O0105 CROHN’S DISEASE AS AN INNATE IMMUNE DEFECT —THE CRITICAL ROLE OF CARD15 IN INTESTINAL ANTIBACTERIAL DEFENSE
Bibliographic record
Abstract
Introduction: Recently, with the discovery of different mutations in the CARD15 gene in a subgroup of patients with ileal Crohn’s disease, a first clue to the understanding of the molecular mechanisms of this disorder was found. CARD15 is an intracel-lular protein functioning as a bacterial sensor via its leucine rich repeat recognizing bacterial muramyl-dipeptide, the minimal signature found in peptidoglycan of gram−/+ or /− bacteria. Methods: Intestinal epithelial cells (HIEC, Caco-2, HT-29) were stimulated by various pro-inflammatory cytokines and the induction of CARD4/CARD15 was monitored by semi-quantiative and quantitative real-time PCR. The functionality of CARD15 was tested by intracellular micro-injection of MDP or MDP-fugene-transfection by means of the activation of NFkB (EMSA, immunofluorescence). WT-CARD15 and mutated CARD15 (3020InsC) were transfected (electroporation method) into HIEC and Caco-2 cells and the response to MDP activation was measured using an NFkB reporter assay. The generation of a pro-inflammatory and anti-bacterial response to MDP-stimulation was compared between WT and mutant transfected enterocytes using a real time PCR approach. Results: Normal, non-transformed intestinal epithelial cells clearly expressed CARD4, but no CARD15 signal was observed. However, pro-inflammatory stimuli induced a high degree of CARD15 expression in an NFkB dependent (TNF) or –independent (IFN) manner. Cytokine-induced CARD15 in enterocytes was functional, since after stimulation with MDP a marked activation of NFkB pathway was observed, as demonstrated by the nuclear translocation of the p65 subunit and by EMSA. Cells not expressing CARD15 were unresponsive to MDP. Using an NFkB reporter assay, we observed that enterocytes transfected with a mutant form of CARD15 failed to activate NFkB. Whereas WT transfectants generated a marked pro-inflammatory response with enhanced anti-bacterial defence activation in response to MDP, mutants were not able to generate either a pro-inflammatory or anti-bacterial (defensin) response. Conclusion: There is clear experimental evidence that in CD patients with a CARD15 mutation, the inducible innate immune system on the intestinal epithelial cell level is defective. In the present study, we observed that in response to bacterial products, normal enterocytes generate a pro-inflammatory and an extremely potent immediate anti-bacterial response. This antibacterial ability is clearly impaired in CD patients rendering them more susceptible to elicit an inappropriate inflammatory reaction to various bacterial stimuli.
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.001 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".