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1.

Figure 3. Proliferation of myeloma cells in the presence of increasing number of macrophages. From: Macrophages and mesenchymal stromal cells support survival and proliferation of multiple myeloma cells.

Myeloma cell lines were cocultured in the presence of increasing number of macrophages (MQ). For both U266 (A) and NCI-H929 (B), increasing number of macrophages was associated with increased proliferation index. 1×106 PBMC was added as additional control.

Jaehyup Kim, et al. Br J Haematol. ;158(3):336-346.
2.
Figure 4

Figure 4. The effect of IL6 neutralizing antibody on proliferation of U266 cells in co-culture with stromal cells. From: Macrophages and mesenchymal stromal cells support survival and proliferation of multiple myeloma cells.

In the presence of IL6 blocking antibody, the proliferation index of myeloma cells was decreased across all conditions. The changes were more pronounced when myeloma cells were co-cultured in the presence of mesenchymal stromal cells (MSC) and macrophages (MQ).

Jaehyup Kim, et al. Br J Haematol. ;158(3):336-346.
3.

Figure 2. Proliferation assays of U266 and NCI-H929 MM cell lines. From: Macrophages and mesenchymal stromal cells support survival and proliferation of multiple myeloma cells.

The mean value was calculated from triplicate experiments and normalized by dividing with the proliferation index obtained from cell line-only control. Combined co-cultures with mesenchymal stromal cells (MSC) and macrophages (MQ)provided a more robust proliferation stimulus than either cell component alone.

Jaehyup Kim, et al. Br J Haematol. ;158(3):336-346.
4.

Figure 5. Proliferation of U266 myeloma cells in the presence of bortezomib pre-treated stromal components. From: Macrophages and mesenchymal stromal cells support survival and proliferation of multiple myeloma cells.

(A) Treatment scheme for generation of bortezomib (Bort) pre-treated macrophages (MQ) and mesenchymal stromal cells (MSC). (B) Proliferation assays of U266 multiple myeloma cell line in the presence of bortezomib pre-treated stromal components. Bortezomib pre-treatment of both macrophages and MSC resulted in decreased proliferation of myeloma cells. Different combinations of pre-treated stromal components are shown. (C) Proliferation assays of NCI-H929 multiple myeloma cell line in the presence of bortezomib pre-treated stromal components. Different combinations of pre-treated stromal components are shown.

Jaehyup Kim, et al. Br J Haematol. ;158(3):336-346.
5.

Figure 1. Schematic diagram of myeloma cell line proliferation assay and analysis. From: Macrophages and mesenchymal stromal cells support survival and proliferation of multiple myeloma cells.

(A) Experimental outline of co-culture experiments. Macrophages (MQ) are generated by culturing CD14+ monocytes for one week, then mesenchymal stromal cells (MSC) and/or multiple myeloma cell lines are added for 3 days. (B) Sample analysis of proliferation assay. CFSE-stained multiple myeloma cell lines were analysed using flow cytometry.

Jaehyup Kim, et al. Br J Haematol. ;158(3):336-346.
6.

Figure 6. The effect of mesenchymal stromal cells and/or macrophages on apoptosis of myeloma cells. From: Macrophages and mesenchymal stromal cells support survival and proliferation of multiple myeloma cells.

(A) Schematic drawing of co-culture for apoptosis testing. (B) U266 cultured on mesenchymal stromal cells (MSCs)+macrophages (MQ), macrophages alone, MSCs alone and cell line only control. (C) NCI-H929 cultured on MSCs+macrophages, macrophages alone, MSCs alone and cell line only control. (D) Primary B cell (CD19 positive cells) apoptosis assay. MSCs, macrophages and MSCs+macrophages all decreased apoptosis of co-cultured multiple myeloma cells. In the case of primary B cells, only macrophages and MSCs+macrophages combination were capable of protecting cells from undergoing apoptosis.

Jaehyup Kim, et al. Br J Haematol. ;158(3):336-346.
7.

Figure 7. Transcriptomic comparisons of distinct subsets of monocytes and macrophages. From: Macrophages and mesenchymal stromal cells support survival and proliferation of multiple myeloma cells.

Cluster analysis based on quantitative RT-PCR showed that MSC-educated macrophages (MEM) exhibit similar phenotype to BM CD14+ monocytes isolated from MM bone marrow (MM BM CD14) or MM BM CD14+ cell-derived macrophages (MM BM MDM) (A). SPP1 encodes SPP1 (osteopontin, OPN), CD274 encodes CD274 (Programmed cell death ligand 1, PDL1), TNFSF13B encodes TNFSF13B (B cell activating factor, BAFF) and TNFRSF13B encodes TNFRSF13B (Transmembrane activator and calcium-modulator and cyclophilin ligand interactor, TACI). In contrast, peripheral blood CD14+ monocytes and peripheral blood monocyte-derived macrophages (MDM) generated a separate cluster. Same data visualized using principal component analysis (B). MSC-educated macrophages shared a similar distribution with MM BM CD14-derived macrophages.

Jaehyup Kim, et al. Br J Haematol. ;158(3):336-346.

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