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NOD2 Mutations Affect Muramyl Dipeptide Stimulation of Human B Lymphocytes and Interact with Other IBD-Associated Genes

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Abstract

Background

Genetic and functional studies have associated variants in the NOD2/CARD15 gene with Crohn’s disease.

Aims

This study aims to replicate the association of three common NOD2 mutations with Crohn’s disease, study its effect on NOD2 expression in B cells and its interaction with other IBD-associated genes.

Methods

A total of 294 IBD patients (179 familial IBD, 115 sporadic IBD) and 298 unrelated healthy controls were from central Pennsylvania. NOD2 mutations were analyzed by primer-specific amplification, PCR based-RFLP, and validated with the ABI SNPlexM genotyping system. Gene–gene interaction was studied using a statistical model for epistasis analysis.

Results

Three common NOD2 mutations are associated with Crohn’s disease (p = 5.08 × 10−7, 1.67 × 10−6, and 1.87 × 10−2 for 1007fs, R720W, and G908R, respectively), but not with ulcerative colitis (p = 0.1046, 0.1269, and 0.8929, respectively). For IBD overall, 1007finsC (p = 4.4 × 10−5) and R720W (p = 9.24 × 10−5) were associated with IBD, but not G908R (p = 0.1198). We revealed significant interactions of NOD2 with other IBD susceptibility genes IL23R, DLG5, and OCTN1. We discovered that NOD2 was expressed in both normal human peripheral blood B cells and in EBV-transformed B cell lines. Moreover, we further demonstrated that muramyl dipeptide (MDP) stimulation of B lymphocytes up-regulated expression of NF-κB-p50 mRNA.

Conclusion

NOD2 is expressed in peripheral B cells, and the up-regulation of NOD2 expression by MDP was significantly impaired by NOD2 mutations. The finding suggests a possible role of NOD2 in the immunological response in IBD pathogenesis.

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References

  1. Xavier RJ, Rioux JD. Genome-wide association studies: a new window into immune-mediated diseases. Nat Rev Immunol. 2008;8:631–643.

    Article  PubMed  CAS  Google Scholar 

  2. Zhang H, Massey D, Tremelling M, et al. Genetics of inflammatory bowel disease: clues to pathogenesis. Br Med Bull. 2008;87:17–30.

    Article  PubMed  CAS  Google Scholar 

  3. Hugot JP, Chamaillard M, Zouali H, et al. Association of NOD2 leucine-rich repeat variants with susceptibility to Crohn’s disease. Nature. 2001;411:599–603.

    Article  PubMed  CAS  Google Scholar 

  4. Wellcome Trust Case Control C. Genome-wide association study of 14,000 cases of seven common diseases and 3,000 shared controls. Nature. 2007;447:661–678.

    Article  Google Scholar 

  5. Hampe J, Franke A, Rosenstiel P, et al. A genome-wide association scan of nonsynonymous SNPs identifies a susceptibility variant for Crohn disease in ATG16L1. Nature Genet. 2007;39:207–211.

    Article  PubMed  CAS  Google Scholar 

  6. Duerr RH, Taylor KD, Brant SR, et al. A genome-wide association study identifies IL23R as an inflammatory bowel disease gene. Science. 2006;314:1461–1463.

    Article  PubMed  CAS  Google Scholar 

  7. Radford-Smith G, Pandeya N. Associations between NOD2/CARD15 genotype and phenotype in Crohn’s disease—Are we there yet? World J Gastroenterol. 2006;12:7097–7103.

    PubMed  Google Scholar 

  8. Csillag C, Nielsen OH, Borup R, et al. CARD15 status and familial predisposition for Crohn’s disease and colonic gene expression. Dig Dis Sci. 2007;52:1783–1789.

    Article  PubMed  CAS  Google Scholar 

  9. Henckaerts L, Van Steen K, Verstreken I, et al. Genetic risk profiling and prediction of disease course in Crohn’s disease patients. Clin Gastroenterol Hepatol. 2009;7:e972.

    Article  Google Scholar 

  10. Tohno M, Shimazu T, Ueda W, et al. Molecular cloning of porcine RP105/MD-1 involved in recognition of extracellular phosphopolysaccharides from Lactococcus lactis ssp. cremoris. Mol Immunol. 2007;44:2566–2577.

