Regulation of IL-8 and IL-1beta expression in Crohn's disease associated NOD2/CARD15 mutations.
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.
Li
Jing
J
The Martin Boyer Laboratories, Gastroenterology Section, Department of Medicine, The University of Chicago, IL 60637, USA.
Moran
Thomas
T
Swanson
Eric
E
Julian
Christina
C
Harris
Jeremy
J
Bonen
Denise K
DK
Hedl
Matija
M
Nicolae
Dan L
DL
Abraham
Clara
C
Cho
Judy H
JH
D000119:Acetylmuramyl-Alanyl-Isoglutamine / Q000378:metabolism; D003424:Crohn Disease / Q000235:genetics / Q000378:metabolism*; D016207:Cytokines / Q000378:metabolism; D017931:DNA Primers; D024202:Electrophoretic Mobility Shift Assay; D005786:Gene Expression Regulation; D006801:Humans; D007113:Immunity, Innate / Q000235:genetics; D007375:Interleukin-1 / Q000378:metabolism*; D016209:Interleukin-8 / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics*; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D046228:Microarray Analysis; D009154:Mutation / Q000235:genetics*; D053473:Nod2 Signaling Adaptor Protein; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000235:genetics*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D016207:Cytokines; D017931:DNA Primers; D007375:Interleukin-1; D016209:Interleukin-8; D047908:Intracellular Signaling Peptides and Proteins; C429018:NOD2 protein, human; D053473:Nod2 Signaling Adaptor Protein; D014409:Tumor Necrosis Factor-alpha; D000119:Acetylmuramyl-Alanyl-Isoglutamine