The Cryoprotecting Effects of Trehalose and Dimethyl Sulfoxide on Alginate-Chitosan Encapsulated Mesenchymal Stem Cells

Trehalose is a promising cryoprotectant for the long-term storage of mesenchymal stem cells (MSCs). Trehalose is relatively safe for the human body and a potential substitute to reduce the toxicity of dimethyl sulfoxide (DMSO), a common cryoprotectant. Previously, we developed an alginate-chitosan microcapsule containing human stem cells. However, the strategy for long-term storage of microencapsulated cells is not established yet. The current study was conducted to evaluate the potencies of trehalose and DMSO for cryopreservation of MSC-loaded alginate-chitosan microcapsules. The microcapsules were preserved with trehalose and/or DMSO and then stored in an isopropanol container at -80°C for stimulating a controlled cooling process. After a week of cryopreservation, microscopic observations showed that cryopreserved microcapsules maintained similar morphologies with freshly-prepared microcapsules. The only physical difference was a slight shrinking of the outer chitosan layer in the cryopreserved microcapsule, which was probably due to dehydration. Among cryopreserved microcapsules, DMSO 10% treatment gave the highest viability (88%), and then a combination of trehalose (50 mM) + DMSO (2%) treatment was 75%, and trehalose only treatment was 50%. DMSO (2%) treatment was excluded due to uncommon low cell recovery. Altogether, it was concluded that trehalose (50 mM) could be used to lower DMSO concentration, whereas low DMSO concentration (2%) or trehalose (50mM) alone was not sufficient to protect microencapsulated MSCs from negative effects of freezing and thawing. This study provides brief evidence of trehalose and DMSO potencies for cryopreserving stem cell-loaded microcapsule.