The vacuolar-type ATPase inhibitor archazolid increases tumor cell adhesion to endothelial cells by accumulating extracellular collagen
Archazolid increases the amount of extracellular collagen on HUVECs. (A) 24-well plates were coated with 10 μg/ml collagen (col), fibronectin (fn) or laminin (ln) or were left uncoated (co). Untreated MDA-MB-231 or PC-3 cells were stained with CellTracker Green CMFDA Dye and added into ECM-coated or uncoated wells. After 10 min of incubation, non-adherent cells were washed off. Adhesion was quantified by measuring the fluorescence signal using a plate reader. Data are expressed as mean ± SEM (n = 3). *p ≤ 0.05 versus uncoated control. (B) A cell adhesion assay onto collagen was performed with untreated MDA-MB-231 or PC-3 cells (co) and MDA-MB-231 or PC-3 cells on which the integrin β1 subunit was blocked by a monoclonal antibody (ab). Data are expressed as mean ± SEM (n = 3). *,#p ≤ 0.05 versus control. (C) Confluent HUVECs were treated with 1 nM archazolid (arch) or DMSO (co) for 24 h. Surface collagen (green) was detected by immunofluorescence staining of viable cells and nuclei (blue) were visualized by Hoechst 33342 staining. Scale bar represents 20 μm. One representative image out of three independently performed experiments is shown. The increase in extracellular collagen was quantified. Data are expressed as mean ± SEM (n = 3). *p ≤ 0.05 versus DMSO control.