Primary colonospheres maintain stem cell‐like key features after cryopreservation
Bibliographic record
Abstract
The development of efficient and repeatable protocols for biobanking and prolonged storage of cancer stem cells (CSCs), with minimum alterations in biological function, is valuable and desired, particularly for retrospective analysis and clinical applications. In particular, data regarding the effect of cryopreservation on CSCs's functional features is scarce. In this regard, few studies have been shown that 3D spheroid structures, which enriched for CSCs, can keep their biological phenotype and genetic profiles. Here, for the first time, we present data on cryopreservation of CT-26 colonospheres, with the focus on essential stem cell-like properties after thawing. Tumor biopsy-derived colonospheres were frozen in standard freezing media (90% fetal bovine serum + 10% dimethyl sulfoxide) and stored in liquid nitrogen for 10 months. Then, cryopreservation effect on preservation of CSCs-related features was verified using real-time polymerase chain reaction for evaluation of stemness genes and flow cytometry for the putative colorectal CSC surface biomarkers. The self-renewal capacity of thawed spheres was also compared with their fresh counterparts using serial formation assay. Finally, tumorigenic capacity of both groups was evaluated in immunocompetence mouse model. Our data indicated that postthawed colonospheres had high viability without drastic alteration in biological and structural features and maintained self-renewal potential after sequential passages. Real-time analysis showed that both fresh and frozen colonospheres displayed similar expression pattern for key stemness genes: SOX2 and OCT4. Cryopreserved spheroids expressed CD133, CD166, and DCLK1 CSCs surface biomarkers at elevated levels when compared with parental as non-cryopreserved counterparts. Our electron scanning microscopy micrographs clearly demonstrated that postthawed colonospheres retain their integrity and cell surface morphology and characteristics. We also found that both fresh and frozen spheroids were equally tumorigenic. This study represented an effective strategy for reliable storage of intact CT-26 colonospheres; this can provide researchers with a functionally reliable repository of murine colorectal CSCs for their future CSCs projects.
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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 | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 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) | 0.001 | 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".