Figure 7. MMP-2 cleavage of Vn and FN increases OvCa adhesion.

(A) Human omentum and peritoneum stained with Vn- or FN-specific antibodies (left panel). HPMCs were subjected to immunoblotting (right panel) using Vn- and FN-specific antibodies. Lane 1, HPMCs. HT-1080 CM was used as positive control. Original magnification, ×400. (B) Full-length FN was incubated with activated MMP-2, and fragments were resolved on a 10% Tris-HCl gel by silver stain analysis. (C) Adhesion assay of SKOV3ip1 cells to full-length and MMP-2–cleaved Vn or FN coated plates as described in Figure 2. (D) FN and MMP-2–cleaved FN were run on a native gel and transferred to nitrocellulose. An adhesion assay was performed with 1.0 × 10⁷ SKOV3ip1 cells for 4 hours, the membrane was washed, fixed, and bound cells were stained. (E) Competition assays were conducted. SKOV3ip1 cells were preincubated with full-length Vn, MMP-2–cleaved Vn, full-length FN, or MMP-2–cleaved FN, and an adhesion assay to 3D coculture was conducted. (F) SKOV3ip1 cells were transfected with a siRNA specific for α₅ or β₃ integrin, and adhesion assay was performed to MMP-2–cleaved Vn or FN coated plates. (G) Fluorescently labeled SKOV3ip1 cells were pretreated with either an MMP-2 or a mouse isotype IgG antibody followed by treatment with α₅, β₁, α_vβ₃, β₄ integrin, or mouse isotype IgG antibodies. Subsequently, an adhesion assay was performed on the 3D model. *P < 0.01, **P < 0.001. Each graph and picture is representative of 3 independent experiments. Scale bar: 100 μm.