Fig. 5

Functional consequences of inhibition of RhoA. a F-actin polymerisation in RhoA siRNA transfected MDA-MB231. Two days after si-RNA transfection, MDA-MB-231 were grown on glass bottom slides and actin cytosqueletton was revealed by a phalloïdin-fluorescein (1 μg/mL) labelling. Pictures present fluorescence microscope series of adherent MDA-MB-231 transfected with non-specific siRNA (ns-SiRNA), RhoA-specific (RhoA si-RNA) unstimulated or stimulated with SDF-1α (100 ng/mL). RhoA inhibition reverted the increased of stress fiber in the treated sample. b RhoA specific si-RNA transfected MDA-MB-231 plasticity on Matrigel. Two days after transfection, with non-specific si-RNA (ns-SiRNA) or RhoA specific (RhoA si-RNA) MDA-MB231 were seeded on a 96-wells plate, coated with Matrigel. Microscopic pictures of cellular networks after SDF-1α stimulation (100 ng/ml) were taken after 18 h of culture. Quantitative evaluation of the cellular interconnection is presented. The evaluation was made by counting the number of cellular interconnections on 10 different fields. RhoA inhibition reversed the interconnection number increase in the treated sample. c Wound Closure assay. Two days after transfection, with non-specific si-RNA (ns-SiRNA) or RhoA specific (RhoA si-RNA) MDA-MB231 migration ability was tested after a scratch with or without SDF-1α (100 ng/ml). RhoA inhibition supressed the effect of SDF-1α on MDA-MB231 motility. d Cell cycle analysis. Two days after transfection, with non-specific si-RNA (ns-SiRNA) or RhoA specific (RhoA si-RNA) MDA-MB231 were treated with or without SDF-1α (50 ng/ml) for 48 h and position in cell cycle were evaluated with NIM-DAPI by flow cytometry. The inhibition of RhoA doesn’t have any effect on the position of the cell cycle position of MDA-MB231. The results presents in this figure are representative of three different experiments. e Adherence of MDA-MB231 RhoA specific si-RNA transfected cells to BMHC. Stable eGFP-MDA-MB231 cells were seeded on the plate and allow to adhere for one hour. As displayed Si-RhoA transfected cancer cells displayed significantly increased adhesion compared to controls