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Updated: May 5, 2026
Megakaryocyte Differentiation and Platelet Formation from Human Cord Blood-derived CD34+ Cells
Published on: December 27, 2017
Absence of micronucleus formation in CHO-K1 cells cultivated in platelet lysate enriched medium
Martina Bernardi1, Valentina Adami2, Elena Albiero1
1Hematology Project Foundation Research Laboratories, Contrà S. Francesco 41, Vicenza, Italy; Advanced Cellular Therapy Laboratory, Department of Cellular Therapy and Hematology, San Bortolo Hospital, Via Rodolfi 37, Vicenza, Italy.
Abstract:
Human platelet lysate (PL) represents an effective substitute of fetal bovine serum (FBS) for mesenchymal stromal cell (MSC) cultivation. Compared to FBS, PL favors MSC proliferation significantly shortening the population doubling time and avoiding the risks related to the use of animal derivatives. Growth factors contained in the platelets are released upon platelet disruption following freezing/thawing cycles or as we have recently described by using ultrasound. We have investigated whether the increased cell proliferation achieved by using PL could induce mitotic stress and whether the potential formation of free radicals during PL production by ultrasound could cause chromosomal instability in mammalian cells. We have applied an image analysis assisted high content screening (HCS) in vitro micronucleus assay in the Chinese Hamster Ovarian K1 (CHO-K1) rodent mammalian cell line. PL was produced by sonication; for the micronucleus assay, CHO-K1 cells were exposed to increasing concentrations of PL. Cytokinesis was blocked by cytochalasin B, nuclei were stained with bisbenzimide and images were acquired and analyzed automatically using an HCS system, both with a 20× and a 10× objective. Our results suggest that growth stimulus induced by the use of PL did not significantly increase micronucleus formation in CHO-K1 cells compared to negative control. Micronucleus testing in conjunction with HCS could represent a valid tool to evaluate the safety of ancillary materials used in the production of cell-based medicinal products.
Insights
Human platelet lysate (PL) enhances mesenchymal stromal cell (MSC) proliferation without inducing significant chromosomal instability. This study used high content screening (HCS) to assess PL safety for cell-based therapies.
Area of Science:
- Cell Biology
- Biotechnology
- Toxicology
Background:
- Human platelet lysate (PL) is a superior alternative to fetal bovine serum (FBS) for culturing mesenchymal stromal cells (MSCs).
- PL promotes MSC proliferation and reduces risks associated with animal-derived products.
- Platelet disruption methods, like sonication, release growth factors but may also generate free radicals.
Purpose of the Study:
- To evaluate if PL-induced MSC proliferation causes mitotic stress.
- To determine if PL production via ultrasound leads to chromosomal instability in mammalian cells.
- To assess the safety of PL as a component in cell-based medicinal products.
Main Methods:
- An in vitro micronucleus assay was performed on Chinese Hamster Ovarian K1 (CHO-K1) cells.
- PL was produced using sonication and cells were exposed to varying PL concentrations.
- High content screening (HCS) with automated image analysis was employed to detect micronucleus formation.
Main Results:
- PL-stimulated growth did not significantly increase micronucleus formation in CHO-K1 cells compared to controls.
- The use of PL did not appear to induce significant chromosomal instability.
- HCS-based micronucleus testing proved effective for evaluating ancillary material safety.
Conclusions:
- PL is a safe alternative to FBS for MSC cultivation, showing no significant genotoxicity.
- High content screening micronucleus assay is a valuable tool for assessing the safety of materials used in cell therapy production.
- Further research can validate PL's role in regenerative medicine manufacturing.
