Efficient generation of CD34+ progenitor-derived dendritic cells from G-CSF-mobilized peripheral mononuclear cells does not require hematopoietic stem cell enrichment.
As a result of their potent antigen-presentation function, dendritic cells (DC) are important tools for cell therapy programs. In vitro-generated DC from enriched CD34+ hematopoietic stem cells (HSC; enriched CD34 DC) have already proven their efficiency in Phase I/II clinical trials. Here, we investigated whether enrichment of CD34+ HSC before the onset of culture was absolutely required for their differentiation into DC. With this aim, we developed a new two-step culture method. PBMC harvested from G-CSF-mobilized, healthy patients were expanded for 7 days during the first step, with early acting cytokines, such as stem cell factor, fetal liver tyrosine kinase 3 ligand (Flt-3L), and thrombopoietin. During the second step, expanded cells were then induced to differentiate into mature DC in the presence of GM-CSF, Flt-3L, and TNF-alpha for 8 days, followed by LPS exposure for 2 additional days. Our results showed that the rate of CD34+/CD38+/lineageneg cells increased 19.5+/-10-fold (mean+/-sd) during the first step, and the expression of CD14, CD1a, CD86, CD80, and CD83 molecules was up-regulated markedly following the second step. When compared with DC generated from enriched CD34+ cells, which were expanded for 7 days before differentiation, DC derived from nonenriched peripheral blood stem cells showed a similar phenotye but higher yields of production. Accordingly, the allogeneic stimulatory capacity of the two-step-cultured DC was as at least as efficient as that of enriched CD34 DC. In conclusion, we report herein a new two-step culture method that leads to high yields of mature DC without any need of CD34+ HSC enrichment.
Paczesny
Sophie
S
Hematology Department, Children's Hospital, CHU Nancy, France, and Department of Pediatrics, University of Michigan Cancer Center, 1500 East Medical Center Drive, Ann Arbor, MI 48109-0942, USA. sophiep@umich.edu
Li
Yin-Ping
YP
Li
Na
N
Latger-Cannard
Véronique
V
Marchal
Luc
L
Ou-Yang
Jing-Ping
JP
Bordigoni
Pierre
P
Stoltz
Jean-François
JF
Eljaafari
Assia
A
D018952:Antigens, CD34 / Q000378:metabolism*; D000943:Antigens, Differentiation / Q000032:analysis; D002454:Cell Differentiation; D003713:Dendritic Cells / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology*; D006412:Hematopoietic Stem Cells / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000187:drug effects / Q000502:physiology; D011994:Recombinant Proteins; D046509:Tissue Culture Techniques / Q000379:methods*
D018952:Antigens, CD34; D000943:Antigens, Differentiation; D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor