Fig. 8
AZD0530 treatment suppresses cancer vascular dissemination. a HUVEC layer replacement assay. Equal numbers of cancer cells were co-cultivated with confluent HUVEC cells for 24 h in the presence of AZD0530 (5 μm) or vehicle (DMSO). Cells were fixed and labeled with antibodies against pan-cytokeratin (red) and VE-cadherin (green). Scale bars, 20 μm. Blue, nuclear staining. b–d Quantification of displaced areas described in a. HUVEC + WT cancer cells (b). HUVEC + LIF+/− cancer cells (c). HUVEC + cLIF cancer cells (d). Invaded areas were calculated using CellSens imaging software (Olympus). Data are presented with scatter dot plot (mean ± SEM). Each black dot represents one captured image. **p < 0.01, ***p < 0.001, Mann–Whitney test. e Western blot analysis of p-YAP, VE-cadherin, and CD31 expression in co-cultivated cancer cells and HUVEC cells, as described in a. Total protein lysates were harvested 24 h post AZD0530 treatment using GAPDH as a loading control. f–h Quantification of disseminated tumor-like structures in zebrafish embryonic xenograft models. WT xenografts (f). LIF+/− xenografts (g). cLIF xenografts (h). The number of disseminated tumor-like structures was determined on day 4 after treatment with AZD0530 (15 μm) or vehicle (DMSO). Data are presented with scatter dot plot (mean ± SEM). Each black dot represents one fish embryo. **p < 0.01, ***p < 0.001, Mann–Whitney test
