Scalable bioprocess for controlled differentiation of embryonic stem cells to hematopoietic progenitors
Bibliographic record
Abstract
Therapeutic application of pluripotent embryonic stem (ES) cells will require advances in cell culture technology that improve our ability to generate target cells. Control of the cell culture environment is of critical importance, as cell fate decisions can be influenced by cell extrinsic factors like cell-cell interactions, soluble cytokines, and physicochemical parameters. However current ES cell differentiation culture systems that make use of static tissue culture plates are limited in terms of measurement and control of the culture environment, and scalability of cell production. In this thesis project, a novel method to differentiate ES cells in scalable, controlled stirred suspension culture was developed. ES cells were differentiated as three dimensional tissue structures termed embryoid bodies (EBs). Successful EB formation was found to depend on the aggregation of ES cells. However cell aggregation, beyond that required for EB formation, was found to impair cell yield. E-cadherin, a cell-cell adhesion molecule, was important in this process. To control cell aggregation, ES cells were encapsulated within agarose microcapsules. ES cells within the same capsule were permitted to aggregate and induce EB formation, but the surrounding agarose matrix prevented developing EBs from contacting and agglomerating with one another. This approach permitted efficient EB formation, growth, and differentiation in stirred culture. The ability to measure and control the culture environment in stirred suspension bioreactors makes them a valuable tool for investigating exogenous factors and optimizing conditions for target cell generation. Physicochemical factors like oxygen tension and pH are of particular interest because they can influence cell proliferation and differentiation in a cost effective way. Using the novel stirred suspension culture system, the role of oxygen tension in hematopoietic cell generation was investigated. Hematopoeitic progenitor generation was optimal at 4% oxygen tension. The mechanism of hypoxia-enhanced hematopoietic progenitor generation was investigated. By comparing the function and expression of VEGF and its receptors VEGFR1 and VEGFR2 under normoxic and hypoxic culture, activation of VEGFR2 by VEGF was found to have both enhancing and inhibiting effects, depending on the stage of development. VEGFR1, secreted in a soluble form, was found to mediate VEGFR2 signaling by competitively binding VEGF.
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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 | 0.001 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| 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".