The Effect of Stromal Cells and Hematopoietic Growth Factors on Proliferation and Differentiation of Megakaryocytic Progenitors from Umbilical Cord Blood / 대한소아혈액종양학회지

ABSTRACT

PURPOSE: CD34+ cells from umbilical cord blood (UCB) were cultured with stromal cells and growth factors, and cell proliferation and megakaryocytic differentiation were observed. The purposes of this study are to find out the optimal culture condition for the megakaryocytic differentiation of CD34+ cells from UCB. METHODS: CD34+ cells of mobilized peripheral blood (PB) and UCB were cultured in IMDM at a concentration of 1x105 cells/mL for 11 days. Thrombopoietin (TPO) 5 ng/mL and 50 ng/mL, flt-3 ligand (FL) 50 ng/mL and stem cell factor (SCF) 50 ng/mL were used as growth factors. Stromal cells cultured from bone marrow (BM) mononuclear cells appeared as a single layer of fusiform adherent cells with expression of SH-2 and ASMA. RESULTS: When PB CD34+ cells were cultured with growth factors only, the cell number increased in TPO 5 ng/mL and 50 ng/mL, TPO and FL, TPO, FL and SCF group increased upto mean 2.0 (in TPO 5 ng/mL), 2.9 (in TPO 50 ng/mL), 2.4 (TPO and FL), 2.8 (TPO, FL and SCF) fold, and the expression of CD61a were mean 11.5%, 12.5%, 13.2% and 17.0%, respectively. When the stromal cells were added to the growth factors, the cell number increased upto mean 4.6, 4.9, 9.2 and 68.5 fold and CD61a were expressed in mean 43.1%, 48.4%, 35.6%, and 6.2% of cultured cells. For UCB CD34+ cells, the cell number were increased upto 7.5, 7.6, 8.2 and 23.6 fold with growth factors and stromal cells. Expression of CD61a were mean 26.0%, 34.3%, 31.5% and 23.5%, respectively. CONCLUSION: Stromal cell enhanced the cellular proliferation and megakaryocytic differentiation of UCB CD34+ cells. The combination of TPO, FL and SCF increased total cell number upto the highest value but the proportion of the CD61a+ cells were relatively low. The combination of TPO and FL induced both cellular proliferation and megakaryocytic differentiation in a large amount.