neutralizes the influence of smoking on surgical rates.
Crohn's disease (CD) and ulcerative colitis are the major forms of chronic inflammatory bowel diseases in the western world, and occur in young adults with an estimated prevalence of more than one per thousand inhabitants. The causes of inflammatory bowel diseases remain unknown, but genetic epidemiology studies suggest that inherited factors may contribute in part to variation in individual susceptibility to Crohn's disease. A genome-wide search performed on two consecutive and independent panels of families with multiple affected members, using a non-parametric two-point sibling-pair linkage method, identified a putative CD-susceptibility locus on chromosome 16 (P less than 0.01 for each panel). The localization was centered around loci D16S409 and D16S419 by using multipoint sibpair analysis (P less than 1.5x10(-5)). This region of the genome contains candidate genes which may be relevant to the pathogenic mechanism of inflammatory bowel diseases.
Some families have multiple members with inflammatory bowel disease (IBD). Do clinical features of familial differ from sporadic cases? Is there concordance between affected family members? Do environmental factors affect familial clustering?
In 67 families, each with three or more first-degree relatives with IBD, the 213 affected family members were interviewed, and their case records were reviewed.
The clinical manifestations of familial cases did not differ from IBD reported in a large series. There was concordance for type of disease (P < 0.001) but not greater than expected concordance for age at diagnosis, site and extent of disease, or transmural aggressiveness of Crohn's disease. A significant association between smoking with Crohn's disease and non-smoking with ulcerative colitis was found (P < 0.001), even in families with both disorders. The intervals between diagnosis of successive family cases varied so greatly that a single etiologic factor with a constant latent period seems to be unlikely. When IBD affected successive generations, parents were diagnosed at a later age than the children (P < 0.001) and after the child in 12 of 49 cases.
Analysis has not identified any clinically significant differences between familial and sporadic cases of IBD. Tobacco smoking affects disease type in familial cases.
Crohn's disease has a bimodal age distribution of disease onset diagnosis. The peaks (20 and 50 years) may represent different phenotypes or different genetic and/or environmental influences between younger- and older-onset individuals. The aim of this study was to examine the influences of age at diagnosis of Crohn's disease on disease site, type, and course.
Records of 552 consecutive patients with Crohn's disease were reviewed retrospectively.
Younger age at diagnosis (younger than 20 years), compared with an older age (40 years or older), was associated with a greater prevalence of a family history of Crohn's disease (29.9% vs. 13.6%), greater small bowel involvement (88.7% vs. 57.5%), more stricturing disease (45.8% vs. 28.8%), and a higher frequency of surgery (70.6% vs. 55.3%). Older age at diagnosis was associated with a greater prevalence of colonic disease (84.8% vs. 71.2%) and the inflammatory subtype (54.5% vs. 34.4%). A conditional logistic regression analysis confirmed an independent effect of age at diagnosis on ileal disease and surgery for intractable disease.
In Crohn's disease, early age at diagnosis is associated with more complicated disease and a greater likelihood of having affected relatives. Stratification of Crohn's disease by age at diagnosis provides support for the concept of genetic heterogeneity.
Crohn's disease (CD) and ulcerative colitis (UC), the chronic inflammatory bowel diseases (CIBD), are common causes of gastro-intestinal disease in the Western world, with a combined prevalence of 100-200/100,000 (ref. 1). Epidemiological studies, particularly concordance rates in twin pairs and siblings, strongly implicate genetic susceptibility in the pathogenesis of CIBD. In fact, the relative contribution of genetic factors to the pathogenesis of CD may be greater than in schizophrenia, asthma or hypertension, and at least equivalent to that in insulin-dependent diabetes. Systematic screening of the entire human genome now provides a strategy for the identification of susceptibility genes in complex polygenic disorders. We undertook a two-stage genome search for susceptibility genes in inflammatory bowel disease involving 186 affected sibling pairs from 160 nuclear families. We provide strong evidence for the presence of susceptibility loci for both CD and UC on chromosome 3, 7 and 12. We obtained the highest lod score (5.47; P = 2.66 x 10(-7) with the marker D12S83 and lod scores of 3.08 and 2.69 for D7S669 and D3S1573, respectively. Our data suggest that CD and UC are closely related, but distinct, polygenic disorders that share some, but not all, susceptibility genes.
