Primitive hematopoietic progenitors within mobilized blood are spared by uncontrolled rate freezing

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Abstract
Uncontrolled-rate freezing techniques represent an attractive alternative to controlled-rate cryopreservation procedures which are time-consuming and require high-level technical expertise. In this study, we report our experience using uncontrolled-rate cryopreservation and mechanical freezer storage at -140 degrees C. Twenty-eight PBPC samples (10 cryovials, 18 freezing bags) from 23 patients were cryopreserved in a cryoprotectant solution composed of phosphate-buffered saline (80%, v/v) supplemented with human serum albumin (10%, v/v) and dimethylsulfoxide (10%, v/v). The cryopreservation procedure required on average 1.5 h. The mean (+/- s.e.m.) storage time of cryovials and bags was 344+/-40 and 299+57 days, respectively. Although cell thawing was associated with a statistically significant reduction of the absolute number of nucleated cells (vials: 0.3x10(9) vs. 0.2x10(9), P< or =0.02; bags: 14x10(9) vs. 11x10(9), P< or =0.0003), the growth of committed progenitors was substantially unaffected by the freezing-thawing procedure, with mean recoveries of CFU-Mix, BFU-E, and CFU-GM ranging from 60+/- 29% to 134+/-15%. Mean recoveries of LTC-IC from cryovials and bags were 262+/-101% and 155+/-27% (P< or =0.2), respectively. In 14 out of 23 patients who underwent high-dose chemotherapy and PBPC reinfusion, the pre-and post-freezing absolute numbers of hematopoietic progenitors cryopreserved in bags were compared. A significant reduction was detected for CFU-Mix (11 vs. 7.4x10(5)), but no significant loss of BFU-E (180 vs. 150x10(5)), CFU-GM (400 vs. 290x10(5)) and LTC-IC (15 vs. 16x10(5)) could be demonstrated. When these patients were reinfused with uncontrolled-rate cryopreserved PBPC, the mean number of days to reach 1x10(9)/l white blood cells and 50x10(9)/l platelets were 9 and 13, respectively. In conclusion, the procedure described here is characterized by short execution time, allows a substantial recovery of primitive and committed progenitors and is associated with prompt hematopoietic recovery following myeloablative therapy even after long-term storage.

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