Endogenously produced protein regulators provide feedback signals that regulate the ex vivo expansion of human hematopoietic stem and progenitor cells
Bibliographic record
Abstract
The absence of effective strategies for the ex vivo expansion of human blood (hematopoietic) stem cells (HSCs) limits the development of many stem cell-based therapies. The focus of this study was to investigate in vitro processes responsible for regulating HSC proliferation and to utilize this information in the design of a robust methodology for expanding HSCs. Herein we show the existence of a negative feedback control mechanism whereby differentiated blood cells secrete soluble factors that limit HSC expansion. We demonstrate that global culture manipulation strategies including subpopulation selection and media dilution/exchange modulate this feedback mechanism to enable stem cell expansion. Using this approach, we were able to generate increased numbers of long term culture-initiating cells (LTC-ICs; 14.6-fold), rapid non-obese diabetic/severe combined immunodeficient (NOD/SCID) repopulating cells (R-SRCs; 12.1-fold), and long-term NOD/SCID repopulating cells (LT-SRCs; 5.2-fold), compared with input; outputs significantly higher than those obtained in unmanipulated control cultures. In order to enable this culture methodology for therapeutic applications, a closed-system bioprocess was designed which incorporated in-line subpopulation selection and media dilution/exchange processes. Experiments showed that the bioprocess was able to expand colony forming cells (CFCs), LTC-ICs and LT-SRCs in a manner consistent with results using standard tissue culture dishes. Studies to optimize the bioprocess operating conditions were also performed. In these studies it was found that non-specific cell loss, which occurred during the subpopulation selection step, could be decreased by increasing flow rate through the selection element. The ability to decrease cell loss is important since it should facilitate higher expansions of hematopoietic stem and progenitor cells within the bioprocess. Furthermore, optimization of the subpopulation selection process through the identification of specific inhibitory factor secreting cells may further augment the measured expansions. To initiate this goal, the gel microdrop (GMD) assay was developed as a means to measure protein secretion from individual cells in the context of cell surface phenotype. The development and use of the GMD assay represents the first step in the design of a second generation bioprocess which should have an even greater capacity for the ex vivo expansion of HSCs.
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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 | 0.000 |
| 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 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".