We report here the purification and cDNA cloning of Apaf-1, a novel 130 kd protein from HeLa cell cytosol that participates in the cytochrome c-dependent activation of caspase-3. The NH2-terminal 85 amino acids of Apaf-1 show 21% identity and 53% similarity to the NH2-terminal prodomain of the Caenorhabditis elegans caspase, CED-3. This is followed by 320 amino acids that show 22% identity and 48% similarity to CED-4, a protein that is believed to initiate apoptosis in C. elegans. The COOH-terminal region of Apaf-1 comprises multiple WD repeats, which are proposed to mediate protein-protein interactions. Cytochrome c binds to Apaf-1, an event that may trigger the activation of caspase-3, leading to apoptosis.
Inflammatory bowel disease (IBD) is characterized by a chronic relapsing intestinal inflammation, typically starting in early adulthood. IBD is subdivided into two subtypes, on the basis of clinical and histologic features: Crohn disease and ulcerative colitis (UC). Previous genomewide searches identified regions harboring susceptibility loci on chromosomes 1, 3, 4, 7, 12, and 16. To expand our understanding of the genetic risk profile, we performed a 9-cM genomewide search for susceptibility loci in 268 families containing 353 affected sibling pairs. Previous linkages on chromosomes 12 and 16 were replicated, and the chromosome 4 linkage was extended in this sample. New suggestive evidence for autosomal linkages was observed on chromosomes 1, 6, 10, and 22, with LOD scores of 2.08, 2.07, 2.30, and 1.52, respectively. A maximum LOD score of 1.76 was observed on the X chromosome, for UC, which is consistent with the clinical association of IBD with Ullrich-Turner syndrome. The linkage finding on chromosome 6p is of interest, given the possible contribution of human leukocyte antigen and tumor necrosis-factor genes in IBD. This genomewide linkage scan, done with a large family cohort, has confirmed three previous IBD linkages and has provided evidence for five additional regions that may harbor IBD predisposition genes.
Ced-4 and Apaf-1 belong to a major class of apoptosis regulators that contain caspase-recruitment (CARD) and nucleotide-binding oligomerization domains. Nod1, a protein with an NH2-terminal CARD-linked to a nucleotide-binding domain and a COOH-terminal segment with multiple leucine-rich repeats, was identified. Nod-1 was found to bind to multiple caspases with long prodomains, but specifically activated caspase-9 and promoted caspase-9-induced apoptosis. As reported for Apaf-1, Nod1 required both the CARD and P-loop for function. Unlike Apaf-1, Nod1 induced activation of nuclear factor-kappa-B (NF-kappaB) and bound RICK, a CARD-containing kinase that also induces NF-kappaB activation. Nod1 mutants inhibited NF-kappaB activity induced by RICK, but not that resulting from tumor necrosis factor-alpha stimulation. Thus, Nod1 is a leucine-rich repeat-containing Apaf-1-like molecule that can regulate both apoptosis and NF-kappaB activation pathways.
Chronic inflammatory bowel disease (IBD) presents as two major clinical forms, Crohn's disease (CD) and ulcerative colitis (UC). Genetic epidemiological studies and animal models suggest that inherited factors play significant roles in the susceptibility to both forms of IBD. From four genome-wide scans, putative susceptibility loci on chromosome 16 (IBD1 for CD), and on chromosomes 1, 3, 4, 6, 7, 10, and 12 for IBD, have been identified. Several other groups, including ours, have confirmed linkage to the loci on chromosomes 12 and 16. The aim of this study is to identify other potential susceptibility loci for CD with a genome-wide search approach. In our sample of 222 individuals from 46 families (20 Jewish and 26 non-Jewish), with a total of 65 sibpairs diagnosed with CD, we observed a novel locus with suggestive linkage [multipoint logarithm of the odds score (Mlod) > 2] at chromosome 14q11.2 (Mlod = 2.8, p = 0.0002). In addition, suggestive linkage was observed in our Jewish families at chromosome 17q21-q23 (Mlod = 2.1, p = 0.01) and chromosome 5q33-q35 (Mlod = 2.2, p = 0.0003). The syntenic regions of the latter locus are mapped within two putative loci on mouse chromosomes 11 and 18, which were identified in a mouse IBD model induced by dextran sulfate sodium (29). Our preliminary results provide potential evidence for several susceptibility loci contributing to the risk of CD. The observation of man-mouse synteny may accelerate the identification of CD susceptibility gene(s) on human chromosome 5.
