目的 探索利用先扩增后诱导的“两步法”从脐血（CB）CD34+细胞高效大量地获得树突细胞（DC）。方法 疫磁珠法从CB分选获得CD34+细胞，以干细胞因子（SCF）、IL23、Flt23配体（FL）、Tpo组合刺激，扩增7d、10 d和14d（依次为Ⅰ、Ⅱ和Ⅲ组） 后以GM-CSF+IL-4+TNF-α诱导8d或5d获得DC，通过相差显微镜、电镜观察形态，流式细胞仪检测表型，混合淋巴细胞培养、ELISA法检测培养液上清IL-12含量评价其功能。结果 CBCD34+细胞经SCF+IL-3+FL+Tpo刺激扩增7d、10d 和14d后细胞总数分别扩增了（53.39±20.59）倍、（307.17±119.59）倍和（1117.25±335.49）倍。经GM-CSF+IL-4+TNF-α诱导8d后所得CD1a+细胞是扩增前细胞数的（21.40±16.70）倍、（143.20±60.35）倍和（150.80±42.16）倍，Ⅱ、Ⅲ组明显多于Ⅰ组（P<0.05），但Ⅱ、Ⅲ组间无显著性差异（P>0.05）。所得DC的形态、表型及刺激异基因T细胞增殖能力、IL-12分泌量，三组无显著性差异（P>0.05）。当诱导时间缩短至5d时，各组DC功能均显著下降（P<0.05）。结论 CBCD34+细胞扩增7～10再诱导8d可以高效大量获得具有正常功能的DC，而扩增时间超过10d并不能显著增加DC产量，诱导时间少于 8d将降低所得DC的功能。

To investigate the clinically applicable conditions that support substantial expansion of both primitive and more mature hematopoietic cells of umbilical cord blood (UCB) for transplantation in adults, enriched CD34+ cells from 8 fresh UCB samples and 4 expanded UCB products were cultured in defined serum-free medium (QBSF-60) in the presence of a cytokine combination of SCF, Flt-3-ligand (FL), thrombopoietin (TPO), IL-3 for up to 2 weeks. Fresh medium with cytokines was supplemented or exchanged at day 4, day 7, and day 10. The proliferative response was assessed at day 7, day 10, and day 14 by evaluating the following parameters: nucleated cell (NC), clonogenic progenitors (colony-forming unit-granulocyte-macrophage [CFU-GM], burst-forming unit-erythrocyte [BFU-E], CFU-GEMM, and high-proliferative potential colony-forming cell [HPP-CFC]), immunophenotypes (CD34+ cells and CD34+ subpopulations), and LTCIC. Simultaneously numerical expansion of various stem/progenitor cells, including primitive CD34+ CD38- HLA-DR- subpopulation and LTCIC, CD34+ cells, and clonogenic progenitors to mature nucleated cells, were continuously observed during the culture. An average 103.32±71.37×106 CD34+ cells (range 10.12×106-317.9×106) could be obtained from initial 1.72±1.13×106 UCB CD34+ cells after 10-14 days cultured under the described conditions. Sufficient CD341 cells (＞50.0×106) for transplantation in adults would be available in all but one UCB collections after 10-14 days expansion. The expanded CD34+ cells sustained most of the in vitro characteristics of initial unmanipulated CD34+ cells, including clonogenic efficiency (of both primitive and committed progenitors), the proportion of CD34+ CD38-HLA-DR- subpopulation, and the expansion potential. Initial addition of IL-3 to the cocktail of SCF+ FL+ TPO had positive effects on the expansion of both primitive and, especially, the more mature hematopoietic cells. It accelerated the expansion speed and shortened the optimal culture time from 14 days to 10 days. These results indicated that our proposed shortterm culture system, consisting of QBSF-60 serum-free medium with a simple early acting cytokine combination of SCF+ FL+TPO, could substantially support simultaneous expansion of various stem/progenitor cell populations involved in the different phases of engraftment. It would be a clinically applicable protocol for ex vivo expansion of CD34+ UCB cells.