MT1-MMP directs the 3D collagen-invasive activity of hMSCs. (A) hMSCs (2 × 105) electroporated with Scr siRNA, MMP-directed siRNAs, or cotransfected with MT1-MMP siRNA and either a control expression vector (MT1siRNA + Vec) or mouse MT1-MMP expression vector (MT1siRNA + mMT1) were cultured within 3D type I collagen gels (2.2 mg/mL) in growth medium for 4 days. In the bottom row of panels, hMSCs in 3D collagen gels were cultured with GM6001 (25μM), TIMP-1 (7.5 μg/mL), or TIMP-2 (2.5 μg/mL). For reference, the arrow to the left of the panels in the top row marks the edge of the embedded island of hMSCs. Arrowheads mark cells migrating into surrounding collagen from the central collagen island. Images shown are representatives of 3 experiments performed. Bar, 100 μm. (B) hMSCs transfected with Scr or MT1-MMP siRNAs were cultured in 3D collagen for 4 days and stained with phalloidin (red) as well as monoclonal antibody 9A4 directed against a type I collagen cleavage neoepitope (green) and visualized by fluorescence microscopy. Bar, 50 μm. (C) The number of invading cells and invasion depth in the 3D collagen gels are expressed as the mean ± SD in 5 randomly selected fields in a single representative experiment of 3 or more performed.
