Background: Human umbilical cord blood (UCB) has been recognized as a source of hematopoietic stem cells for transplantation and the CD34+ cell dose is important factor predicting the hematologic recovery. However, the cell dose and CD34 (+) cells in one unit UCB are limited. This study compared the ex vivo culture conditions for expansion of total nucleated cells (TNC) and CD 34+ cells using isolated from UCB CD34 (+) and CD 133 (+) subset cells.

Material and method: The isolation of CD34 (+) and CD133 (+) cells from UCB was done by AutoMACS and the purity was more than 95%. We set three different cell culture conditions in stroma-free, serum-free medium. The culture conditions were 1) TPO, stem cell factor (SCF), Flt3-1 2) TPO/SCF/FLT3-1 and IL 3 10ng/dl, 3) TPO/SCF/FLT3-1 and IL 6 10ng/dl. After 7 days and 14days culture, we have been evaluated TNC and CD 34 (+) cell number for expansion potential.

Results: The TNC after 7 days and 14 days culture, in the CD 34(+) subset, the TPO/SCF/Flt3-1 condition was 15.6 fold and 8.0 fold and the TPO/SCF/Flt3-1 + IL-3 condition was 31.6 fold and 20.1 fold and the TPO/SCF/Flt3-1 + IL-6 condition was 25.5 fold and 12.0 fold, respectively. In the CD133 (+) subset, the TPO/SCF/Flt3-1 condition was 7.1 fold and 8.4 fold and in the TPO/SCF/Flt3-1 + IL-3 condition was 17.9 fold and 24.8 fold and in TPO/SCF/Flt3-1 + IL-6 condition was 15.9 fold and 19.8 fold, respectively. In terms of CD34 (+) cell population in the expanded cells, in the CD34 (+) subset, the TPO/SCF/Flt3-1 condition was 40%, the TPO/SCF/Flt3-1 + IL-3 condition was 64% and in the TPO/SCF/Flt3-1 + IL-6 condition was 40%, respectively. In the CD 133 (+) subset, the TPO/SCF/Flt3-1 condition was 83% and in TPO/SCF/Flt3-1 + IL-3 condition was 48% and in TPO/SCF/Flt3-1 + IL-6 condition was 62%, respectively.

Conclusions: To obtain more CD 34(+) cell population among ex vivo expanded cells, 7 days culture was more effective than 14 days culture and CD 133(+) subset was better than CD 34(+) subset. In terms of cell expansion ratio TPO/SCF/Flt3-1 + IL-3 was better than TPO/SCF/Flt3-1 and TPO/SCF/Flt3-1+ IL-6, which would be support the relevance of 7 days culture and CD133 (+) subset and IL-3 for expansion of hematopoietic stem cell ex vivo expansion.

Fig 1:
Fig 1:. The expansion ratio of both CD (34+)(a) and CD133 (+)(b) subset after 7 days and 14 days cultures in the 3 different cell culture conditions
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The expansion ratio of both CD (34+)(a) and CD133 (+)(b) subset after 7 days and 14 days cultures in the 3 different cell culture conditions

Fig 1:
Fig 1:. The expansion ratio of both CD (34+)(a) and CD133 (+)(b) subset after 7 days and 14 days cultures in the 3 different cell culture conditions
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The expansion ratio of both CD (34+)(a) and CD133 (+)(b) subset after 7 days and 14 days cultures in the 3 different cell culture conditions

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Fig 2:
Fig 2:. The CD34(+) cell percentages in the different culture conditions both CD34(+) and CD133(+) subsets.
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The CD34(+) cell percentages in the different culture conditions both CD34(+) and CD133(+) subsets.

Fig 2:
Fig 2:. The CD34(+) cell percentages in the different culture conditions both CD34(+) and CD133(+) subsets.
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The CD34(+) cell percentages in the different culture conditions both CD34(+) and CD133(+) subsets.

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Topics:
interleukin-3, ms-like tyrosine kinase 3, cd34 antigens, interleukin-6, cell culture techniques, stem cell factor, transplantation, cell count, hematopoietic stem cells, erythroblasts

Author notes

Corresponding author

2005, The American Society of Hematology
2005
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