Ex Vivo Expansion of Primitive Hematopoietic Cells by Using Hematopoietic Growth Factors

Sung, Ki Woong; Lee, Jun Ah; Yoo, Keon Hee; Kim, Dong Hyun; Jung, Hye Lim; Koo, Hong Hoe; Shin, Hee Young; Ahn, Hyo Seop

Abstract

PURPOSE
If primitive hematopoietic cells expanded ex vivo can be used in stem cell transplantation, duration for hematologic recovery will be shortened. In this study, CD34 cells were cultured with various hematopoietic growth factors which are known to stimulate proliferation of primitive hematopoietic cells.
METHODS
CD34 cells isolated from cord blood were cultured and expanded ex vivo with IL-3, stem cell factor (SCF), flt-3 ligand (FL), thrombopoietin (TPO), granulocyte-colony stimulating factor (G-CSF). To find optimal combination of growth factors, CD34 cells were cultured for 9 days with various combinations of growth factors. To find optimal duration of culture, CD34 cells were cultured for 5, 7, 9, 12 days. To evaluate expansion of primitive hematopoietic cells, the number of CD34 cells, colony forming cells and long-term culture-initiating cells (LTC-IC) were counted by flow cytometry, colony forming cell assay and limiting dilution assay respectively.
RESULTS
Primitive hematopoietic cells were successfully expanded from CD34 cells of cord blood. Maximal expansion of LTC-IC and CFU- GEMM were 2.58 and 2.37 fold respectively after 9 days of culture, and were obtained with the combination of IL-3 SCF FL TPO. When CD34 cells were cultured for 5, 7, 9, 12 days with the combination of IL-3 SCF FL TPO, expansion of LTC-IC was maximal (2.95 fold) after 9 days culture. After reaching maximal expansion of LTC-IC, the number of nucleated cells increased, but that of primitive hematopoietic cells decreased.
CONCLUSION
Primitive hematopoietic cells can be successfully expanded ex vivo by using hematopoietic growth factors. Duration for hematologic recovery after stem cell transplantation will be shortened by using primitive hematopoietic cells expanded ex vivo.