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0221U106768,
0121U113738
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Cryobiology profile of 3D mesenchymal stem cells spheroids
Petrenko Oleksandr Yu., д.б.н.
20-12-2021
Institute of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine
The aim of the work was formation of functionally active spheroids based on MSCs and assessment of their viability, metabolic activity and osmotic characteristics. Monolayer and three-dimensional cell culture, light and fluorescence microscopy, cryopreservation, volumetric method and physical-mathematical modeling were used in the study. Key features of MSCs spheroid formation using "hanging drop" method with different seeding concentrations and culture time have been studied. It was found that the concentration of 5 thousand cells / drop and 3 days of cultivation are optimal for the formation of spheroids from viable and functionally active cells. Prolonged culture time led to a decrease in the metabolic activity of MSCs within spheroids. MSCs obtained from spheroids had higher proliferative potential and the ability to form colonies compared to monolayer cells. The possibility of spheroid cryopreservation using a standard protocol developed for cells in suspension was assessed. It was found that slow two-step freezing under the protection of cryoprotectant DMSO resulted in partial disruption of spheroids integrity and death of more than a half of cells in their composition. To improve the spheroids cryopreservation protocol on the basis of empirical data along with physical and mathematical modeling, the filtration and permeability coefficients of spheroids for water molecules and 1 M DMSO were calculated. Obtained results will be used for the development of an effective cryopreservation protocol considering optimal time of exposure of spheroids in cryoprotective solution and cooling rates.
Kovalenko Igor F.
Kovalenko Svitlana Ye.
Mazur Svitlana P.
Revenko Olena B.
Rogulska Olena Yu.
Tykhvynska Olga O.
Trufanova Natalia A.
2021-12-20
Updated: 2025-12-16
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