Figure 3.

E/P-selectins and their ligand PSGL-1 are crucial for MΦ-mediated protection of MM cells. (a) Expression of PSGL-1 on human MM cell lines and primary MM cells, and expression of E/P-selectins on monocyte-derived MΦs and primary MM–BM MΦs by flow cytometry analysis. Percentage of melphalan (15 μM)-induced apoptotic MM cells (ARP-1, ARK, CAG, or primary MM cells from three patients’s) in direct coculture with MΦs in the presence or absence of (b) P/E-selectin-blocking antibody (αP/E sel, 20 μg/ml) or (c) PSGL-1-blocking antibody (αPSGL-1, 20 μg/ml). Equal amounts of control IgG were used as controls. (d) Melphalan (15 μM)-induced apoptotic, PSGL-1-knocked down (PSGL-1 shRNA) ARP-1 or ARK cells in coculture with MΦs. Wild-type (control) and control shRNA-transfected (control shRNA) cells served as controls. (e) Melphalan-induced apoptotic MM cells (ARP-1, ARK or CAG) in the presence of E-selectin–Fc fusion protein (E-sel-Fc, 2 μg/ml). (f) Expression of E-selectin on monocytes and monocyte-derived MΦs from 10 blood donors by flow cytometry analysis. Numbers indicate the mean values of mean fluorescent intensity (MFI). Summarized results from four independent experiments are shown. *P<0.05, **P<0.01.