[Preliminary study on extensive amplification of human dendritic cells differentiated from cord blood CD34+ progenitor cells by two-step culture].
OBJECTIVE
To Explore a two-step culture system to generate a large number of dendritic cells (DC) differentiated from cord blood (CB) CD(34)(+) cells.


METHODS
Enriched CB CD(34)(+) cells with immunoadsorption were primarily cultured in the presence of stem cell factor (SCF), Flt-3 ligand (FL), thrombopoietin (Tpo) and interleukin-3 (IL-3) for 7 (group I), 10 (group II) or 14 days (group III) respectively, and then further cultured with GM-CSF, IL-4 and TNF-alpha for 5 - 8 days to induce DC. The expansion and cell function were evaluated by flow cytometry (FCM) and mix-lymphocyte reaction (MLR), and detection of IL-12 in the supernatant by using ELISA.


RESULTS
The total nucleated cells with 53.39 +/- 20.59-, 307.17 +/- 119.59- and 1117.25 +/- 335.49-folds expansion could be respectively obtained after 7 - 14 days of expansion culture. After DC induction, CD(1a)(+) cells were 21.40 +/- 16.70-, 143.2 +/- 60.35- and 150.8 +/- 42.16-fold increase as compared to the initial nucleated cells. Comparing with that in group I, the CD(1a)(+) cells were much more in groups II and III; but there was no difference between the latter two groups (P > 0.05). The cultured cells in the three groups showed almost the same allo-stimulatory capability and IL-12 excretion when the second culture duration maintained 8 days, while the capability and excretion were greatly decreased when the duration shortened to 5 days (P < 0.05).


CONCLUSIONS
A plenty of functionally mature DC could be obtained from the CD(34)(+) cells in the two-step culture system of 7 - 10 days HSC expansion followed by 8 days DC induction.
State Key Laboratory of Experimental Hematology, Institute of Hematology and Blood Diseases Hospital, CAMS & PUMC, Tianjin 300020, China.
Ya-fei
YF
Wang
Qian
Q
Li
Heng-xing
HX
Meng
Zhen
Z
Yu
Jin-hua
JH
Liu
Wen
W
Cui
Yu
Y
Zhou
Yu-jie
YJ
Mai
Sheng-guo
SG
You
Lu-gui
LG
Qiu
D018952:Antigens, CD34 / Q000032:analysis*; D002454:Cell Differentiation; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000166:cytology* / Q000502:physiology; D005312:Fetal Blood / Q000166:cytology*; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D018664:Interleukin-12 / Q000096:biosynthesis; D008213:Lymphocyte Activation
D018952:Antigens, CD34; D018664:Interleukin-12