Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Holzinger, L.
Cryopreservation and Thawing of Peripheral Blood Stem Cells
[ Diplomarbeit ] Medical University of Graz; 2010. pp. 78
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- Autor*innen der Med Uni Graz:
- Betreuer*innen:
- Ritter-Sovinz Petra
- Schwinger Wolfgang
- Altmetrics:
- Abstract:
- Background: Cryopreservation is the storage method of choice for autologous peripheral blood stem cells (PBSC) and allogeneic umbilical cord blood stem cells intended for haematopoietic stem cell transplantation. The number of haematopoietic stem cells (CD34+) correlates with haematological engraftment. However, the viability of haematopoietic stem cells may be influenced by different variables during cryopreservation and thawing. The aim of the study was to investigate influence of dimethyl sulfoxide (DMSO), temperature and incubation time on different haematological cell types. To our knowledge, no commonly accepted algorithm for processing and short-time storage after thawing of cryopreserved PBSC exists. Study Design and Methods: Frozen cryovials were thawed in a 37 °C water bath and subsequently DMSO was washed out either prior to (protocol I) or after an incubation period (protocol II). Cell samples were separately incubated at three different temperature stages (6 °C, 20 °C, 37 °C) and analysed after four different sampling delay periods (immediately at 0 min and after 30, 60 and 120 min) for cell counts, cell subtypes (FACS), viability (Annexin V/PI) and repopulating abilities (2nd CFU, LTC-IC). Statistical analysis was performed using a two-way analysis of variance method. Results: Significant declines in cell counts were seen for some of T lymphocyte subpopulations (CD3+, CD3+/4+, CD3+/8+) at 20 °C and 37 °C mainly if cells were incubated in the presence of DMSO; this decline was more pronounced with increasing incubation time. Although in this study protocol, CD34+ numbers did not decrease, frequency of LTC-IC excessively decreased after 60 min (II) and 120 min (I) of incubation at 37 °C, respectively. Conclusions: These results suggest that post-thaw processing and infusion of cryopreserved peripheral blood stem cell products should not take longer than 60 minutes in order to avoid cell loss and compromise of viability of haematopoietic stem cells. Immediate wash-out of DMSO additionally reduces detrimental effects on cell counts and repopulating potential. Thawed products should be infused at lowest temperatures possible, as warming-up to 37 °C body temperature negatively affects quality of graft.