Evaluation of trehalose and sucrose as cryoprotectants for hematopoietic stem cells of umbilical cord blood

Abstract

Bone marrow transplantation (BMT) is a therapeutic procedure that involves transplantation of hematopoietic stem cells (HSC). To date, there are three sources of HSC for clinical use: bone marrow; mobilized peripheral blood; and umbilical cord blood (UCB). Depending on the stem cell source or type of transplantation, these cells are cryopreserved. The most widely used cryoprotectant is dimethylsulfoxide (Me 2SO) 10% (v/v), but infusion of Me 2SO-cryopreserved cells is frequently associated with serious side effects in patients. In this study, we assessed the use of trehalose and sucrose for cryopreservation of UCB cells in combination with reduced amounts of Me 2SO. The post-thawed cells were counted and tested for viability with Trypan blue, the proportion of HSC was determined by flow cytometry, and the proportion of hematopoeitic progenitor cells was measured by a colony-forming unit (CFU) assay. A solution of 30 mmol/L trehalose with 2.5% Me 2SO (v/v) or 60 mmol/L sucrose with 5% Me 2SO (v/v) produced results similar to those for 10% (v/v) Me 2SO in terms of the clonogenic potential of progenitor cells, cell viability, and numbers of CD45 +/34 + cells in post-thawed cord blood cryopreserved for a minimum of 2 weeks. Thus, cord blood, as other HSC, can be cryopreserved with 1/4 the standard Me 2SO concentration with the addition of disaccharides. The use of Me 2SO at low concentrations in the cryopreservation solution may improve the safety of hematopoietic cell transplantation by reducing the side effects on the patient.