Rapid induction of CD40 on a subset of granulocyte colony-stimulating factor-mobilized CD34(+) blood cells identifies myeloid committed progenitors and permits selection of nonimmunogenic CD40(-) progenitor cells.
CD40 antigen is a costimulatory molecule highly expressed on dendritic cells (DC) and activated B cells, which induces T-cell proliferation through the binding with CD40L receptor. In this study, we evaluated CD40 expression on normal CD34(+) blood cells and functionally characterized CD34(+)CD40(+) and CD34(+)CD40(-) cell subsets. CD40, CD80, and CD86 antigens were constitutively expressed on 3.2% +/- 4.5%, 0%, and 1.8% +/- 1.2% CD34(+) blood cells, respectively. However, after 24 hours in liquid culture with medium alone, or with tumor-necrosis-factor-alpha (TNF-alpha), or with allogeneic mononuclear cells 10.8% +/- 3.8%, 75.3% +/- 15.0% and 53. 7% +/- 17.0% CD34(+) blood cells, respectively, became CD40(+). After incubation for 24 hours with TNF-alpha CD34(+)CD40(+) blood cells expressed only myeloid markers and contained less than 5% CD86(+) and CD80(+) cells. Also, a 24-hour priming with TNF-alpha or ligation of CD40 significantly increased the CD34(+) blood cells alloantigen presenting function. Finally, purified CD34(+)CD40(+) blood cells stimulated an alloreactive T-cell response in MLC, were enriched in granulocytic, monocytic, and dendritic precursors, and generated high numbers of DC in 11-14 d liquid cultures with GM-CSF, SCF, TNF-alpha and FLT-3L. In contrast, CD34(+)CD40(-) cells were poorly immunogenic, contained committed granulocytic and erythroid precursors and early progenitors, and differentiated poorly toward the DC lineage. In conclusion, a short incubation with TNF-alpha allows the selection of CD40(+) blood progenitors, which may be a useful source of DC precursors for antitumor vaccine studies, and also a CD34(+)CD40(-) blood cell fraction that could be exploited in innovative strategies of allogeneic transplantation across HLA barriers.
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Institute of Hematology and Medical Oncology, "Seràgnoli," University of Bologna, Bologna, Italy. drond@med.unibo.it
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D000328:Adult; D015703:Antigens, CD / Q000096:biosynthesis / Q000097:blood*; D018952:Antigens, CD34 / Q000097:blood*; D019013:CD40 Antigens / Q000096:biosynthesis / Q000097:blood*; D002478:Cells, Cultured; D018920:Coculture Techniques; D003114:Colony-Forming Units Assay; D015672:Erythroid Precursor Cells / Q000166:cytology; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006098:Granulocytes / Q000166:cytology; D006133:Growth Substances / Q000494:pharmacology*; D019650:Hematopoietic Stem Cell Mobilization / Q000379:methods; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000502:physiology; D008213:Lymphocyte Activation; D007959:Lymphocyte Culture Test, Mixed; D008565:Membrane Proteins / Q000494:pharmacology; D019089:Stem Cell Factor / Q000494:pharmacology; D013601:T-Lymphocytes / Q000276:immunology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D015703:Antigens, CD; D018952:Antigens, CD34; D019013:CD40 Antigens; D006133:Growth Substances; D008565:Membrane Proteins; D019089:Stem Cell Factor; D014409:Tumor Necrosis Factor-alpha; C084881:flt3 ligand protein; D016179:Granulocyte Colony-Stimulating Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor