No simple way to averaging out: Pooled mesenchymal stromal cells do not reflect average donor characteristics

Title (eng)
No simple way to averaging out: Pooled mesenchymal stromal cells do not reflect average donor characteristics
Author
Dea Kukaj
Author
Sabine Niebert
Author
Christoph Biehl
Author
Ursula Reichart
Author
Christiane Schueler
Author
Janina Burk
Abstract (eng)
Mesenchymal stromal cells (MSCs) are promising candidates for numerous regenerative therapies. Still, clinical translation is complicated by the heterogeneity of MSCs and related shortcomings in preclinical research. Pooling MSCs from multiple donors is increasingly being advocated as an effective way to mitigate donor variability. However, it remains unclear whether the range of individual cell characteristics is equally reflected in pooled cultures, or if pooling rather leads to a homogenized cell population dominated by the fittest donor, which would lead to skewed results. This study investigates whether MSC pools are functionally representative for their respective donor MSCs and whether dominant donors emerge over time. MSCs from nine human donors were categorized into low-, middle-, and high-fitness groups. Individual MSCs were then pooled according to their fitness groups, complemented by a mixed-fitness pool. Functional assays for proliferation, metabolic activity, differentiation, migration and senescence were performed to evaluate the pools versus the individual MSCs. Donor representation within pools was tracked using fluorescence microscopy and qPCR. The high-fitness pool, as well as its individual donor MSCs, displayed the most rapid proliferation and highest metabolic activity. However, while for proliferation, the pool data aligned well with the individual donor data, all other assays revealed discrepancies between the pooled cultures and individual donor cells. Interestingly, particularly the mixed fitness pool showed inferior metabolic activity and differentiation potential in comparison with the respective individual donor MSCs. Cell tracking showed that over one passage, even pools composed of donors with similar cell fitness became dominated by the donor with the highest cellular fitness. The discrepancy between pooled and individual donor data emphasizes the importance of biological replicates to capture donor variation and ensure that MSC research reflects natural diversity.
Keywords (eng)
Mesenchymal Stromal CellsHuman MSCsAdipose-derivedDonor VariabilityHeterogeneityCellular FitnessCell PoolingFunctional CharacterizationCell Tracking
Type (eng)
journal article
Language
[eng]
Persistent identifier
https://phaidra.vetmeduni.ac.at/o:5005
DOI
10.1016/j.reth.2025.09.012
Is in series
Title (eng)
Regenerative Therapy
Volume
30
ISSN
2352-3204
Issued
2025
Number of pages
11
Publication
Elsevier
Version type (eng)
version of record (published version)
Date issued
2025
Access rights (eng)
open access
License
CC BY 4.0 International
Rights statement (eng)
© 2025 The Author(s)