    Article  PubMed  CAS  Google Scholar 

  11. Cobrin GM, Abreu MT. Defects in mucosal immunity leading to Crohn’s disease. Immunol Rev. 2005;206:277–295.

    Article  PubMed  CAS  Google Scholar 

  12. Girardin SE, Boneca IG, Viala J, et al. Nod2 is a general sensor of peptidoglycan through muramyl dipeptide (MDP) detection. J Biol Chem. 2003;278:8869–8872.

    Article  PubMed  CAS  Google Scholar 

  13. Wang Z, Liu T, Lin Z, et al. A general model for multilocus epistatic interactions in case-control studies. PLoS One. 2010;5:e11384.

    Article  PubMed  Google Scholar 

  14. Yu W, Lin Z, Kelly AA, et al. Association of Nk2–3 polymorphism with Crohn’s disease and expression of Nk2–3 is up-regulated in B cell lines and intestinal tissues with Crohn’s disease. J Crohns Colitis. 2009;3:189–195.

    Article  PubMed  Google Scholar 

  15. Lin Z, Cui X, Li H. Multiplex genotype determination at a large number of gene loci. Proc Natl Acad Sci USA. 1996;93:2582–2587.

    Article  PubMed  CAS  Google Scholar 

  16. De la Vega FM, Lazaruk KD, Rhodes MD, et al. Assessment of two flexible and compatible SNP genotyping platforms: TaqMan SNP Genotyping Assays and the SNPlex genotyping system. Mutat Res. 2005;573:111–135.

    Article  PubMed  Google Scholar 

  17. Tobler AR, Short S, Andersen MR, et al. The SNPlex genotyping system: a flexible and scalable platform for SNP genotyping. J Biomol Tech. 2005;16:398–406.

    PubMed  Google Scholar 

  18. Inohara N, Ogura Y, Fontalba A, et al. Host recognition of bacterial muramyl dipeptide mediated through NOD2. Implications for Crohn’s disease. J Biol Chem. 2003;278:5509–5512.

    Article  PubMed  CAS  Google Scholar 

  19. Cho JH. Significant role of genetics in IBD: the NOD2 gene. Rev Gastroenterol Disord. 2003;3:S18–S22.

    PubMed  Google Scholar 

  20. Mahurkar S, Banerjee R, Rani SV, et al. Common variants in NOD2 and IL23R are not associated with inflammatory bowel disease in Indian patients. J Gastroenterol Hepatol. 2011;26:694–699.

    Article  PubMed  CAS  Google Scholar 

  21. Inoue N, Tamura K, Kinouchi Y, et al. Lack of common NOD2 variants in Japanese patients with Crohn’s disease. Gastroenterology. 2002;123:86–91.

    Article  PubMed  CAS  Google Scholar 

  22. Vermeire S, Van Assche G, Rutgeerts P. Inflammatory bowel disease and colitis? new concepts from the bench and the clinic. Curr Opin Gastroenterol. 2011;27:32–37.

    Article  PubMed  Google Scholar 

  23. Ogura Y, Inohara N, Benito A, et al. Nod2, a Nod1/Apaf-1 family member that is restricted to monocytes and activates NF-kappaB. J Biol Chem. 2001;276:4812–4818.

    Article  PubMed  CAS  Google Scholar 

  24. Gutierrez O, Pipaon C, Inohara N, et al. Induction of NOD2 in myelomonocytic and intestinal epithelial cells via nuclear factor-kappa B activation. J Biol Chem. 2002;277:41701–41705.

    Article  PubMed  CAS  Google Scholar 

  25. Muzio M, Scielzo C, Bertilaccio MT, et al. Expression and function of toll like receptors in chronic lymphocytic leukaemia cells. Br J Haematol. 2009;144:507–516.

    Article  PubMed  CAS  Google Scholar 

  26. Petterson T, Jendholm J, Mansson A, Bjartell A, Riesbeck K, Cardell LO. Effects of NOD-like receptors in human B lymphocytes and crosstalk between NOD1/NOD2 and Toll-like receptors. J Leukoc Biol. 2011;89:177–187.

    Article  PubMed  CAS  Google Scholar 

  27. Netea MG, Ferwerda G, de Jong DJ, et al. The frameshift mutation in NOD2 results in unresponsiveness not only to NOD2- but also NOD1-activating peptidoglycan agonists. The J Biol Chem. 2005;280:35859–35867.

    Article  CAS  Google Scholar 

  28. Chamaillard M, Philpott D, Girardin SE, et al. Gene-environment interaction modulated by allelic heterogeneity in inflammatory diseases. Proc Natl Acad Sci USA. 2003;100:3455–3460.

