Age-of-diagnosis dependent ileal immune intensification and reduced alpha-defensin in older versus younger pediatric Crohn Disease patients despite already established dysbiosis

Article metrics

Abstract

Age-of-diagnosis associated variation in disease location and antimicrobial sero-reactivity has suggested fundamental differences in pediatric Crohn Disease (CD) pathogenesis. This variation may be related to pubertal peak incidence of ileal involvement and Peyer’s patches maturation, represented by IFNγ-expressing Th1 cells. However, direct mucosal evidence is lacking. We characterize the global pattern of ileal gene expression and microbial communities in 525 treatment-naive pediatric CD patients and controls (Ctl), stratifying samples by their age-of-diagnosis. We show a robust ileal gene signature notable for higher expression of specific immune genes including GM-CSF and INFγ, and reduced expression of antimicrobial Paneth cell α-defensins, in older compared to younger patients. Reduced α-defensin expression in older patients was associated with higher IFNγ expression. By comparison, the CD-associated ileal dysbiosis, characterized by expansion of Enterobacteriaceae and contraction of Lachnospiraceae and Ruminococcaceae, was already established within the younger group and did not vary systematically with increasing age-of-diagnosis. Multivariate analysis considering individual taxa, however did demonstrate negative associations between Lachnospiraceae and IFNγ, and positive associations between Bacteroides and α-defensin expression. These data provide evidence for maturation of mucosal Th1 immune responses and loss of epithelial antimicrobial α-defensins which are associated with specific taxa with increasing age-of-diagnosis in pediatric CD.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

References

  1. 1.

    Ruel, J., Ruane, D., Mehandru, S., Gower-Rousseau, C. & Colombel, J. F. IBD across the age spectrum-is it the same disease? Nature reviews. Gastroenterol. & Hepatol. 11, 88–98 (2014).

  2. 2.

    Siegel, C. A. et al. Real-time tool to display the predicted disease course and treatment response for children with Crohn's disease. Inflamm. Bowel Dis. 17, 30–38 (2011).

  3. 3.

    Gower-Rousseau, C. et al. Epidemiology of inflammatory bowel diseases: new insights from a French population-based registry (EPIMAD). Dig. Liver Dis. : Off. J. Ital. Soc. Gastroenterol. Ital. Assoc. Study Liver 45, 89–94 (2013).

  4. 4.

    Heyman, M. B. et al. Children with early-onset inflammatory bowel disease (IBD): analysis of a pediatric IBD consortium registry. J. Pediatr. 146, 35–40 (2005).

  5. 5.

    Meinzer, U. et al. Ileal involvement is age dependent in pediatric Crohn's disease. Inflamm. Bowel Dis. 11, 639–644 (2005).

  6. 6.

    Levine, A. et al. Pediatric modification of the Montreal classification for inflammatory bowel disease: the Paris classification. Inflamm. Bowel Dis. 17, 1314–1321 (2011).

  7. 7.

    Uhlig, H. H. et al. The diagnostic approach to monogenic very early onset inflammatory bowel disease. Gastroenterology 147, 990–1007 e1003 (2014).

  8. 8.

    Muise, A. M., Snapper, S. B. & Kugathasan, S. The age of gene discovery in very early onset inflammatory bowel disease. Gastroenterology 143, 285–288 (2012).

  9. 9.

    de Bie, C. I. et al. Diagnostic workup of paediatric patients with inflammatory bowel disease in Europe: results of a 5-year audit of the EUROKIDS registry. J. Pediatr. Gastroenterol. Nutr. 54, 374–380 (2012).

  10. 10.

    Cornes, J. S. Number, size, and distribution of Peyer's patches in the human small intestine: Part I The development of Peyer's patches. Gut 6, 225–229 (1965).

  11. 11.

    Van Kruiningen, H. J., Ganley, L. M. & Freda, B. J. The role of Peyer's patches in the age-related incidence of Crohn's disease. J. Clin. Gastroenterol. 25, 470–475 (1997).

  12. 12.

