Fig. 6

Tumor cell invasion is favored by the presence of EDA+ fibronectin in the 3D ECM. A MDA-MB-231 cell oriented migration depends on the presence of EDA+ fibronectin in the 3D ECM. Cell-tracker labeled MDA-MB-231 tumor cells were seeded on top of decellularized 3D ECMs generated by the indicated MEFs in the absence or presence of 5 ng/ml TGFβ₁. Cell migration was recorded overnight by taking IF images every 15 min using life microscopy (Additional file 1: Fig. S8A). MDA cell movement was tracked using ImageJ software, and displacement features, such as the angle of each displacement, were measured. Oriented migration was plotted as the percentage of cell movements in the maximum orientation (up to 21˚ deviation from the mode). B MDA-MB-231 cell invasion is increased on 3D ECMs with EDA+ fibronectin. The indicated MEF lines were allowed to produce 3D ECMs in the presence of 5 ng/ml TGFβ₁ on invasion-insert membranes. ECMs were decellularized, and MDA cells (in DMEM with 0.1% FBS) were seeded on top. DMEM with 10% FBS was added to the lower chamber as a chemoattractant. Cells were allowed to invade for 16 h and fixed with 4% PFA. Invading cells were stained with DAPI and quantified. C MDA-MB-231 cell invasion through EDA+ fibronectin matrices is interfered by irigenin treatment during matrix formation. MDA invasion through decellularized 3D ECMs produced by the indicated MEF lines activated with 5 ng/ml TGFβ₁ and either treated or not with 50 μM irigenin, was quantified as in B. D EpRas invasion is increased on 3D ECMs containing EDA+ fibronectin. Invasion insert membranes were covered with indicated 3D ECMs as described in B and EpRas were induced to invade decellularized ECMs for 48 h and quantified as in B