Figure 1
Figure 1. Myeloma promotes MGC formation by inducing cell fusion in human dendritic cells. (A) Immunofluorescense analysis of MGCs in tumor-DC cocultures. Myeloma cells (U266) were plated with or without monocyte-derived DCs at a tumor/DC ratio of 1:100 in methylcellulose cultures. Cells were harvested for cytospins after an incubation of 2 weeks. Slides were acetone-fixed and stained for CD138 (green), CD11c (red), and DAPI (blue). MGCs were counted in 10 random high-power fields. Shown is a representative area with 3 CD138−CD11c+ giant cells with varying numbers of nuclei (dotted arrows) and CD138+ tumor cells (thin arrows). Original magnification, ×40. (B) Myeloma (U266 and ARK), breast cancer (MCF-7) or mantle cell lymphoma cell line (NCEB1) were plated with and without DCs at a ratio of 1:100 in methylcellulose cultures. Cells were harvested for cytospins after an incubation of 2 weeks. Slides were acetone-fixed and stained for CD138, CD11c, and DAPI as described in panel A. The numbers of MGCs were counted in 10 random high-power fields (HPF). Results are the mean ± SEM of 3 separate experiments. *P < .05. (C) Myeloma cells (U266) were plated with either monocytes (Monocytes), GM-CSF–treated monocytes (Macrophages), or immature DCs (Mo-DCs) cultured from the same donor at a tumor/myeloid cell ratio of 1:100 in methylcellulose cultures. Cells were harvested for cytospins after 2 weeks. Slides were acetone-fixed and stained as described previously. Numbers of MGCs were counted in 10 random HPF. Results are the mean ± SEM of 3 separate experiments for the cell line. *P < .05. (D) Kinetics of MGC formation in the experimental conditions in panel C were monitored by microscopy. *P < .05. (E) Bone marrow mononuclear cells (BMMNCs) from a myeloma patient were cocultured with DCs in methylcellulose cultures at a tumor/DC ratio of 1:100. Cytospins were made after an incubation of 2 weeks. Slides were acetone-fixed and stained for CD138, CD11c, and DAPI and counted for MGCs (arrows) as described previously. Data are representative of similar experiments on 3 separate patients. Original magnification, ×20. (F) Enumeration of MGCs in BMMNC + DC cocultures from a representative myeloma patient is shown. Data are representative of similar experiments on 3 different patients. BMMNCs alone and DCs alone were used as controls.

Myeloma promotes MGC formation by inducing cell fusion in human dendritic cells. (A) Immunofluorescense analysis of MGCs in tumor-DC cocultures. Myeloma cells (U266) were plated with or without monocyte-derived DCs at a tumor/DC ratio of 1:100 in methylcellulose cultures. Cells were harvested for cytospins after an incubation of 2 weeks. Slides were acetone-fixed and stained for CD138 (green), CD11c (red), and DAPI (blue). MGCs were counted in 10 random high-power fields. Shown is a representative area with 3 CD138−CD11c+ giant cells with varying numbers of nuclei (dotted arrows) and CD138+ tumor cells (thin arrows). Original magnification, ×40. (B) Myeloma (U266 and ARK), breast cancer (MCF-7) or mantle cell lymphoma cell line (NCEB1) were plated with and without DCs at a ratio of 1:100 in methylcellulose cultures. Cells were harvested for cytospins after an incubation of 2 weeks. Slides were acetone-fixed and stained for CD138, CD11c, and DAPI as described in panel A. The numbers of MGCs were counted in 10 random high-power fields (HPF). Results are the mean ± SEM of 3 separate experiments. *P < .05. (C) Myeloma cells (U266) were plated with either monocytes (Monocytes), GM-CSF–treated monocytes (Macrophages), or immature DCs (Mo-DCs) cultured from the same donor at a tumor/myeloid cell ratio of 1:100 in methylcellulose cultures. Cells were harvested for cytospins after 2 weeks. Slides were acetone-fixed and stained as described previously. Numbers of MGCs were counted in 10 random HPF. Results are the mean ± SEM of 3 separate experiments for the cell line. *P < .05. (D) Kinetics of MGC formation in the experimental conditions in panel C were monitored by microscopy. *P < .05. (E) Bone marrow mononuclear cells (BMMNCs) from a myeloma patient were cocultured with DCs in methylcellulose cultures at a tumor/DC ratio of 1:100. Cytospins were made after an incubation of 2 weeks. Slides were acetone-fixed and stained for CD138, CD11c, and DAPI and counted for MGCs (arrows) as described previously. Data are representative of similar experiments on 3 separate patients. Original magnification, ×20. (F) Enumeration of MGCs in BMMNC + DC cocultures from a representative myeloma patient is shown. Data are representative of similar experiments on 3 different patients. BMMNCs alone and DCs alone were used as controls.

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