A unique challenge faced by the intestinal immune system is the requirement for tolerance to luminal bacteria while simultaneously defending against pathogenic organisms. The dysregulation of this balance can lead to uncontrolled activation of the gastrointestinal immune system, a feature of Crohn's disease and ulcerative colitis. The recognition and response to microbial organisms is mediated in part by pattern recognition receptors (PRR), which include nucleotide oligomerization domain 2 (NOD2), an intracellular bacterial recognition receptor for peptidoglycan (PGN), a component of both gram positive and gram negative cell walls. Crohn's disease is associated with loss-of-function polymorphisms in NOD2 (CARD15). However, the specific NOD2 dysfunction(s) leading to the development of Crohn's disease is not yet understood. Upon acute stimulation of NOD2, either alone or in combination with other bacterial receptors, peripherally-derived antigen presenting cells (APC) secrete proinflammatory cytokines. Intestinal macrophages and dendritic cells (DC) are in large part derived from circulating monocytes that migrate into the intestinal tissues and undergo differentiation based on the intestinal environment. The intestinal immune system is an environment which results in chronic stimulation. Therefore, it is critical to understand the consequences of chronic stimulation through NOD2. We have recently found that chronic stimulation through NOD2 in primary peripheral monocyte-derived human macrophages results in reduced production of pro- inflammatory cytokines upon restimulation either through NOD2 or through other PRR. This downregulation in production of pro-inflammatory cytokine secretion is similar to the tolerance, or significantly reduced secretion of pro-inflammatory cytokines, exhibited by intestinal macrophages stimulated with bacterial products. On the other hand, intestinal APC from individuals with inflammatory bowel disease (IBD) have excess production of pro-inflammatory cytokines. We hypothesize that the tolerance induced in primary human APC upon stimulation through NOD2 is mediated by a combination of mechanisms that selectively affect specific signaling pathways (e.g. pro-inflammatory cytokines), and that some of these mechanisms are different than those mediated by chronic stimulation of Toll like receptors (TLR). This proprosal will define the tolerance defects in human peripheral macrophages expressing the various Crohn's disease- associated NOD2 mutations, the mechanisms mediating peripheral macrophage tolerance through chronic NOD2 versus TLR stimulation, and then extend these findings directly into human intestinal macrophages.

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We have recently found that chronic stimulation through NOD2 in primary peripheral monocyte-derived human macrophages results in reduced production of proinflammatory cytokines upon restimulation either through NOD2 or through other pattern recognition receptors. We seek to define the combination of NOD2-mediated mechanisms that selectively affect the downregulation of these pro-inflammatory cytokines, and determine if these mechanisms are different from those induced upon chronic stimulation of Toll like receptors. We will then extend these findings directly into human intestinal macrophages and determine if individuals harboring Crohn's disease associated NOD2 mutations are defective in these mechanisms.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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Gastrointestinal Mucosal Pathobiology Study Section (GMPB)
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Hamilton, Frank A
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Yale University
Internal Medicine/Medicine
Schools of Medicine
New Haven
United States
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Hedl, Matija; Abraham, Clara (2016) A TPL2 (MAP3K8) disease-risk polymorphism increases TPL2 expression thereby leading to increased pattern recognition receptor-initiated caspase-1 and caspase-8 activation, signalling and cytokine secretion. Gut 65:1799-1811
Zheng, Shasha; Hedl, Matija; Abraham, Clara (2015) Twist1 and Twist2 Contribute to Cytokine Downregulation following Chronic NOD2 Stimulation of Human Macrophages through the Coordinated Regulation of Transcriptional Repressors and Activators. J Immunol 195:217-26
Zheng, Shasha; Hedl, Matija; Abraham, Clara (2015) TAM receptor-dependent regulation of SOCS3 and MAPKs contributes to proinflammatory cytokine downregulation following chronic NOD2 stimulation of human macrophages. J Immunol 194:1928-37
Lahiri, Amit; Hedl, Matija; Abraham, Clara (2015) MTMR3 risk allele enhances innate receptor-induced signaling and cytokines by decreasing autophagy and increasing caspase-1 activation. Proc Natl Acad Sci U S A 112:10461-6
Hedl, Matija; Lahiri, Amit; Ning, Kaida et al. (2014) Pattern recognition receptor signaling in human dendritic cells is enhanced by ICOS ligand and modulated by the Crohn's disease ICOSLG risk allele. Immunity 40:734-46
Lahiri, Amit; Abraham, Clara (2014) Activation of pattern recognition receptors up-regulates metallothioneins, thereby increasing intracellular accumulation of zinc, autophagy, and bacterial clearance by macrophages. Gastroenterology 147:835-46
Wu, Xingxin; Lahiri, Amit; Haines 3rd, G Kenneth et al. (2014) NOD2 regulates CXCR3-dependent CD8+ T cell accumulation in intestinal tissues with acute injury. J Immunol 192:3409-18
Hedl, Matija; Abraham, Clara (2014) A TNFSF15 disease-risk polymorphism increases pattern-recognition receptor-induced signaling through caspase-8-induced IL-1. Proc Natl Acad Sci U S A 111:13451-6
Hedl, Matija; Zheng, Shasha; Abraham, Clara (2014) The IL18RAP region disease polymorphism decreases IL-18RAP/IL-18R1/IL-1R1 expression and signaling through innate receptor-initiated pathways. J Immunol 192:5924-32
Zheng, Shasha; Abraham, Clara (2013) NF-?B1 inhibits NOD2-induced cytokine secretion through ATF3-dependent mechanisms. Mol Cell Biol 33:4857-71

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