    Jung, C., Hugot, J. P. & Barreau, F. Peyer's Patches: the immune sensors of the intestine. Int. J. Inflam. 2010, 823710 (2010).

  13. 13.

    Rhee, S. J., Walker, W. A. & Cherayil, B. J. Developmentally regulated intestinal expression of IFN-gamma and its target genes and the age-specific response to enteric Salmonella infection. J. Immunol. 175, 1127–1136 (2005).

  14. 14.

    Salvati, V. M. et al. Enhanced expression of interferon regulatory factor-1 in the mucosa of children with celiac disease. Pediatr. Res. 54, 312–318 (2003).

  15. 15.

    Salzman, N. H. et al. Enteric defensins are essential regulators of intestinal microbial ecology. Nat. Immunol. 11, 76–83 (2010).

  16. 16.

    Wehkamp, J. et al. Reduced Paneth cell alpha-defensins in ileal Crohn's disease. Proc. Natl Acad. Sci. USA. 102, 18129–18134 (2005).

  17. 17.

    Ramasundara, M., Leach, S. T., Lemberg, D. A. & Day, A. S. Defensins and inflammation: the role of defensins in inflammatory bowel disease. J. Gastroenterol. Hepatol. 24, 202–208 (2009).

  18. 18.

    Kugathasan, S. et al. Prediction of complicated disease course for children newly diagnosed with Crohn's disease: a multicentre inception cohort study. Lancet, https://doi.org/10.1016/S0140-6736(17)30317-3 (2017).

  19. 19.

    Haberman, Y. et al. Pediatric Crohn disease patients exhibit specific ileal transcriptome and microbiome signature. J. Clin. Invest. 124, 3617–3633 (2014).

  20. 20.

    Gevers, D. et al. The treatment-naive microbiome in new-onset Crohn's disease. Cell. Host. Microbe 15, 382–392 (2014).

  21. 21.

    Dubinsky, M. C. et al. Increased immune reactivity predicts aggressive complicating Crohn's disease in children. Clin. Gastroenterol. Hepatol. 6, 1105–1111 (2008).

  22. 22.

    Han, X. et al. Granulocyte-macrophage colony-stimulating factor autoantibodies in murine ileitis and progressive ileal Crohn's disease. Gastroenterology 136, 1261–1271 (2009). e1261-1263.

  23. 23.

    Bray, N. L., Pimentel, H., Melsted, P. & Pachter, L. Near-optimal probabilistic RNA-seq quantification. Nat. Biotechnol. 34, 525–527 (2016).

  24. 24.

    Chen, J., Bardes, E. E., Aronow, B. J. & Jegga, A. G. ToppGene Suite for gene list enrichment analysis and candidate gene prioritization. Nucleic Acids Res. 37, W305–W311 (2009).

  25. 25.

    Kaimal, V., Bardes, E. E., Tabar, S. C., Jegga, A. G. & Aronow, B. J. ToppCluster: a multiple gene list feature analyzer for comparative enrichment clustering and network-based dissection of biological systems. Nucleic Acids Res. 38, W96–W102 (2010).

  26. 26.

    Saito, R. et al. A travel guide to Cytoscape plugins. Nat. Methods 9, 1069–1076 (2012).

  27. 27.

    Caporaso, J. G. et al. Ultra-high-throughput microbial community analysis on the Illumina HiSeq and MiSeq platforms. Isme. J. 6, 1621–1624 (2012).

  28. 28.

    McDonald, D. et al. An improved Greengenes taxonomy with explicit ranks for ecological and evolutionary analyses of bacteria and archaea. Isme. J. 6, 610–618 (2012).

  29. 29.

    Vandussen, K. L. et al. Genetic variants synthesize to produce paneth cell phenotypes that define subtypes of Crohn's disease. Gastroenterology 146, 200–209 (2014).

  30. 30.

    Farin, H. F. et al. Paneth cell extrusion and release of antimicrobial products is directly controlled by immune cell-derived IFN-gamma. J. Exp. Med. 211, 1393–1405 (2014).

  31. 31.

