Fig. 2

The combination of maraviroc and tocilizumab inhibits MDA-MB-231-LN cell migration. a Migration assay of MDA-MB-231-LN cells (top chamber) in treatment with maraviroc (M), tocilizumab (T), and the combination of maraviroc and tocilizumab (M+T) with 180 ul of conditioned medium (CM) (bottom chamber) from lymphatic endothelial cells (LECs) cultured with serum-free medium (SFM) or tumor-conditioned medium (TCM) of MDA-MB-231-LN cells in the RTCA system. The cell index was measured continuously for 48 h. The representative migration is shown (*P < 0.001, n = 3). b Schematic diagram of (TCM-LEC)CM. CM was prepared by growing LECs in 30% SFM or TCM of MDA-MB-231-LN cells with treatment with either vehicle or tocilizumab for 4 days and the medium was replaced with 3 ml SFM with 2% FBS. After 48 h, the supernatant was centrifuged and filtered. The conditioned medium, (TCM-LEC)CM was used for the migration assay. c Migration assay of MDA-MB-231-LN cells pre-labeled with Cell Tracker Green and the migration was measured using the Oris cell migration kit. Labeled MDA-MB-231-LN cells (50,000) in complete medium were added to each well of a 96-well plate containing stoppers to prevent the cells from settling in the center region of the wells. Cells were allowed to adhere for 24 h, after which the stoppers were carefully removed. CM as described in b was added, and the cells that migrated to the center of the well were observed for 48 h. d ELISA of human CCL5 (Quantikine ELISA, R&D System) in the CM described in b (*P < 0.001, n = 3). Veh, Vehicle