The cryopreservation protocol optimal for progenitor recovery is not optimal for preservation of marrow repopulating ability

The efficiency of five different cryopreservation protocols (our original c ontrolled-rate and noncontrolled-rate protocols) was evaluated on the basis of the recovery after thawing of very primitive pluripotent hemopoietic st em cells (MRA(CFU-GM)), pluripotent progenitors (CFU-Sd12) and committed gr anulocyte-monocyte progenitors (CFU-GM) in mouse bone marrow. Although the nucleated cell recovery and viability determined immediately after the thaw ing and washing of the cells were found to be similar, whether controlled-r ate or noncontrolled-rate cryopreservation protocols were used, the recover y of MRA(CFU-GM), CFU-Sd12 and CFU-GM varied depending on the type of proto col and the cryoprotector (DMSO) concentrations used. It was shown that the controlled-rate protocol was more efficient, enabling better MRA(CFU-GM), CFU-Sd12 and CFU-GM recovery from frozen samples. The most efficient was th e controlled-rate protocol of cryopreservation designed to compensate for t he release of fusion heat, which enabled a better survival of CFU-Sd12 and CFU-GM when combined with a lower (5%) DMSO concentration. On the contrary, a satisfactory survival rate of very primitive stem cells (MRA(CFU-GM)) wa s achieved only when 10% DMSO was included with a five-step protocol of cry opreservation, These results point to adequately used controlled-rate freez ing as essential for a highly efficient cryopreservation of some of the cat egories of hematopoietic stem and progenitor cells, At the same time, it wa s obvious that a higher DMSO concentration was necessary for the cryopreser vation of very primitive stem cells, but not, however, for more mature prog enitor cells (CFU-S, CFU-GM). These results imply the existence of a mechan ism that decreases the intracellular concentration of DMSO in primitive MRA cells, which is not the case for less primitive progenitors.