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Cryopreservation of rat MSCs by use of a programmed freezer with magnetic field
Shunichi Kojima1, Masato Kaku, Toshitsugu Kawata
1Department of Orthodontics and Craniofacial Developmental Biology, Hiroshima University, Graduate School of Biomedical Science, Japan.
Abstract:
Mesenchymal stem cells (MSCs) can be used for the regeneration of various tissues and cryopreservation of MSCs is so important for regenerative medicine. The purpose of this study was to evaluate the influences of cryopreservation on MSCs by use of a programmed freezer with a magnetic field (CAS freezer). MSCs were isolated from bone marrow of rat femora. The cells were frozen by a CAS freezer with 10% dimethyl sulfoxide (Me2SO) and cryopreserved for 7 days at a temperature of -150 °C. Immediately after thawing, the number of survived cells was counted. The cell proliferation also examined after 48 h culture. Next, MSCs were frozen by two different freezers; CAS freezer and a conventional programmed freezer without magnetic field. Then, osteogenic and adipogenic differentiations of cryopreserved cells were examined. As a result, survival and proliferation rates of MSCs were significantly higher in CAS freezer than in the non-magnetic freezer. Alizarin positive reaction, large amount of calcium quantification, and greater alkaline phosphatase activity were shown in both the non-cryopreserved and CAS groups after osteogenic differentiation. Moreover, Oil Red O staining positive reaction and high amount of PPARγ and FABP4 mRNAs were shown in both the non-cryopreserved and CAS groups after adipogenic differentiation. From these findings, it is shown that a CAS freezer can maintain high survival and proliferation rates of MSCs and maintain both adipogenic and osteogenic differentiation abilities. It is thus concluded that CAS freezer is available for cryopreservation of MSCs, which can be applied to various tissue regeneration.
Insights
Cryopreservation using a magnetic field (CAS freezer) significantly enhances mesenchymal stem cell (MSC) survival and proliferation. This method preserves MSC differentiation potential for regenerative medicine applications.
Area of Science:
- Biomedical Engineering
- Cell Biology
- Regenerative Medicine
Background:
- Mesenchymal stem cells (MSCs) are crucial for tissue regeneration.
- Effective cryopreservation is vital for the clinical application of MSCs in regenerative medicine.
Purpose of the Study:
- To evaluate the impact of cryopreservation using a novel magnetic field programmed freezer (CAS freezer) on MSC viability and function.
- To compare the efficacy of the CAS freezer against a conventional programmed freezer.
Main Methods:
- Rat bone marrow-derived MSCs were cryopreserved using the CAS freezer with 10% dimethyl sulfoxide (Me2SO) at -150 °C for 7 days.
- Cell survival and proliferation were assessed post-thaw.
- Osteogenic and adipogenic differentiation potentials of MSCs cryopreserved with the CAS freezer and a conventional freezer were examined.
Main Results:
- The CAS freezer demonstrated significantly higher MSC survival and proliferation rates compared to the conventional freezer.
- MSCs cryopreserved with the CAS freezer retained robust osteogenic differentiation capacity, indicated by Alizarin Red staining and alkaline phosphatase activity.
- Adipogenic differentiation was also well-preserved in CAS freezer-preserved MSCs, evidenced by Oil Red O staining and gene expression of PPARγ and FABP4.
Conclusions:
- The CAS freezer effectively preserves high survival and proliferation rates of mesenchymal stem cells.
- Cryopreservation with the CAS freezer maintains the osteogenic and adipogenic differentiation abilities of MSCs.
- The CAS freezer is a viable technology for MSC cryopreservation, supporting its use in regenerative medicine.