Epidemiological studies have shown that genetic factors contribute to the pathogenesis of the idiopathic inflammatory bowel diseases (IBD), Crohn disease (CD) and ulcerative colitis (UC). Recent genome scans and replication studies have identified replicated linkage between CD and a locus on chromosome 16 (the IBD1 locus), replicated linkage between IBD (especially UC) and a locus on chromosome 12q (the IBD2 locus), and replicated linkage between IBD (especially CD) and a locus on chromosome 6p (the IBD3 locus). Since the estimated locus-specific lambdas values for the regions of replicated linkage do not account for the overall lambdas in CD, and since the published genome scans in IBD show at least nominal evidence for linkage to regions on all but two chromosomes, we performed an independent genome scan using 751 microsatellite loci in 127 CD-affected relative pairs from 62 families. Single-point nonparametric linkage analysis using the GENEHUNTER-PLUS program shows evidence for linkage to the adjacent D14S261 and D14S283 loci on chromosome 14q11-12 (LOD = 3.00 and 1.70, respectively), and the maximal multipoint LOD score is observed at D14S261 (LOD = 3.60). In the multipoint analysis, nominal evidence for linkage (P<.05) is observed near D2S117 (LOD = 1.25), near D3S3045 (LOD = 1.31), between D7S40 and D7S648 (LOD = 0.91), and near D18S61 (LOD = 1.15). Our finding of significant linkage to D14S261 and the finding of suggestive linkage to the same locus in an independent study (multipoint LOD = 2.8) satisfies criteria for confirmed linkage, so we propose that the region of interest on chromosome 14q11-12 should be designated the IBD4 locus.
The chronic inflammatory bowel diseases (IBDs)-Crohn disease (CD) and ulcerative colitis (UC)-are idiopathic, inflammatory disorders of the gastrointestinal tract. These conditions have a peak incidence in early adulthood and a combined prevalence of approximately 100-200/100,000. Although the etiology of IBD is multifactorial, a significant genetic contribution to disease susceptibility is implied by epidemiological data revealing a sibling risk of approximately 35-fold for CD and approximately 15-fold for UC. To elucidate the genetic basis for these disorders, we undertook a genomewide scan in 158 Canadian sib-pair families and identified three regions of suggestive linkage (3p, 5q31-33, and 6p) and one region of significant linkage to 19p13 (LOD score 4.6). Higher-density mapping in the 5q31-q33 region revealed a locus of genomewide significance (LOD score 3.9) that contributes to CD susceptibility in families with early-onset disease. Both of these genomic regions contain numerous genes that are important to the immune and inflammatory systems and that provide good targets for future candidate-gene studies.
The immune response to microbial pathogens is initiated by recognition of specific pathogen components by host cells both at the cell surface and in the cytosol. While the response triggered by pathogen products at the surface of immune cells is well characterized, that initiated in the cytosol is poorly understood. Nod1 is a member of a growing family of intracellular proteins with structural homology to apoptosis regulators Apaf-1/Ced-4 and a class of plant disease-resistant gene products. Here we show that bacterial lipopolysaccharides, but not other pathogen components tested, induced TLR4- and MyD88-independent NF-kappaB activation in human embryonic kidney 293T cells expressing trace amounts of Nod1. Nod2, another Nod family member, also conferred responsiveness to bacterial components but with a response pattern different from that observed with Nod1. As it was reported for plant disease-resistant R proteins, the leucine-rich repeats of Nod1 and Nod2 were required for lipopolysaccharide-induced NF-kappaB activation. A lipopolysaccharide binding activity could be specifically coimmunopurified with Nod1 from cytosolic extracts. These observations suggest that Nod1 and Nod2 are mammalian counterparts of plant disease-resistant gene products that may function as cytosolic receptors for pathogen components derived from invading bacteria.
There is evidence for the IBD1 Crohn's disease (CD) susceptibility locus on chromosome 16 in several but not all populations studied. Genetic and phenotypic heterogeneity may underlie ability to replicate IBD1. We determined if age and severity stratification could identify a clinical subgroup at risk for IBD1.
Linkage analysis at microsatellites spanning chromosome 16 was performed in 2 groups of CD pedigrees: group 1, 57 pedigrees with at least one affected relative classified as having "severe" disease, by history of surgical resection or immunomodulator therapy, and with disease diagnosed before age 22; and group 2, 33 pedigrees with no history of early-onset, severe CD.
Group 1 pedigrees demonstrated genomewide significant linkage evidence for the IBD1 locus (nonparametric multipoint logarithm of the odds [Mlod], 3.84; P = 1.3 x 10(-5)) with linkage evidence greater than all 90 pedigrees (Mlod, 2.12; P = 9.0 x 10(-4)). Group 2 pedigrees had near zero nonparametric 2-point and Mlod scores for the IBD1 region. Heterogeneity between groups 1 and 2 was significant (P = 0.002).
