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Blood, Vol. 113, Issue 15, 3503-3511, April 9, 2009
ATM-ATR–dependent up-regulation of DNAM-1 and NKG2D ligands on multiple myeloma cells by therapeutic agents results in enhanced NK-cell susceptibility and is associated with a senescent phenotype
Blood Soriani et al.
113: 3503
Supplemental materials for: Soriani et al
Files in this Data Supplement:
- Figure S1. DNAM-1 and NKG2D ligands are up-regulated by therapeutic treatment (JPG, 97.5 KB)
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MM cell lines were treated for 48 h with different therapeutic agents at doses selected by the MTT proliferation assay. In the figure we report data relative to PVR and MICA expression on treated (dashed lines) or untreated cells (solid lines) as assessed by immunofluorescence and flow cytometry. The grey histogram represents the isotype control antibody. Similar results were obtained upon 24 h drug-treatment.
- Figure S2. Dose-response curve for PVR, MICA and MICB surface expression on SKO-007(J3) cell line treated with doxorubicin or melphalan (JPG, 93.3 KB)
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SKO-007(J3) cells were treated with different doses of doxorubicin or melphalan for 48 h. The expression of PVR, MICA and MICB was analyzed by immunofluorescence and flow cytometry. On the left y-axis is shown the median fluorescence intensity (MFI) values of ligand expression obtained by subtracting the MFI of the isotype control antibody whether on the right y-axis is shown the percentage of viable cells assessed by PI staining.
- Figure S3. Modulation of DNAM-1 and NKG2D ligand expression on patient-derived PCs following therapeutic treatment (JPG, 100 KB)
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(A) Patients were classified according to Durie & Salmon’s staging system. (B) Malignant PCs from BM samples were selected by gating on CD38+CD138+ cells and were analyzed by immunofluorescence and flow cytometry for NKG2D and DNAM-1 ligand expression. The MFI of specific ligand subtracted for MFI of isotype control gated on CD38+CD138+ malignant PCs population is reported. (C) Ligand surface expression was analyzed as above on untreated and 48h drug-treated PCs. The percentage of CD38+CD138+ cells expressing the indicated ligand before and after the drug-treatment is shown. The corresponding increase in mRNA levels has been tested after 24 h by real-time PCR performed as described in the Materials and Methods. The relative mRNA amount for the different ligands was evaluated on untreated (□) and melphalan (▒) (20 µM) or bortezomib (■) (5 nM) treated PCs. The mRNA levels have been also assessed on PCs (P7 and P10) treated for 24 h with doxorubicin (▨) (0.05 µM). Data expressed as arbitrary units were normalized with β-actin, and referred to untreated cells considered as calibrator. Data are presented as the means plus or minus standard deviations from triplicates.
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