Generation of large numbers of dendritic cells from mouse bone marrow cultures supplemented with granulocyte-macrophage colony stimulating factor

Inaba, K., Inaba, M., Romani, N., Aya, H., Deguchi, M., Ikehara, S., Muramatsu, S., and Steinman, R.M. Generation of large numbers of dendritic cells from mouse bone marrow cultures supplemented with granulocyte-macrophage colony stimulating factor. J. Exp. Med. 176: 1693-1702, 1992https://digitalcommons.rockefeller.edu/historical-scientific-reports/1033/thumbnail.jp

Dendritic cell progenitors phagocytose particulates, including bacillus calmette-guerin organisms, and sensitize mice to mycobacterial antigens in vivo

Dendritic cells, while effective in sensitizing T cells to several different antigens, show little or no phagocytic activity. To the extent that endocytosis is required for antigen processing and presentation, it is not evident how dendritic cells would present particle-associated peptides. Evidence has now been obtained showing that progenitors to dendritic cells can internalize particles, including Bacillus Calmette-Guerin (BCG) mycobacteria. The particulates are applied for 20 h to bone marrow cultures that have been stimulated with granulocyte/macrophage colony-stimulating factor (GM-CSF) to induce aggregates of growing dendritic cells. Cells within these aggregates are clearly phagocytic. If the developing cultures are exposed to particles, washed, and “chased” for 2 d, the number of major histocompatibility complex class II-rich dendritic cells increases substantially and at least 50% contain internalized mycobacteria or latex particles. The mycobacterialaden, newly developed dendritic cells are much more potent in presenting antigens to primed T cells than corresponding cultures of mature dendritic cells that are exposed to a pulse of organisms. A similar situation exists when the BCG-charged dendritic cells are injected into the footpad or blood stream of naive mice. Those dendritic cells that have phagocytosed organisms induce the strongest T cell responses to mycobacterial antigens in draining lymph node and spleen. The administration of antigens to GM-CSF-induced, developing dendritic cells (by increasing both antigen uptake and cell numbers) will facilitate the use of these antigen-presenting cells for active immunization in sit

Unusual phenotype of B cells in the thymus of normal mice

[No abstract available

Granulocytes, macrophages, and dendritic cells arise from a common major histocompatibility complex class II-negative progenitor in mouse bone marrow

Inaba, K., Inaba, M., Deguchi, M., Hagi, K., Yasumizu, R., Ikehara, S., Muramatsu, S., and Steinman, R.M. Granulocytes, macrophages, and dendritic cells arise from a common major histocompatibility complex class II-negative progenitor in mouse bone marrow. Proc. Natl. Acad. Sci. USA. 90: 3038-3042, 1993https://digitalcommons.rockefeller.edu/historical-scientific-reports/1034/thumbnail.jp

Macrophages phagocytose thymic lymphocytes with productively rearranged T cell receptor α and β genes

[No abstract available

Granulocytes, macrophages, and dendritic cells arise from a common major histocompatibility complex class II-negative progenitor in mouse bone marrow

The developmental origin of dendritic cells, a specialized system of major histocompatibility complex (MHC) class II-rich antigen-presenting cells for T-cell immunity and tolerance, is not well characterized. Granulocyte-macrophage colony-stimulating factor (GM-CSF) is known to stimulate dendritic cells, including growth and development from MHC class II-negative precursors in suspension cultures of mouse bone marrow. Here we studied colony formation in semi-solid methylcellulose cultures, a classical bioassay system in which GM-CSF induces the formation of mixed granulocyte-macrophage colonies. When colonies were induced from MHC class II-negative precursors, a small subset (1-2%) of typical dendritic cells developed alongside macrophages and granulocytes. The dendritic cells were distinguished by their cytologic features, high levels of MHC class II products, and distinct intracellular granule antigens. By using differential adherence to plastic, enriched populations of the various myeloid cell types were isolated from colonies. Only the dendritic cells stimulated a primary T-cell immune response, the mixed leukocyte reaction, and the potency was comparable to typical dendritic cells isolated from spleen. Macrophages from mixed or pure colonies were inactive as stimulator cells. Therefore, three distinct pathways of myeloid development - granulocytes, macrophages, and dendritic cells - can develop from a common MHC class II-negative progenitor under the aegis of GM-CSF

Generation of large numbers of dendritic cells from mouse bone marrow cultures supplemented with granulocyte/macrophage colony-stimulating factor

Antigen-presenting, major histocompatibility complex (MHC) class II-rich dendritic cells are known to arise from bone marrow. However, marrow lacks mature dendritic cells, and substantial numbers of proliferating less-mature cells have yet to be identified. The methodology for inducing dendritic cell growth that was recently described for mouse blood now has been modified to MHC class II-negative precursors in marrow. A key step is to remove the majority of nonadherent, newly formed granulocytes by gentle washes during the first 2-4 d of culture. This leaves behind proliferating clusters that are loosely attached to a more firmly adherent “stroma.” At days 4-6 the clusters can be dislodged, isolated by 1-g sedimentation, and upon recuhure, large numbers of dendritic cells are released. The latter are readily identified on the basis of their distinct cell shape, ultrastructure, and repertoire of antigens, as detected with a panel of monoclonal antibodies. The dendritic cells express high levels of MHC class II products and act as powerful accessory cells for initiating the mixed leukocyte reaction. Neither the clusters nor mature dendritic cells are generated if macrophage colony-stimulating factor rather than granulocyte/macrophage colonystimulating factor (GM-CSF) is applied. Therefore, GM-CSF generates all three lineages of myeloid cells (granulocytes, macrophages, and dendritic cells). Since \u3c5 × 10 6 dendritic cells develop in 1 wk from precursors within the large hind limb bones of a single animal, marrow progenitors can act as a major source of dendritic cells. This feature should prove useful for future molecular and clinical studies of this otherwise trace cell type

Accessory cell - T lymphocyte interactions: antigen dependent and independent clustering

Inaba, K., and Steinman, R.M. Accessory cell - T lymphocyte interactions: antigen dependent and independent clustering. J. Exp. Med. 163: 247-261, 1986https://digitalcommons.rockefeller.edu/historical-scientific-reports/1018/thumbnail.jp

Monoclonal antibodies to LFA-1 and TO CD4 inhibit the mixed leukocyte reaction after the antigen-dependent clustering of dendritic cells and T lymphocytes

[No abstract available

Accessory cell-T lymphocyte interactions. antigen-dependent and -independent clustering

[No abstract available