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Development of dendritic cells in vitro from murine fetal liver–derived lineage phenotype-negative c-kit +hematopoietic progenitor cells by Yanyun Zhang, Yi Zhang, Yong Wang, Masafumi Ogata, Shin-ichi Hashimoto, Nobuyuki Onai, and Kouji Matsushima Blood Volume 95(1):138-146 January 1, 2000 ©2000 by American Society of Hematology Schematic representation of culture conditions of purified murine FL-derived Lin− c-kit + HPCs. Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology The culture of murine 13 dpc FL-derived Lin− c-kit + HPCs.(A) Using a phase contrast microscope, morphologic analyses were performed on cultured murine 13 dpc FL-derived Lin− c-kit + HPCs stimulated with GM-CSF + SCF + Flt3L + mTNFα at day 7, 14, and 21. Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology Generation of DCs from 13dpc FL-derived Lin− c-kit + HPCs.13 dpc FL-derived Lin− c-kit + HPCs were cocultured with stromal cell lines PA6 or OP9 cells in the presence of GM-CSF + SCF + Flt3L for 12 to 14 days, and restimulated by GM-CSF + mTNFα for an addit... Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology Immunophenotypic and morphologic analyses of DC-like cells.13dpc FL-derived Lin− c-kit + HPCs were first supplemented with PA6 cells + GM-CSF + SCF + Flt3L for 12 to 14 days and then stimulated with GM-SCF + mTNFα for an additional 3 to 5 days. Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology The capacity of the cultured cells to enhance allogeneic MLR.Allogeneic MLR was performed using purified T cells (3 × 105 cells/well in 96-round-well plate) as responder cells. Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology The role of stromal cells in the induction of DCs from FL-derived Lin− c-kit + HPCs.13 dpc FLderived HPCs were cultured in the presence of cytokines, GM-CSF + SCF + Flt3L, with or without stromal cell line OP9 or PA6 cells in a transwell culture system. Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology Immunofluorescence analysis on DC precursors generated from murine FL-derived Lin− c-kit +HPCs.Immunofluorescence analysis was performed on the nonadherent cells from FL-derived HPCs cocultured with PA6 cells + GM-CSF + SCF + Flt3L for 12 to 14 days. Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology Endocytic activity of FL HPC-derived DC precursors and mature DCs.The capacity of FITC-DX uptake was analyzed by a cell sorter as described in “Materials and Methods.” The FITC intensity of cells as a control or the test of FITC-DX uptake was indicated. Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology Generation of NK1.1+ cells from FL-derived Lin− c-kit + HPCs.13 dpc FL-derived Lin− c-kit + HPCs were cocultured with PA6 cells in the presence or absence of SCF + Flt3L with or without GM-CSF in a transwell culture system. Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology Immunophenotypic analysis on NK1.1+ cells generated from FL-derived Lin− c-kit + HPCs.Lin− c-kit + HPCs from 13 dpc FL were cultured in the presence of PA6 cells + SCF + Flt3L for 14 days. Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology Capacity to generate DC precursors from NK1.1+ cells.13 dpc FL-derived HPCs were co-cultured with PA6 cells + SCF + Flt3L in transwell plates. Yanyun Zhang et al. Blood 2000;95:138-146 ©2000 by American Society of Hematology
