Fig. 1

Endothelial cells influence the phenotype and migratory potential of cancer cells. a CFSE fluorescently labelled cancer cell lines were plated on a tight HUVEC monolayer on the inner surface of a Boyden migration chamber for migration through the endothelial cell layer. After 48 h, the cells remaining in the insert of the chamber were removed, and the number of migrated cancer cells was quantified by fluorescence microscopy and compared to its own control (migration in absence of endothelial monolayer) . All experimental conditions were tested in three replicates and each condition in biological triplicates. (***p ≤ 0.0001, *p ≤ 0.05 versus control by unpaired student’s t-test, error bar: standard error of the mean). b Different cancer cell lines were exposed to human umbilical vein endothelial cell (HUVEC)-conditioned medium for 4 days. After exposure to the HUVEC supernatant, cancer cell migration was evaluated using a Boyden chamber migration assay. The migration rate of each cell line exposed to the HUVEC-conditioned medium was compared to its own control in non-conditioned medium (**p ≤ 0.01, *p ≤ 0.05 versus ctrl by unpaired student’s t-test). c SK-BR-3 cell morphology was visualized by phase contrast imaging (top) after exposure to unconditioned or HUVEC-conditioned medium. Expression of actin stress fibres (phalloidin), focal adhesions (paxillin) and tight junctions (ZO-1) was analysed by immunofluorescence staining (middle, bottom). d SK -BR-3 cells were exposed to unconditioned (ctrl) or conditioned media derived from confluent or subconfluent HUVEC and HDMEC cultures for 4 days. The effect on the migration of SK-BR-3 cells was evaluated using a Boyden chamber assay (**p ≤ 0.05, ***p ≤ 0.001 versus ctrl by unpaired student’s t-test, error bar: standard error of the mean). e Expression of fibronectin (FN1) was detected by immunofluorescent staining of SK-BR-3 cells after 4 days of exposure to unconditioned (ctrl) or conditioned medium derived from confluent or subconfluent HUVEC cultures