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Introduction
There are several hematopoietic-stem cells (HSCs) thawing methods for bone marrow reconstitution. They intend to avoid cell death and patient's side effects due to the dimethyl sulfoxide (DMSO). We propose another thawing method that diminishes cell death and therefore a more rapid hematological recovery.
Material and Methods
The standard thawing-removing DMSO method for cord blood units was described by Rubinstein in 1995 (method 1). Ours (method 2) pretend to increase more than 10 fold the dilution of cryopreserved HSCs in the standard washing solution (5% albumin + dextran 40). Methods 1 and 2 were compared to determine viability by means of total-nucleated-cell (TNC) count by trypan blue and flow cytometry at the time of collection, cryopreservation, thawing, and after removing the DMSO, as well as the patient's day of engraftment.
Results
Results are shown in Table 1 and 2.
Conclusions
A greater dilution of cryopreserved HSCs in the washing solution, as a new thawing method, may decrease cell death; therefore a greater number of HSCs will be infused to the patient. Studies with a larger number of thawing procedures are needed to make this assertion.
Table 1. Patient's demographic data.
Global
| Method 1
| Method 2
| P
| |
Total (n)
| 26
| 13
| 13
| |
Transplant (n) Autologous Allogeneic
| 12 14
| 5 8
| 7 6
| .43
|
Receptor age (years) Median (range)
| 26 (3-56)
| 27 (5-55)
| 25 (3-56)
| .75
|
Figure 1. Viability loss
Figure 2. Neutrophil engrafment
Figure 3. Platelel engrafment
Table 2. Overall results in the study.
Global
| Rubinstein's Method
| New Method
| P
| |
Viability before thawing. Mean (SD)
| 97.28% (6.55)
| 99.01% (1.08)
| 95.56% (9.04)
| .30
|
Viability after thawing. Mean (SD)
| 78.92% (11.69)
| 75.46% (12.5)
| 82.39% (10.12)
| .15
|
Viability loss. Mean (SD)
| 18.36% (12.11)
| 23.55% (12.74)
| 13.18% (9.21)
| .02
|
Engrafment days >20,000 Platelets. Median (range)
| 11 (6-23)
| 11 (6-23)
| 11 (7-19)
| .60
|
Engrafment days >500 Neutrophils. Median (range)
| 13.5 (6-28)
| 13 (6-28)
| 14 (9-17)
| .30
|