Early apoptosis largely accounts for functional impairment of CD34+ cells in frozen-thawed stem cell grafts

被引:51
作者
De Boer, F
Dräger, AM
Pinedo, HM
Kessler, FL
Monnee-Van Muijen, M
Weijers, G
Westra, G
Van der Wall, E
Netelenbos, T
Oberink, JW
Huijgens, PC
Schuurhuis, GJ
机构
[1] Free Univ Amsterdam, Med Ctr, Dept Hematol, NL-1081 HV Amsterdam, Netherlands
[2] Free Univ Amsterdam, Med Ctr, Dept Med Oncol, NL-1081 HV Amsterdam, Netherlands
来源
JOURNAL OF HEMATOTHERAPY & STEM CELL RESEARCH | 2002年 / 11卷 / 06期
关键词
D O I
10.1089/152581602321080619
中图分类号
R5 [内科学];
学科分类号
1002 ; 100201 ;
摘要
Quality assessment of stem cell grafts is usually performed by flow cytometric CD34(+) enumeration or assessment of clonogenic output of fresh material. Previously, we identified the occurrence of early apoptosis, not detectable with the permeability marker 7-amino actinomycin D (7-AAD), in purified frozen-thawed CD34(+) cells, using the vital stain Syto16. Syto(high)/7-AAD(-) cells were defined as viable, Syto16(low)/7-AAD(-) cells as early apoptotic and Syto16(low)/7-AAD(+) as dead. This was confirmed in a subsequent study using frozen-thawed transplants of lymphoma patients. In the present study on grafts from multiple myeloma and lymphoma patients, we investigated the functional consequences of the early apoptotic process. The mean Syto16-defined viability was 41 and 42%, respectively, for both graft groups, compared to 78% and 72%, respectively, using 7-AAD only. The established early apoptosis marker annexin V missed roughly 50% of the early apoptosis detected with Syto16. In contrast, viability of CD34(+) cells in nonmanipulated whole blood transplants from a matched group of lymphoma. patients, after 72 h of storage at 4degreesC, was more than 90%, even with the Syto16 assay. CFU recovery (median 26-33%) after cryopreservation matched CD34(+) recovery after Syto16, but not 7-AAD correction. In contrast, colony-forming unit (CFU) recovery in the whole blood transplant was close to 100%. Furthermore, early apoptotic CD34(+) cells had lost migratory ability toward stromal cell derived factor-1alpha (SDF-1alpha). The establishment of a Syto16(high)/7-AAD(-) proportion of CD34(+) cells offers a new approach for. a more correct determination of the number of viable nonapoptotic CD34(+) cells in stem cell grafts. Further development of this assay should allow its incorporation into the routine CD34(+) assessment of post-thawed samples in clinical flow cytometry laboratories.
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收藏
页码:951 / 963
页数:13
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