Serum-free human MSC medium supports consistency in human but not in equine adipose-derived multipotent mesenchymal stromal cell culture.
Abstract: For clinical applications of multipotent mesenchymal stromal cells (MSCs), serum-free culture is preferable to standardize cell products and prevent contamination with pathogens. In contrast to human MSCs, knowledge on serum-free culture of large animal MSCs is limited, despite its relevance for preclinical studies and development of veterinary cellular therapeutics. This study aimed to evaluate the suitability of a commercially available serum-free human MSC medium for culturing equine adipose-derived MSCs in comparison with human adipose MSCs. Enzyme-free isolation by explant technique and expansion of equine and human cells in the serum-free medium were feasible. However, serum-free culture altered the morphology and complicated handling of equine MSCs, with cell aggregation and spontaneous detachment of multilayers, compared to culture in standard medium supplemented with fetal bovine serum. Furthermore, proliferation and the surface immunophenotype of equine cells were more variable compared to the controls and appeared to depend on the lot of the serum-free medium. Particularly the expression of CD90 was different between experimental groups (P < 0.05), with lower percentages of CD90 cells found in equine MSC samples cultured in serum-free medium (5.21-83.40%) compared to standard medium (86.20-99.50%). Additionally, small subpopulations expressing MSC exclusion markers such as CD14 (0.28-11.60%), CD34 (0.00-9.87%), CD45 (0.35-10.50%), or MHCII (0.00-3.67%) were found in equine samples after serum-free culture. In contrast, human samples displayed a more consistent morphology and a consistent CD29 (98.60-99.90%), CD73 (94.60-98.40%), CD90 (99.60-99.90%), and CD105 (97.40-99.80%) immunophenotype after culture in serum-free medium. The obtained data demonstrate that the serum-free medium was suitable for human MSC culture but did not lead to entirely satisfactory results in equine MSCs. This underlines that requirements regarding serum-free culture conditions are species-specific, indicating a need for serum-free media to be optimized for MSCs from relevant animal species. © 2017 International Society for Advancement of Cytometry.
© 2017 International Society for Advancement of Cytometry.
Publication Date: 2017-09-19 PubMed ID: 28926198DOI: 10.1002/cyto.a.23240Google Scholar: Lookup
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- Journal Article
Summary
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The research article involves a study conducted to assess the suitability of a commercially available serum-free human mesenchymal stromal cells (MSCs) medium for growing equine adipose-derived MSCs. The study found that while the serum-free medium supports consistent human MSCs cultures, it does not yield the same predictable results for equine MSCs, suggesting that the serum-free culture conditions requirements vary between species.
Objective
- The objective of the research was to evaluate the effectiveness of a specific serum-free human MSCs medium in culturing equine adipose-derived MSCs.
Methodology
- The researchers applied an enzyme-free isolation process using the explant technique. They then expanded equine and human cells in the serum-free medium.
- Both equine and human MSC cultures were compared to see if there were consistencies or disparities.
Findings
- The study found that the serum-free medium could alter the shape of equine MSCs and made their handling more complex, such as their tendency to aggregate and the spontaneous detachment of cell multilayers. These effects were not seen in human MSCs.
- It was also observed that the proliferation and the surface immunophenotype of equine cells were more inconsistent compared to the controls. This variability appeared to be influenced by the lot of the serum-free medium used.
- Equine MSCs produced in serum-free medium also showed reduced expression of CD90 and inconsistency in expressing MSC exclusion markers compared to those produced in standard medium supplemented with fetal bovine serum.
- On the other hand, human MSCs cultured in serum-free medium exhibited stable cell morphology and consistent immunophenotype across various markers.
Conclusion
- In conclusion, the study suggests that the serum-free medium is suitable for human MSCs culture but not for equine MSCs.
- The study proposes that serum-free culture conditions are species-specific and that there is a need for serum-free media to be optimized for MSCs from relevant animal species.
Cite This Article
APA
Schubert S, Brehm W, Hillmann A, Burk J.
(2017).
Serum-free human MSC medium supports consistency in human but not in equine adipose-derived multipotent mesenchymal stromal cell culture.
Cytometry A, 93(1), 60-72.
https://doi.org/10.1002/cyto.a.23240 Publication
Researcher Affiliations
- Saxonian Incubator for Clinical Translation (SIKT), University of Leipzig, Philipp-Rosenthal-Straße 55, Leipzig 04103, Germany.
- Faculty of Veterinary Medicine, Institute of Physiology, University of Leipzig, An den Tierkliniken 7, Leipzig 04103, Germany.
- Saxonian Incubator for Clinical Translation (SIKT), University of Leipzig, Philipp-Rosenthal-Straße 55, Leipzig 04103, Germany.
- Faculty of Veterinary Medicine, Large Animal Clinic for Surgery, University of Leipzig, An den Tierkliniken 21, Leipzig 04103, Germany.
