Assessment of the Developmental Potential of Rat Embryonic Stem-Like (ES-Like) Cells by Diploid and Tetraploid Embryo Aggregation Confirms Their In Vivo Multipotency.
Bibliographic record
Abstract
The rat is a model of major importance to biomedical research. The existence of genuine embryonic stem (ES) cells in the rat is highly anticipated and desirable because it would allow gene-targeting technology (i.e. knockout) in this species and permit further elucidation of gene function. At present, this powerful technology is only available in mice, and in fact, derivation of ES cells in the rat has remained elusive for many years. However, recently we have demonstrated that rat ES-like cells could be efficiently derived and cultured in vitro for extended periods while maintaining pluripotent cell markers such as alkaline phosphatase, stage-specific embryonic antigen-1, and both mRNA and protein expression of the transcription factor Oct4. These rat ES-like cells were transfected with a reporter gene, injected into diploid host blastocysts, and transferred to pseudopregnant females. The results obtained demonstrated that rat ES-like cells can contribute to multiple developing extraembryonic tissues of both midgestation and term fetuses. However, their in vivo contribution appears limited only to the extraembryonic tissues, as their presence in tissues of the embryo proper could not be detected. We hypothesized that rat ES-like cells could contribute to a wider range of tissues if host embryos at an earlier developmental stage than the blastocyst were used for testing the developmental potential. To this end, we adapted two assays frequently used in mice to generate embryonic chimeras, that is, diploid and tetraploid aggregation. The specific objectives were to: 1) develop a robust in vitro embryo culture system 2) generate tetraploid embryos 3) carry out diploid (2n<->ES) and tetraploid (4n<->ES) aggregations and compare the development rates to the morula \ blastocyst stage 4) determine the implantation rates following embryo transfer and 5) examine the contribution of the rat ES-like cells to resulting fetuses. The results obtained showed efficient development of control embryos to the morula \ blastocyst stage, indicating an efficient culture system. Tetraploid embryos were very successfully generated. No difference was observed in the rates of development to the morula \ blastocyst stage between controls, 2n<->ES, and 4n<->ES embryo aggregations. The implantation rates were similar in all groups, suggesting that the presence of rat ES-like cells in aggregates does not adversely affect implantation following embryo transfer. Importantly, these rat ES-like cells were found to have contributed in vivo to the extraembryonic tissues at midgestation, confirming similar results obtained previously using blastocyst injection. In conclusion, for the first time, we have shown that embryo aggregations are suitable to determine the developmental potential of rat ES-like cells, suggesting that such aggregations can be used to screen for pluripotent rat ES-like cell lines. This research therefore constitutes another step forward in the search for genuine rat ES cells. Supported by a Natural Sciences and Engineering Research Council of Canada Industrial Postgraduate Scholarship to S.-P. Demers and by Clonagen inc.
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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 | 0.001 | 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 | 0.001 |
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".