Danet G H, Lee H W, Luongo J L, Simon M C, Bonnet D A
Howard Hughes Medical Institute, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-6160, USA.
Exp Hematol. 2001 Dec;29(12):1465-73. doi: 10.1016/s0301-472x(01)00750-0.
The relationship between phenotype and function in ex vivo-cultured human hematopoietic stem cells (HSC) remains poorly understood. We investigated the effects of a short-term serum-free culture on the relationship between stem cell phenotype, cell division history, and function in human CD34(+) cells.
G-CSF-mobilized peripheral CD34(+) cells were cultured for 4 days with stem cell factor, flt-3 ligand, and thrombopoietin. The phenotype (CD34, CD38, HLA-DR, c-kit), cell division history, colony-forming cell (CFC), long-term culture-initiating cell (LTC-IC), and NOD/SCID repopulating activities were evaluated at Day 0 and 4.
We observed a loss of CD38, HLA-DR, and c-kit surface expression resulting in a drastic increase in CD34(+)CD38(-), CD34(+)HLA-DR(-), and CD34(+)c-kit(-/low) cells at Day 4. In contrast, the frequency of Thy-1(+) cells was maintained. We observed a 1.3-fold expansion of CFC, a 4.8-fold increase in LTC-IC, and an overall maintenance of the NOD/SCID repopulating cell activity. CD34(+)CD38(-) and CD34(+)HLA-DR(-) cells detected at Day 4 displayed the most active pattern of division (4 to 5 divisions) whereas 60% of CD34(+)Thy-1(+) cells divided 0 to 2 times during the same period. At Day 4, the NOD/SCID repopulating activity was associated with Thy-1(+) cells with no more than 2 divisions.
Our results show that the relationship between stem cell phenotype and function is dramatically altered in cultured CD34(+) cells. Thy-1 expression and cell division history appear to be superior to CD38, HLA-DR, and c-kit, or to homing molecules (CXCR4, VLA-4) as predictors of the repopulating activity of cultured peripheral CD34(+) cells.
体外培养的人类造血干细胞（HSC）的表型与功能之间的关系仍未得到充分理解。我们研究了短期无血清培养对人类CD34(+)细胞中干细胞表型、细胞分裂历史和功能之间关系的影响。
将粒细胞集落刺激因子（G-CSF）动员的外周血CD34(+)细胞与干细胞因子、fms样酪氨酸激酶3配体（flt-3 ligand）和血小板生成素一起培养4天。在第0天和第4天评估表型（CD34、CD38、人类白细胞抗原DR（HLA-DR）、原癌基因c-kit）、细胞分裂历史、集落形成细胞（CFC）、长期培养起始细胞（LTC-IC）以及非肥胖糖尿病/重症联合免疫缺陷（NOD/SCID）重建造血活性。
我们观察到CD38、HLA-DR和c-kit表面表达缺失，导致第4天CD34(+)CD38(-)、CD34(+)HLA-DR(-)和CD34(+)c-kit(-/低)细胞急剧增加。相比之下，Thy-1(+)细胞的频率保持不变。我们观察到CFC扩增了1.3倍，LTC-IC增加了4.8倍，并且NOD/SCID重建造血细胞活性总体维持。第4天检测到的CD34(+)CD38(-)和CD34(+)HLA-DR(-)细胞显示出最活跃的分裂模式（4至5次分裂），而同期60%的CD34(+)Thy-1(+)细胞分裂0至2次。在第4天，NOD/SCID重建造血活性与分裂不超过2次的Thy-1(+)细胞相关。
我们的结果表明，培养的CD34(+)细胞中干细胞表型与功能之间的关系发生了显著改变。作为培养的外周血CD34(+)细胞重建造血活性的预测指标，Thy-1表达和细胞分裂历史似乎优于CD38、HLA-DR、c-kit或归巢分子（CXCR4、极迟抗原4（VLA-4））。