    Article  PubMed  CAS  Google Scholar 

  29. Bonen DK, Ogura Y, Nicolae DL, et al. Crohn’s disease-associated NOD2 variants share a signaling defect in response to lipopolysaccharide and peptidoglycan. Gastroenterology. 2003;124:140–146.

    Article  PubMed  CAS  Google Scholar 

  30. Rahman MK, Midtling EH, Svingen PA, et al. The pathogen recognition receptor NOD2 regulates human FOXP3+T cell survival. J Immunol. 2010;184:7247–7256.

    Article  PubMed  CAS  Google Scholar 

  31. Beynon V, Cotofana S, Brand S, et al. NOD2/CARD15 genotype influences MDP-induced cytokine release and basal IL-12p40 levels in primary isolated peripheral blood monocytes. Inflamm Bowel Dis. 2008;14:1033–1040.

    Article  PubMed  Google Scholar 

  32. Li J, Moran T, Swanson E, et al. Regulation of IL-8 and IL-1beta expression in Crohn’s disease associated NOD2/CARD15 mutations. Hum Mol Genet. 2004;13:1715–1725.

    Article  PubMed  CAS  Google Scholar 

  33. Sabbah A, Chang TH, Harnack R, et al. Activation of innate immune antiviral responses by NOD2. Nat Immunol. 2009;10:1073–1080.

    Article  PubMed  CAS  Google Scholar 

  34. Netea MG, Ferwerda G, de Jong DJ, et al. NOD2 3020insC mutation and the pathogenesis of Crohn’s disease: impaired IL-1beta production points to a loss-of-function phenotype. Neth J Med. 2005;63:305–308.

    PubMed  CAS  Google Scholar 

  35. van Heel DA, Ghosh S, Hunt KA, et al. Synergy between TLR9 and NOD2 innate immune responses is lost in genetic Crohn’s disease. Gut. 2005;54:1553–1557.

    Article  PubMed  Google Scholar 

  36. van Heel DA, Ghosh S, Butler M, et al. Muramyl dipeptide and toll-like receptor sensitivity in NOD2-associated Crohn’s disease. Lancet. 2005;365:1794–1796.

    Article  PubMed  Google Scholar 

  37. Comalada M, Peppelenbosch MP. Impaired innate immunity in Crohn’s disease. Trends Mol Med. 2006;12:397–399.

    Article  PubMed  CAS  Google Scholar 

  38. Noguchi E, Homma Y, Kang X, et al. A Crohn’s disease-associated NOD2 mutation suppresses transcription of human IL10 by inhibiting activity of the nuclear ribonucleoprotein hnRNP-A1. Nat Immunol. 2009;10:471–479.

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

The authors thank Tony Lin for his assistant with RFLP genotyping. This work is supported by a grant from the Philadelphia Health Care Trust (to W.A Koltun) and a Surgery Initiation Grant from Pennsylvania State University College of Medicine (to Z. Lin).

Conflict of interest

None.

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Authors and Affiliations

  1. Division of Colon and Rectal Surgery, Department of Surgery, Pennsylvania State University College of Medicine, H137, 500 University Drive, Hershey, PA, 17033, USA

    Zhenwu Lin, John P. Hegarty, Gerrit John, Rishabh Sehgal, Danielle M. Pastor, Yunhua Wang, Leonard R. Harris III, Lisa S. Poritz & Walter A. Koltun

  2. Department of Public Health Science, Pennsylvania State University College of Medicine, Hershey, PA, USA

    Arthur Berg & Zhong Wang

  3. Department of General Internal Medicine, Christian-Albrechts-University, Kiel, Germany

    Stefan Schreiber

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  1. Zhenwu Lin
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  2. John P. Hegarty
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  4. Arthur Berg
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  5. Zhong Wang
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Corresponding authors

Correspondence to Zhenwu Lin or Walter A. Koltun.

Additional information

Zhenwu Lin, John P. Hegarty and Gerrit John equally contributed to this article.

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Lin, Z., Hegarty, J.P., John, G. et al. NOD2 Mutations Affect Muramyl Dipeptide Stimulation of Human B Lymphocytes and Interact with Other IBD-Associated Genes. Dig Dis Sci 58, 2599–2607 (2013). https://doi.org/10.1007/s10620-013-2696-8

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  • DOI: https://doi.org/10.1007/s10620-013-2696-8

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