A16 ESOPHAGEAL ORGANOID PROLIFERATION AND DIFFERENTIATION ARE ALTERED BY LOSS OF MSH2
Bibliographic record
Abstract
Abstract Background The stratified epithelium of the esophagus includes Krt15+ basal stem cells that display self-renewing and regenerative capacity, and multipotency. However, the mechanisms that specifically control their functions remain unknown. Interestingly, RNA sequencing and GSEA revealed an enrichment of a gene set associated with DNA repair in Krt15+ cells vs Krt15- cells. We also observed that Msh2 (DNA mismatch repair pathway) is the most significantly upregulated gene in Krt15+ stem cells. Aims To determine the effect of Msh2 loss on self-renewal and differentiation of esophageal organoids. Methods Esophageal epithelial cells were isolated from a wild-type mouse. Using flow cytometry, esophageal Krt15+ (GFP+) and Krt15- (GFP-) cells were sorted from Krt15-CrePR1 (R26mT/mG) mice. All cell populations were grown as organoids and Msh2 was depleted using a CRISPR/Cas9 approach. Impact of Msh2 loss on self-renewal and differentiation in esophageal epithelial organoids was evaluated through organoid formation assays, WST-1 proliferation assays and histological analysis. Results At baseline, organoids depleted for Msh2 formed more poorly differentiated and less well-differentiated organoids than controls. Lower expression of differentiation gene Krt13 was also observed in Msh2-depleted organoids, confirming an altered differentiation pattern. Furthermore, these organoids showed a higher organoid formation rate and proliferation by WST-1 assay, suggesting that self-renewal capacity and viability are increased when Msh2 is depleted. Interestingly, following radiation, organoids depleted for Msh2 showed higher residual levels of p-H2AX (DNA damage marker), suggesting that their capacity to cope with DNA damages is altered. As mentioned above, we previously reported that Msh2 is the most upregulated gene in Krt15+ vs Krt15- cells. Therefore, to determine if Msh2 role is distinct in both populations, we depleted Msh2 in Krt15+ and Krt15- cells-derived organoids. Interestingly, our preliminary results suggest that Msh2 deletion led to increased p-H2AX and decreased Krt13 levels in Krt15+ organoids but not in Krt15- organoids. Conclusions Our results show that Msh2 is potentially a key contributor of esophageal stemness in homeostatic and injured conditions. Funding Agencies CIHRCanada Research Chair
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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 | 0.000 |
| Insufficient payload (model declined to judge) | 0.003 | 0.001 |
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".