Regulation of IL-8 and IL-1beta expression in Crohn's disease associated NOD2/CARD15 mutations

Hum Mol Genet. 2004 Aug 15;13(16):1715-25. doi: 10.1093/hmg/ddh182. Epub 2004 Jun 15.


Crohn's disease (CD) is a chronic inflammation affecting the gastrointestinal tract. Three mutations (Arg702Trp, Gly908Arg and Leu1007fsinsC) within the NOD2/CARD15 gene increase CD susceptibility. Here, we define cytokine regulation in primary human mononuclear cells, with muramyl dipeptide (MDP), the minimal NOD2/CARD15 activating component of peptidoglycan. By microarray, MDP induces a broad array of transcripts, including interleukin 1beta (IL-1beta) and interleukin 8 (IL-8). Leu1007fsinsC homozygotes demonstrated decreased transcriptional response to MDP. Electromobility shift assay demonstrated that MDP-induced NF-kappaB activation is mediated via p50 and p65 subunits, but not RelB or c-Rel. In wild-type individuals, MDP-induced IL-8 protein expression with a greater response to high dose (1 micro g/ml) compared with low-dose (10 ng/ml) MDP. At low MDP doses, in all homozygotes, we observed no induction of IL-8 protein. With high doses of MDP, Leu1007fsinsC homozygotes showed no induction. Modest induction of IL-8 protein was observed in Gly908Arg and Arg702Trp homozygotes, indicating varying MDP sensitivity of the CD-associated mutations. In wild-type healthy control, CD and ulcerative colitis individuals, low-dose MDP and TNFalpha alone results in only modest IL-1beta protein induction. With MDP plus TNFalpha, there is a synergistic induction of IL-1beta secretion. In Leu1007fsinsC homozygotes, there is a profound defect in IL-1beta secretion, despite marked induction of IL-1beta mRNA. These findings demonstrate post-transcriptional dependency on the NOD2/CARD15 pathway for IL-1beta secretion with MDP and TNFalpha treatment. Taken together, these studies suggest that a signaling defect of innate immunity to MDP may be an essential underlying defect in the pathogenesis of some CD patients.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Acetylmuramyl-Alanyl-Isoglutamine / metabolism
  • Crohn Disease / genetics
  • Crohn Disease / metabolism*
  • Cytokines / metabolism
  • DNA Primers
  • Electrophoretic Mobility Shift Assay
  • Gene Expression Regulation*
  • Humans
  • Immunity, Innate / genetics
  • Interleukin-1 / metabolism*
  • Interleukin-8 / metabolism*
  • Intracellular Signaling Peptides and Proteins / genetics*
  • Leukocytes, Mononuclear / metabolism
  • Microarray Analysis
  • Mutation / genetics*
  • Nod2 Signaling Adaptor Protein
  • Reverse Transcriptase Polymerase Chain Reaction
  • Signal Transduction / genetics*
  • Tumor Necrosis Factor-alpha / metabolism


  • Cytokines
  • DNA Primers
  • Interleukin-1
  • Interleukin-8
  • Intracellular Signaling Peptides and Proteins
  • NOD2 protein, human
  • Nod2 Signaling Adaptor Protein
  • Tumor Necrosis Factor-alpha
  • Acetylmuramyl-Alanyl-Isoglutamine