Atmospheric Oxygen Inhibits Growth and Differentiation of Marrow‐Derived Mouse Mesenchymal Stem Cells via a p53‐Dependent Mechanism: Implications for Long‐Term Culture Expansion
Open accessAbstract
Large scale expansion of human mesenchymal stem cells (MSCs) is routinely performed for clinical therapy. In contrast, developing protocols for large scale expansion of primary mouse MSCs has been more difficult due to unique aspects of rodent biology. Currently, established methods to isolate mouse MSCs select for rapidly dividing subpopulations that emerge from bone marrow cultures following long-term (months) expansion in atmospheric oxygen. Herein, we demonstrate that exposure to atmospheric oxygen rapidly induced p53, TOP2A, and BCL2-associated X protein (BAX) expression and mitochondrial reactive oxygen species (ROS) generation in primary mouse MSCs resulting in oxidative stress, reduced cell viability, and inhibition of cell proliferation. Alternatively, procurement and culture in 5% oxygen supported more prolific expansion of the CD45(-ve) /CD44(+ve) cell fraction in marrow, produced increased MSC yields following immunodepletion, and supported sustained MSC growth resulting in a 2,300-fold increase in cumulative cell yield by fourth passage. MSCs cultured in 5% oxygen also exhibited enhanced trilineage differentiation. The oxygen-induced stress response was dependent upon p53 since siRNA-mediated knockdown of p53 in wild-type cells or exposure of p53(-/-) MSCs to atmospheric oxygen failed to induce ROS generation, reduce viability, or arrest cell growth. These data indicate that long-term culture expansion of mouse MSCs in atmospheric oxygen selects for clones with absent or impaired p53 function, which allows cells to escape oxygen-induced growth inhibition. In contrast, expansion in 5% oxygen generates large numbers of primary mouse MSCs that retain sensitivity to atmospheric oxygen, and therefore a functional p53 protein, even after long-term expansion in vitro.
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Typically, MSCs are expanded under atmospheric (20.9%) oxygen tension (normoxia), a process that has been shown to impair their proliferative and differentiation capacity.
“Typically, however, MSCs are expanded under atmospheric (20.9%) oxygen tension (i.e., normoxia), a process that has been shown to impair their proliferative and differentiation capacity [25, 26] and does not properly recapitulate or prepare them for the in vivo environment.”
From Short-term physiological hypoxia potentiates the therapeutic function of mesenchymal stem cells (2018) · cites this paper as [26]
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Short-term physiological hypoxia potentiates the therapeutic function of mesenchymal stem cells
Antebi, B., Rodriguez, L. A., Walker, K. P., Asher, A. M., Kamucheka, R. M., Alvarado, L., Mohammadipoor, A., & Cancio, L. C. (2018) · Stem Cell Research & Therapy · doi:10.1186/s13287-018-1007-x · Cited by 137 · see citing works
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- Authors
- Boregowda, S. V., Krishnappa, V., Chambers, J. W., Lograsso, P. V., Lai, W., Ortiz, L. A., & Phinney, D. G.
- Affiliations
- Scripps Research Institute (US)
- National Health Research Institutes (TW)
- University of Pittsburgh (US)
- Corresponding author
- Phinney, Donald G.
- Venue
- STEM CELLS
- Publisher
- Oxford University Press (OUP)
- Published
- 2012-04-09
- Volume
- 30
- Issue
- 5
- Pages
- 975-987
- ISSN
- 1066-5099 (ISSN-L)
- 1549-4918
- PMC
- PMC3683654
- OpenAlex
- W2069668624
- Cited by
- 114
- References
- 53
- Citations by year
- 2025: 3 · 2024: 4 · 2023: 9 · 2022: 7 · 2021: 8 · 2020: 7 · 2019: 7 · 2018: 13 · 2017: 11 · 2016: 7 · 2015: 19 · 2014: 10 · 2013: 3 · 2012: 6
- FWCI
- 5.80
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Licence
- Locations
- Stem Cells (publishedVersion)
- PubMed (publishedVersion)
- Europe PMC (PubMed Central) (acceptedVersion)
- PubMed Central (acceptedVersion)
- Funders
- National Institute of Health (R01 NS052301-01A2)
- Topics
- Health Sciences › Medicine › Genetics › Mesenchymal stem cell research
- Health Sciences › Medicine › Hematology › Hematopoietic Stem Cell Transplantation
- Health Sciences › Medicine › Physiology › Telomeres, Telomerase, and Senescence
- Keywords
- mesenchymal stem cells, atmospheric oxygen, p53, oxidative stress, reactive oxygen species, mitochondrial reactive oxygen species, cell proliferation, cell differentiation, trilineage differentiation, bone marrow, TOP2A, Bax, stem cell selection
- MeSH terms
- Poly-ADP-Ribose Binding Proteins, Animals, Antigens, Neoplasm, Bone Marrow Cells, Cell Differentiation, Cells, Cultured, DNA Topoisomerases, Type II, DNA-Binding Proteins, Humans, Oxygen, Tumor Suppressor Protein p53, Reactive Oxygen Species, Mice, Knockout, Cell Proliferation, bcl-2-Associated X Protein, Mice, Gene Knockdown Techniques, Mesenchymal Stem Cells
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