NOVEL REGULATION OF A PAR2‐MEDIATED CELLULAR MIGRATION PROGRAM BY PRO‐INFLAMMATORY CYTOKINES
Bibliographic record
Abstract
BACKGROUND Mucosal healing is the gold standard for IBD therapy. However, the mechanisms that determine mucosal healing are complex and need to be better understood to explain why many patients fail to achieve this key goal of therapy. The inflammatory environment is characterized by the presence of several serine proteases that signal through the protease‐activated receptors (PARs). PAR2 is ubiquitously expressed in the epithelial cells of the intestinal mucosa and activation serves a protective role consistent with host defence and repair. While our preliminary findings indicate that activation of PAR2 enhances wound closure, we do not know the cellular mechanisms that underlie this response. We hypothesize that activation of PAR2 induces a cellular migration program in intestinal epithelial cells that enhances mucosal healing in the inflammatory milieu . METHODS Circular wounds were introduced in T84 colonic epithelial cells. Wounded monolayers were treated with the PAR2 activating peptide, 2‐furoyl‐LIGRLO (2fLI, 5 μM), or the control reverse‐sequence peptide, 2‐furoyl‐OLRGIL (2fO, 5 μM), and live cell imaging was utilized to record wound closure over a 24 hr period. In other experiments, a cytokine cocktail consisting of IFNγ (10 ng/mL) and TNFα (10 ng/mL), alone or in combination with 2fLI, was applied to wounded monolayers and wound closure assessed. Intracellular effects of PAR2 activation and cytokine treatment were evaluated by measuring phosphorylation of key canonical signaling proteins using the MSD multi‐plex ELISA array system. EdU staining, with or without Mitomycin C (MMC), an inhibitor of proliferation, was used to determine the effect of cellular proliferation on wound closure. RESULTS PAR2 activation by 2fLI promoted wound closure in a concentration‐dependent manner (0.1 μM – 10 μM) compared to 2fO controls at the 12 and 24 hr timepoints. Interestingly, co‐stimulation with 2fLI and the cytokine cocktail resulted in an enhanced wound closure response at the 12 and 24 hr timepoints that was significantly greater than individual treatments ( p <0.001). Co‐stimulation activated the downstream intracellular targets ERK1/2 and JNK compared to 2fO ( p <0.001), but this activation was not greater than stimulation with 2fLI or cocktail alone, suggesting that MAP kinase activation was not responsible for the enhanced effect. MMC pre‐treatment partially reduced wound closure caused by co‐stimulation with 2fLI/IFNγ/TNFα, indicating that the enhanced wound healing effect was due in part to increased proliferation. CONCLUSIONS These data suggest that both PAR2 activation and cytokine treatment promote wound closure in vitro through both enhanced cell migration and proliferation. The enhanced wound healing response to PAR2 activation in the presence of pro‐inflammatory cytokines suggests that the inflammatory milieu is necessary to initiate a proper wound healing response. Support or Funding Information Canadian Institutes of Health Research (CIHR) This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 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) | 0.002 | 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 source (direct Gemma or distilled Codex), 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".