Effect of cell concentration on bone marrow and peripheral blood stem cell cryopreservation

Abstract

The effects of cell concentration during cryopreservation on bone marrow (BM) or peripheral blood (PB)-derived hematopoietic progenitor cells have not been described. The much greater numbers of cells harvested for autologous PB stem cell (PBSC) transplantation requires that the cells be frozen at higher cell concentrations, or in much greater volumes, compared with BM. We cryopreserved 108 PBSC collections from 30 patients at an average (+/- SD) cell concentration of 3.7 +/- 1.9 x 10(8) nucleated cells per mL in 127 +/- 45 mL. The proportion of mononuclear cells was 52.9% +/- 27.2%. The products also contained 2.9 +/- 2.1 x 10(9) platelets/mL and an average red cell proportion of 12.9% +/- 7.2%. The nucleated cell recovery after thawing was 75.4% +/- 13.0%. The nucleated cell concentration during freezing was not predictive for the postthaw recoveries of nucleated cells (P = .38), granulocyte-macrophage colony-forming unit (P = .06) or CD34+ cells (P = .54), or for the viability of mononuclear cells (P = .81). The platelet and red cell concentrations similarly were not predictive for these endpoints. Samples (3 BM, 7 PBSC) from 10 patients were simultaneously cryopreserved at two-fold, and from 5 additional patients (PBSC) at 6- to 24-fold differing cell concentrations. A lower recovery of erythroid burst forming unit was found for samples frozen at higher cell concentrations (P = .04), but no significant differences were found in the other endpoints listed above. The average cell concentration during freezing for each patient's PBSC collections (n = 34 patients) did not predict time to achieve a PB count of > 500 granulocytes/microL (P = .51) or platelet transfusion independence (P = .39). Patients achieved these endpoints of engraftment at medians of 12 and 13 days, respectively. The infusion of these products was generally well tolerated. Similarly, the cell concentration at which BM cells were frozen did not predict for the duration of granulocyte (P = .63) or platelet (P = .36) aplasias for 54 patients undergoing autologous BM transplantation. These data suggest that PBSC or BM cells collected for transplantation may be cryopreserved at very high cell concentrations without loss of engraftment potential or undue infusion-related toxicity.

Cryopreservation impact on blood progenitor cells: influence of diagnoses, mobilization treatments, and cell concentration

“…In contrast to other reports describing no effect of the cell concentration in cryopreservation mixture, 24,28 in our study cell concentration impacted postthawing recovery of CD34+ cells, granulocytes, lymphocytes, and monocytes, but not cell viability. A better understanding of how cryopreservation affects progenitor cells 30 in different ways in line with diagnoses, mobilization, and cell concentration might permit a more rational use of freezing procedures and should help us to explain these discordances.…”

“…Some authors describe decreased counts of CD34+ cells after thawing while others find increased amounts, 5,6,18 differences that, in our opinion, could have been induced by the use of short and heterogeneous series of patient and variable progenitor cell mobilization protocol. Additionally, diversity in the way to express results, percentage, concentration/mL, recovery, or millions/kg could have also contributed 5,8,18,24,25 . Accordingly to previous results described for Sartor and colleagues, 26 in our series the amount of CD34+ cells ×10 6 /L slightly but significantly decreased after thawing when the total group of samples was analyzed, although no significant differences could be demonstrated when diagnoses were studied separately.…”

Leukapheresis components may be cryopreserved at high cell concentrations without additional loss of HPC function

“…The present study showed that the cryopreservation of PBPC components at cell concentrations 3.3 (1.8‐6.2)‐fold the standard had no adverse effect on the in vitro functionality of HPCs after thawing or on hematopoietic recovery after transplantation. Our findings are in agreement with the results published by Rowley et al, 12 and they contrast with the decreased recovery of CFU–GM found by Keung et al 11 when PBPCs were cryopreserved at high cell concentrations. In both studies, hematopoietic engraftment after transplantation was not adversely affected.…”

“…To date, there are no conclusive results as to the implementation of these strategies. To our knowledge, only two studies focused on an analysis of the effect of cell concentration on the cryopreservation of peripheral blood progenitor cells (PBPCs) have been published, with discordant results in terms of functional recovery of PBPCs 11,12 …”

Storage characteristics of cord blood progenitor cells: report of a multicenter study by the cellular therapies team of the Biomedical Excellence for Safer Transfusion (BEST) Collaborative

“…It has been found that when HPCs are stored overnight at RT with a cell concentration of greater than 200 × 10 9 /L, there is a significant reduction of the cell viability and the recovery of CD34+ cells and colony‐forming unit–granulocyte‐macrophage (CFU‐GM) 8 . However, cells stored at concentrations between 400 × 10 9 to 500 × 10 9 /L but cryopreserved within 4 hours of collection showed satisfactory postthaw characteristics 9 …”