Investigating the interaction between the intestinal epithelium and group-3 innate lymphoid cells in response to Clostridioides difficile toxins
Bibliographic record
Abstract
Hypothesis: Upon disruption of the colonic epithelium, Clostridioides difficile toxins A and B (TcdAB) are thought to signal through to underlying immune cells. Amongst these are group-3 innate lymphoid cells (ILC3s), which modulate pathogen defense and tissue repair largely through IL-22 secretion. We hypothesize that interactions between epithelial cells and ILC3s modulate effects on the colonic epithelium in response to TcdAB. Methodology: Confluent, 2-dimensional, epithelial monolayers established from mouse colonoids were treated apically with TcdAB (0.001 micrograms per millilitre (μg/mL)). Transepithelial electrical resistance (TER) was measured over 6 hours to determine TcdAB-induced permeability changes. To assess ILC3s impact on epithelial response to TcdAB, MNK-3 cells were co-cultured underlying monolayers for 2 days before TcdAB treatment. To assess MNK-3 IL-22 production in monolayer co-cultures, IL-22 was measured by ELISA in basolateral media samples. Results: TcdAB significantly reduced monolayer TER. TER response between monocultured monolayers and those co-cultured with MNK-3 cells did not differ. Starting TER varied significantly (252-2039 Ω/cm 2 ) thus, we hypothesized that starting TER may modulate effects of MNK-3 co-culture on TcdAB response. To test this, data were stratified by starting TER (<900 or >900 Ω/cm 2 ) and starting TER did not influence the effect of MNK-3 co-culture on permeability. Co-culture did not affect MNK3 cell viability. IL-22 was not detected in co-cultures, suggesting that culture of MNK-3 cells in organoid media may inhibit the ability of these cells to secrete IL-22. To test this, we observed IL-22 production from MNK-3 cells cultured in monolayer media vs. MNK-3 media alone, followed by treatment with TcdAB (0.001μg/mL) or with IL-23 (50 nanograms (ng)/mL and 10ng/mL). IL-23 treatment elicited significant IL-22 production that did not differ between MNK-3 cells cultured in monolayer media and those cultured in MNK-3 media, indicating normal MNK-3 cell function. In contrast to previous reports, direct treatment of MNK-3 cells with TcdAB did not result in increased IL-22. Conclusions: TcdAB disrupts epithelial barrier integrity of mouse colon monolayers. Co-culture of primary cell monolayers with MNK-3 cells did not influence epithelial integrity in response to TcdAB. MNK-3 cells can produce IL-22 similarly in both MNK-3 and monolayer media, yet IL-22 is not detected in monolayer-MNK-3 co-cultures. Taken together, these results disprove our hypothesis that IL-22 represents a functional link between TcdAB, disrupted colonic epithelium and ILC3s in our co-culture system. Master's degree research funded by Canadian Institutes for Health Research and William H. Davies Medical Research Scholarship This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.
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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 | 0.002 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.001 |
| 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".