Presence of early-onset, more severe CD identifies pedigrees at high risk for IBD1. These pedigrees will have more power to refine the IBD1 locus and identify the causative gene.
The technique of genomewide scanning has been applied successfully in inflammatory bowel disease (IBD). A number of putative susceptibility loci have been identified through genomewide searches including replicated regions of linkage on chromosomes 12, 16, 6 (the HLA region), and 14. We have investigated the contribution of the X chromosome in 145 Belgian affected relative pairs.
In the first stage of the study, 79 (68 CD, 11 mixed) sibling pairs were genotyped at 12 microsatellite markers covering the X chromosome. In the second stage, 10 additional markers in the X-pericentromeric region were studied in the families involved in stage 1 together with 62 additional families (52 sibling pairs, 14 second-degree relative pairs).
In the first stage, evidence for linkage was found over a 30-cM pericentromeric region spanning dXs991, dXs990, and dXs8096 (multipoint maximum LOD score in the CD subgroup, 2.5; P = 0.0003). The remainder of the X chromosome was excluded (exclusion under LOD-2) for a locus with lambda(s) = 2. Fine mapping in the second stage confirmed linkage, and narrowed and shifted the linked region to Xq21.3 around dXs1203 (nonparametric linkage [NPL], 2.90; P = 0.0017). The NPL-1 interval around the linkage peak comprises 19.7 cM.
These data provide suggestive evidence for the presence and chromosomal location of an X-linked susceptibility gene in IBD.
Numerous familial, non-Mendelian (i.e., complex) diseases have been screened by linkage analysis for regions harboring susceptibility genes. Except for rare, high-penetrance syndromes showing Mendelian inheritance, such as BRCA1 and BRCA2, most attempts have failed to produce replicable linkage findings. For example, in multiple sclerosis and other complex diseases, there have been many reports of significant linkage, followed by numerous failures to replicate. In inflammatory bowel disease (IBD), linkage to two regions has elsewhere been reported at genomewide significance levels: the pericentromeric region on chromosome 16 (IBD1) and chromosome 12q (IBD2). As with other complex diseases, the subsequent support for these localizations has been variable. In this article, we report the results of an international collaborative effort to investigate these putative localization by pooling of data sets that do not individually provide convincing evidence for linkage to these regions. Our results, generated by the genotyping and analysis of 12 microsatellite markers in 613 families, provide unequivocal replication of linkage for a common human disease: a Crohn disease susceptibility locus on chromosome 16 (maximum LOD score 5.79). Despite failure to replicate the previous evidence for linkage on chromosome 12, the results described herein indicate the need to further investigate the potential role of this locus in susceptibility to ulcerative colitis. This report provides a convincing example of the collaborative approach necessary to obtain the sample numbers required to achieve statistical power in studies of complex human traits.
Van Gos-sum, M. Van Winckel, and M. Veyrac, as well as gastroenter-ologists from Nord, Pas de Calais, Somme, and Seine Maritime, generously assisted in the recruitment of families for the study. We are grateful to Dr
- Drs J Balanzo
- B Bonaz
- Y Bouhnik
- G Cadiot
- A Cortot
- S Cucchiara
- B Crusius
- J J Delchier
- B Duclos
- J L Dupas
- J P Galmiche
- J P Gendre
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We are grateful to the patients and their families for their participation in this study. Drs. J. Balanzo, B. Bonaz, Y. Bouhnik, G. Cadiot, A. Cortot, S. Cucchiara, B. Crusius, J. J. Delchier, B. Duclos, J. L. Dupas, J. P. Galmiche, J. P. Gendre, D. Golfain, C. Grä nnö, D. Heresbach, A. Lachaux, H. Lautraite, C. Len-aerts, E. Lerebours, V. Levy, R. Lö fberg, H. Malchow, P. Mar-teau, A. Morali, F. Pallone, S. Pena, A. Rotenberg, I. Rousseau, J. Schmitz, F. Shanahan, I. Sobhani, H. Svensson, A. Van Gos-sum, M. Van Winckel, and M. Veyrac, as well as gastroenter-ologists from Nord, Pas de Calais, Somme, and Seine Maritime, generously assisted in the recruitment of families for the study. We are grateful to Dr. C. Bellanné-Chantelot, J. C. Beaudoin, C. Billon, O. Bluteau, T. H. Bui, L. Cazes, C. Guidicelli, P.
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