2007 Fiscal Year Final Research Report Summary
Development of the system to produce large scale generation of human platelets from cord blood CD34^+ cells in vitro
| Project/Area Number |
18591076
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Hematology
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| Research Institution | Sapporo Medical University |
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Principal Investigator |
MATSUNAGA Takuya Sapporo Medical University, School of Medicine, Assistant professor (70260768)
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| Co-Investigator(Kenkyū-buntansha) |
IYAMA Satoshi Sapporo Medical University, School of Medicine, Instructor (50398319)
SATO Tsutomu Sapporo Medical University, School of Medicine, Instructor (40404602)
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| Project Period (FY) |
2006 – 2007
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| Keywords | Cord blood / CD34^+ cells / Artificial platelets |
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| Research Abstract |
We generated platelets (PLT) from cord blood (CB) CD34^+ cells employing a three-phase culture system. We first cultured 500 CB CD34^+ cells on telomerase gene-transduced human stromal cells (hTERT stroma) in serum-free medium supplemented with stem cell factor (SCF), Flt-3/Flk-2 ligand (FL) and thrombopoietin (TPO) for 14 days. We then transferred the cells to hTERT stroma and cultured for another 14 days with fresh medium containing interleukin-11 (IL-11) in addition to the original cytokine cocktail. Subsequently, we cultured the cells in a liquid culture medium containing SCF, FL, TPO and IL-11 for another 5 days to recover PLT fractions from the supernatant, which were then gel filtered to purify the PLT. The calculated yield of PLT from 1.0 unit CB (5x10^6 CD34+ cells) was 1.26 x10^<11> - 1.68 x10^<11> PLT. These numbers of PLT are equivalent to 2.5-3.4 units of random donor-derived PLT or 2/5-6/10 of a single apheresis PLT. The CB-derived PLT exhibited features quite similar to those from peripheral blood in morphology, as revealed by electron micrographs, and in function, as revealed by fibrinogen/ADP aggregation, with the appearance of P-selectin and activated glycoprotein IIb-IIIa antigens. Thus this culture system may be applicable for large scale generation of PLT for future clinical usage.
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