    Biswas, A. et al. Induction and rescue of Nod2-dependent Th1-driven granulomatous inflammation of the ileum. Proc. Natl Acad. Sci. USA. 107, 14739–14744 (2010).

  32. 32.

    Frank, D. N. et al. Molecular-phylogenetic characterization of microbial community imbalances in human inflammatory bowel diseases. Proc. Natl Acad. Sci. USA. 104, 13780–13785 (2007).

  33. 33.

    Simms, L. A. et al. Reduced alpha-defensin expression is associated with inflammation and not NOD2 mutation status in ileal Crohn's disease. Gut 57, 903–910 (2008).

  34. 34.

    Shanahan, M. T. et al. Mouse Paneth cell antimicrobial function is independent of Nod2. Gut, https://doi.org/10.1136/gutjnl-2012-304190 (2013).

  35. 35.

    Perminow, G. et al. Defective paneth cell-mediated host defense in pediatric ileal Crohn's disease. Am. J. Gastroenterol. 105, 452–459 (2010).

  36. 36.

    Koslowski, M. J. et al. Association of a functional variant in the Wnt co-receptor LRP6 with early onset ileal Crohn's disease. PLoS. Genet. 8, e1002523 (2012).

  37. 37.

    Beisner, J. et al. TCF-1 mediated Wnt Signaling regulates Paneth cell innate immune defense effectors HD-5 and -6: implications for Crohn's disease. American journal of physiology. Gastrointestinal and liver physiology, https://doi.org/10.1152/ajpgi.00347.2013 (2014).

  38. 38.

    Yatsunenko, T. et al. Human gut microbiome viewed across age and geography. Nature 486, 222–227 (2012).

  39. 39.

    Gordon, J. I., Dewey, K. G., Mills, D. A. & Medzhitov, R. M. The human gut microbiota and undernutrition. Sci. Transl. Med. 4, 137ps112 (2012).

  40. 40.

    Harris, R. A. et al. Colonic mucosal epigenome and microbiome development in children and adolescents. J. Immunol. Res. 2016, 9170162 (2016).

  41. 41.

    Hollister, E. B. et al. Structure and function of the healthy pre-adolescent pediatric gut microbiome. Microbiome 3, 36 (2015).

  42. 42.

    Assa, A. et al. Mucosa-associated ileal microbiota in new-onset pediatric Crohn's Disease. Inflamm. Bowel Dis. 22, 1533–1539 (2016).

  43. 43.

    Noble, C. L. et al. Characterization of intestinal gene expression profiles in Crohn's disease by genome-wide microarray analysis. Inflamm. Bowel Dis. 16, 1717–1728 (2010).

  44. 44.

    Arijs, I. et al. Predictive value of epithelial gene expression profiles for response to infliximab in Crohn's disease. Inflamm. Bowel Dis. 16, 2090–2098 (2010).

  45. 45.

    Renz, H., Brandtzaeg, P. & Hornef, M. The impact of perinatal immune development on mucosal homeostasis and chronic inflammation. Nat. Rev. Immunol. 12, 9–23 (2012).

Download references

Acknowledgements

This work was supported by the Crohn’s and Colitis Foundation, the Gene and Protein Expression and Bioinformatics cores of the National Institutes of Health (NIH)-supported Cincinnati Children’s Hospital Research Foundation Digestive Health Center (1P30DK078392-01), NIH grant U54 DE023798 and HMP2 (R.J.X., and C.H), the Leona M. and Harry B. Helmsley Charitable Trust (R.J.X., and C.H), the European Crohn’s and Colitis Organization (ECCO, Y.H), the Israel Science Foundation (grant No 908/15), the I-CORE program (Y.H), and European Research Council starting grant (grant No 758313, Y.H). We thank the Crohn’s and Colitis Foundation RISK study publication committee for critical review of this manuscript.

Author information

Correspondence to Yael Haberman.

Ethics declarations

Competing interests

The authors declare no competing interests.

Supplementary information

Suppl. figure 1

Suppl. table1

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Further reading