Fig. 3.

Adhesion and migration of wt- and ko-MCA tumor cells. a Tumor cells were seeded on matrix protein-coated plates. After 4 h incubation and washing, the percent of adherent cells was evaluated by crystal-violet staining measuring OD585; b MCA cells were seeded on BSA-coated plates, the medium containing 10% Exo-depleted FCS or 30 μg/ml wt-sExo or -TEX. After 4 h incubation and washing, the percent of adherent cells was evaluated by crystal-violet staining measuring OD585; a, b mean values±SD (triplicates), significant differences between wt- and ko-cells: *; significant differences between BSA- versus matrix protein-coated plates and between Exo-depleted FCS and sExo or TEX: s; c transwell migration of wt- and ko-MCA cells towards BSA versus 20% FCS; mean % migrating cells±SD (triplicates); significant differences in transwell migration between wt- and ko-MCA tumor cells: *; d in vitro wound healing of wt- and ko-MCA cells was evaluated by the scratch assay. After reaching subconfluence, the monolayer was scratched and wound healing was followed microscopically for 48 h–72 h. Representative examples of wt- and ko-MCA cells seeded on coll I-coated plates (wound edge are indicated by a dashed line) and on BSA-, coll I-, coll IV-, LN111-, LN332- or FN-coated plates; the mean % wound area ± SD (3 assays) are shown; the mean % differences in the wound area compared to BSA-coated plates are listed in the table, significant differences are shadowed; e transwell migration of wt- and ko-MCA cells towards BSA versus wt-sExo and wt- and ko-TEX was evaluated in Boyden chambers after 16 h of incubation; mean % migrating cells±SD (triplicates); significant differences in migration towards BSA versus sExo and TEX: s; f wound healing of ko-MCA cells seeded on BSA-coated plates was evaluated in the presence of medium containing 10% Exo-depleted FCS or wt- and ko-TEX (30 μg/ml) after 24 h; the mean % wound area ± SD (triplicates), significant differences between Exo-depleted FCS versus TEX containing medium: s. Transwell migration and in vitro wound healing of ko-, most strongly dbko-MCA cells is impaired, the difference in wound healing being maintained in matrix protein-coated plates, where distinct to Tspan8ko- and CD151ko-MCA cells, dbko-MCA only migrate on FN-coated plates. Migration is promoted by wt-TEX and, weakly, by Tspan8ko- or CD151ko-TEX in CD151ko-, respectively, Tspan8ko-MCA cells