Cytokine-immobilized microparticle-coated plates for culturing hematopoietic progenitor cells

The purpose of this study was to provide a culture method for an effective expansion of human CD 34 positive hematopoietic progenitor cells (CD 34 (+) HCs) utilizing low molecular weight heparin/protamine microparticles (LH/P MPs) which can be stably coated onto plastic surfaces and cytokines. CD 34 (+) HCs optimally proliferated on LH/P MP-coated plates in the presence of stem cell factor (SCF; 5 ng/ml), thrombopoietin (Tpo; 10 ng/ml), and Flt-3 ligand (Flt-3; 10 ng/ml) in hematopoietic progenitor growth medium (HPGM). After 6 days, the total cells expanded 16.5-fold. Those cytokines were shown to be partially immobilized on the LH/P MP-coated plates, and the immobilized cytokines were gradually released into the medium with half releasing time of 3-4 days. Since flow cytometry analyses revealed that 90% of initial cells and 44.5% of expanded cells were CD 34 positive, CD 34 (+) HCs were estimated to have increased 8.0-fold after 6 days, and to have increased to over 31.9-fold after 12 days. In contrast, cultured CD 34 (+) HCs on non-coated tissue culture plates increased only 2.9-fold in the identical medium after 6 days, and only 5.2-fold after 12 days.