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 controlled-rate and noncontrolled-rate protocols) was evaluated on the basis of the recovery after thawing of very primitive pluripotent hemopoietic stem cells (MRA(CFU-GM), pluripotent progenitors (CFU-Sd12) and committed granulocyte-monocyte progenitors (CFU-GM) in mouse bone marrow. Although the nucleated cell recovery and viability determined immediately after the thawing and washing of the cells were found to be similar, whether controlled-rate or noncontrolled-rate cryopreservation protocols were used, the recovery of MRA(CFU-GM), CFU-Sd12 and CFU-GM varied depending on the type of protocol 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 the controlled-rate protocol of cryopreservation designed to compensate for the 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)) was achieved only when 10% DMSO was included with a five-step protocol of cryopreservation. These results point to adequately used controlled-rate freezing as essential for a highly efficient cryopreservation of some of the categories of hematopoietic stem and progenitor cells. At the same time, it was obvious that a higher DMSO concentration was necessary for the cryopreservation of very primitive stem cells, but not, however, for more mature progenitor cells (CFU-S, CFU-GM). These results imply the existence of a mechanism that decreases the intracellular concentration of DMSO in primitive MRA cells, which is not the case for less primitive progenitors.
Balint
B
B
Institute of Transfusiology of MMA, Belgrade, Yugoslavia.
Ivanović
Z
Z
Petakov
M
M
Taseski
J
J
Jovcić
G
G
Stojanović
N
N
Milenković
P
P
D000818:Animals; D002452:Cell Count; D002470:Cell Survival; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D005260:Female; D006098:Granulocytes / Q000166:cytology; D008264:Macrophages / Q000166:cytology; D008297:Male; D051379:Mice; D008808:Mice, Inbred CBA; D013234:Stem Cells / Q000166:cytology
D003451:Cryoprotective Agents; D004121:Dimethyl Sulfoxide