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Development of Mouse Dendritic Cells from Lineage-Negative c-kit^<low Pluripotent Hemopoietic Stem Cells In Vitro

First Department of Pathology, Kansai Medical University, Moriguchi City, Osaka, Japan

Key Words. c-kit^<low • Dendritic cells • Hemopoietic stem cells

Susumu Ikehara, M.D., Ph.D., First Department of Pathology, Kansai Medical University, 10-15 Fumizono-cho, Moriguchi City, Osaka 570-8506, Japan. Telephone: 81-6-6993-9429; Fax: 81-6-6994-8283; e-mail: ikehara{at}takii.kmu.ac.jp

Dendritic cells (DCs) are essential for the presentation of antigens in the primary immune response. To examine the generation of DCs from hemopoietic stem cells in the bone marrow (BM), lineage-negative (Lin^–)/CD71^– bone marrow cells (BMCs) from C57BL/6 mice were separated into major histocompatibility complex (MHC) class I^high/ c-kit^low and MHC class I^high/c-kit^<low (phenotypically c-kit-negative, but c-kit message only detected by reverse transcriptase-polymerase chain reaction) populations. A large number of cells with the morphological, phenotypical, and functional characteristics of DCs was generated from both c-kit^low and c-kit^<low populations when cultured with a combination of cytokines (GM-CSF, tumor necrosis factor-a [TNF-a], interleukin 7 [IL-7], IL-3, stem cell factor [SCF], and flt3 ligand); the cytokine combination studies revealed that SCF and IL-3 in addition to GM-CSF and TNF-a are essential for DCs to be generated from these primitive populations. To our surprise most (>80%) generated cells expressed high levels of DC surface markers such as DEC205 and MHC class II, and they were potent stimulators in the primary allogeneic T cell activation. The development of DCs from c-kit^<low cells was slower than that from c-kit^low cells. These results indicate that c-kit^<low cells are more primitive than c-kit^low cells, although both c-kit^low cells and c-kit^<low cells can differentiate into DCs. It should be noted that the combination of these cytokines selectively induces DCs from both c-kit^<low and c-kit^low cells in vitro, suggesting that the ex vivo expansion of DCs using these primitive cells would be applicable to immunotherapy.

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