- Saxonian Incubator for Clinical Translation (SIKT), University of Leipzig, Philipp-Rosenthal-Straße 55, Leipzig 04103, Germany.
- Saxonian Incubator for Clinical Translation (SIKT), University of Leipzig, Philipp-Rosenthal-Straße 55, Leipzig 04103, Germany.
- Faculty of Veterinary Medicine, Institute of Physiology, University of Leipzig, An den Tierkliniken 7, Leipzig 04103, Germany.
MeSH Terms
- Adipose Tissue / cytology
- Animals
- Cell Culture Techniques / methods
- Cell Differentiation
- Cell Lineage
- Cell Proliferation
- Cell Separation
- Culture Media, Serum-Free
- Flow Cytometry
- Horses
- Humans
- Immunophenotyping
- Lipopolysaccharide Receptors / metabolism
- Mesenchymal Stem Cells / cytology
- Mesenchymal Stem Cells / immunology
- Real-Time Polymerase Chain Reaction
- Species Specificity
- Thy-1 Antigens / metabolism
Citations
This article has been cited 12 times.- Sahoo A, Damala M, Jaffet J, Prasad D, Basu S, Singh V. Expansion and characterization of human limbus-derived stromal/mesenchymal stem cells in xeno-free medium for therapeutic applications. Stem Cell Res Ther 2023 Apr 15;14(1):89.
- Even KM, Gaesser AM, Ciamillo SA, Linardi RL, Ortved KF. Comparing the immunomodulatory properties of equine BM-MSCs culture expanded in autologous platelet lysate, pooled platelet lysate, equine serum and fetal bovine serum supplemented culture media. Front Vet Sci 2022;9:958724.
- Pilgrim CR, McCahill KA, Rops JG, Dufour JM, Russell KA, Koch TG. A Review of Fetal Bovine Serum in the Culture of Mesenchymal Stromal Cells and Potential Alternatives for Veterinary Medicine. Front Vet Sci 2022;9:859025.
- Nguyen LT, Tran NT, Than UTT, Nguyen MQ, Tran AM, Do PTX, Chu TT, Nguyen TD, Bui AV, Ngo TA, Hoang VT, Hoang NTM. Optimization of human umbilical cord blood-derived mesenchymal stem cell isolation and culture methods in serum- and xeno-free conditions. Stem Cell Res Ther 2022 Jan 10;13(1):15.
- Burk J, Melzer M, Hagen A, Lips KS, Trinkaus K, Nimptsch A, Leopold J. Phospholipid Profiles for Phenotypic Characterization of Adipose-Derived Multipotent Mesenchymal Stromal Cells. Front Cell Dev Biol 2021;9:784405.
- Melzer M, Schubert S, Müller SF, Geyer J, Hagen A, Niebert S, Burk J. Rho/ROCK Inhibition Promotes TGF-β3-Induced Tenogenic Differentiation in Mesenchymal Stromal Cells. Stem Cells Int 2021;2021:8284690.
- Pezzanite L, Chow L, Griffenhagen G, Dow S, Goodrich L. Impact of Three Different Serum Sources on Functional Properties of Equine Mesenchymal Stromal Cells. Front Vet Sci 2021;8:634064.
- Bhat S, Viswanathan P, Chandanala S, Prasanna SJ, Seetharam RN. Expansion and characterization of bone marrow derived human mesenchymal stromal cells in serum-free conditions. Sci Rep 2021 Feb 9;11(1):3403.
- Hagen A, Lehmann H, Aurich S, Bauer N, Melzer M, Moellerberndt J, Patané V, Schnabel CL, Burk J. Scalable Production of Equine Platelet Lysate for Multipotent Mesenchymal Stromal Cell Culture. Front Bioeng Biotechnol 2020;8:613621.
- Gugjoo MB, Hussain S, Amarpal, Shah RA, Dhama K. Mesenchymal Stem Cell-Mediated Immuno-Modulatory and Anti- Inflammatory Mechanisms in Immune and Allergic Disorders. Recent Pat Inflamm Allergy Drug Discov 2020;14(1):3-14.
- Melzer M, Niebert S, Heimann M, Ullm F, Pompe T, Scheiner-Bobis G, Burk J. Differential Smad2/3 linker phosphorylation is a crosstalk mechanism of Rho/ROCK and canonical TGF-β3 signaling in tenogenic differentiation. Sci Rep 2024 May 6;14(1):10393.
- Burk J, Wittenberg-Voges L, Schubert S, Horstmeier C, Brehm W, Geburek F. Treatment of Naturally Occurring Tendon Disease with Allogeneic Multipotent Mesenchymal Stromal Cells: A Randomized, Controlled, Triple-Blinded Pilot Study in Horses. Cells 2023 Oct 24;12(21).
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