Equine platelet lysate as an alternative to fetal bovine serum in equine mesenchymal stromal cell culture – too much of a good thing?
Abstract: Multipotent mesenchymal stromal cells (MSC) are often culture-expanded in vitro. Presently, expansion medium (EM) for MSC is supplemented with fetal bovine serum (FBS). However, increasing cost, variable composition and potential risks associated with bovine antigens call for alternatives. Platelet lysate (PL) has shown promise as an alternative supplement. Objective: To determine how equine umbilical cord blood (CB) MSC proliferate in EM enriched with PL or FBS at various concentrations. Methods: Randomised dose escalation study. Methods: Platelet concentrate was generated from 5 equine whole blood samples through a double centrifugation method and standardised to 1 × 10(12) platelets/l prior to a freeze/thaw cycle to produce PL. Pooled PL or pooled FBS was added to EM at concentrations of 5% to 60%. Proliferation of 4 equine CB-MSC cultures was determined after 4 days using a resazurin semiquantitative assay. Results: Cord blood-MSC proliferated with a dose-dependent response with no significant difference found between PL and FBS up to a 30% concentration. Beyond 30%, proliferation fell in the PL-cultured cells, while continued dose-dependent proliferation was noted in the FBS-cultured cells. Despite reduced cell numbers in high PL concentrations, live/dead staining revealed that adherent cells remained viable. Conclusions: Expansion medium enriched with PL can support short-term equine CB-MSC proliferation at conventional culture concentrations. Based on the unexpected suppression of CB-MSC at higher PL concentrations, an in vivo dose study is indicated to investigate if combinational therapies of CB-MSC and platelet-rich plasma are associated with synergistic or antagonistic effect on CB-MSC function.
© 2015 EVJ Ltd.
Publication Date: 2015-05-04 PubMed ID: 25772755DOI: 10.1111/evj.12440Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This research investigated the use of equine platelet lysate as an alternative to fetal bovine serum in growing equine mesenchymal stromal cells in a lab. The study found that the platelet lysate can support these cells’ growth at typical concentrations but discovered an unexpected suppression of cell proliferation at higher concentrations.
Objective of Study
In laboratory settings, a substance called fetal bovine serum (FBS) is typically used to cultivate multipotent mesenchymal stromal cells (MSC). These cells are commonly used in research because of their ability to differentiate into other types of cells. However, using FBS presents several challenges, such as fluctuating costs, inconsistencies in composition, and possible risks related to bovine antigens. Therefore, the researchers aimed to explore alternatives to FBS, specifically platelet lysate (PL), derived from horse blood.
- The primary goal of the study was to examine how equine umbilical cord blood (CB) MSC proliferates in an expansion medium (EM) enriched with either PL or FBS at different concentrations.
Research Methods
The researchers used a randomised dose escalation study design and generated platelet concentrate from five equine whole blood samples. They standardized the platelet count before undergoing a freeze and thaw cycle to produce the PL. They then added pooled PL or FBS to the expansion medium (EM) at concentrations ranging from 5% to 60%.
- The growth of the CB-MSC cultures was then monitored for 4 days using a resazurin semi-quantitative assay, a commonly used method for monitoring cell growth and viability.
Findings
The research found that both PL and FBS supported CB-MSC proliferation in a dose-dependent way, up to a 30% concentration. However, cell growth decreased in the PL-enriched cells beyond a 30% concentration, whereas cell proliferation in the FBS-enriched cells continued to increase with dosage. Notably, even at high PL concentrations, live/dead staining showed that the remaining adherent cells were still viable.
Implications
These results suggest that PL-enriched expansion medium can support short-term equine CB-MSC proliferation at conventional culture concentrations.
- The unexpected suppression of CB-MSCs at higher PL concentrations, however, prompts further investigation. This finding suggests a need for an in vivo study to examine whether combined treatments of CB-MSC and platelet-rich plasma have a positive or negative effect on the function of CB-MSCs.
Cite This Article
APA
Russell KA, Koch TG.
(2015).
Equine platelet lysate as an alternative to fetal bovine serum in equine mesenchymal stromal cell culture – too much of a good thing?
Equine Vet J, 48(2), 261-264.
https://doi.org/10.1111/evj.12440 Publication
Researcher Affiliations
- Department of Biomedical Sciences, University of Guelph, Ontario, Canada.
- Department of Biomedical Sciences, University of Guelph, Ontario, Canada.
- Department of Clinical Studies, Orthopaedic Research Lab, Aarhus University, Copenhagen, Denmark.
MeSH Terms
- Animals
- Blood Platelets / physiology
- Cattle
- Cell Culture Techniques
- Cell Proliferation
- Cell Survival
- Culture Media
- Horses
- Mesenchymal Stem Cells / physiology
Citations
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- Hagen A, Holland H, Brandt VP, Doll CU, Häußler TC, Melzer M, Moellerberndt J, Lehmann H, Burk J. Platelet Lysate for Mesenchymal Stromal Cell Culture in the Canine and Equine Species: Analogous but Not the Same. Animals (Basel) 2022 Jan 13;12(2).
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