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.
Kishimoto
Satoko
S
Research Institute, National Defense Medical College, Saitama, Japan.
Nakamura
Shingo
S
Nakamura
Shin-ichiro
S
Hattori
Hidemi
H
Oonuma
Fumie
F
Kanatani
Yasuhiro
Y
Tanaka
Yoshihiro
Y
Harada
Yasuji
Y
Tagawa
Masahiro
M
Maehara
Tadaaki
T
Ishihara
Masayuki
M
D018952:Antigens, CD34 / Q000032:analysis* / Q000097:blood; D001667:Binding, Competitive; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods; D049109:Cell Proliferation / Q000187:drug effects*; D003470:Culture Media / Q000737:chemistry / Q000494:pharmacology; D016207:Cytokines / Q000737:chemistry / Q000494:pharmacology*; D017985:Dalteparin / Q000737:chemistry; D003692:Delayed-Action Preparations / Q000737:chemistry / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D006025:Glycosaminoglycans / Q000737:chemistry; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D006493:Heparin / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry / Q000494:pharmacology; D008863:Microspheres; D008970:Molecular Weight; D010316:Particle Size; D011479:Protamines / Q000737:chemistry; D019089:Stem Cell Factor / Q000737:chemistry / Q000494:pharmacology; D013926:Thrombopoietin / Q000737:chemistry / Q000494:pharmacology
D018952:Antigens, CD34; D003470:Culture Media; D016207:Cytokines; D003692:Delayed-Action Preparations; D006025:Glycosaminoglycans; D008565:Membrane Proteins; D011479:Protamines; D019089:Stem Cell Factor; C084881:flt3 ligand protein; D006493:Heparin; D013926:Thrombopoietin; D017985:Dalteparin