Figure 1.

Plasmin induces fibroblast apoptosis in association with fibronectin proteolysis. (A) IMR-90 fibroblasts were cultured to 60% confluence and growth arrested for 24 hours before treatment with plasmin (doses indicated) for 16 hours. Equal amounts of protein from the cell culture supernatant were subjected to SDS-PAGE and Western immunoblotting for fibronectin. (B) Similarly cultured IMR-90 fibroblasts were treated with plasmin (0.5 U/ml) ± TGF-β1 (2.0 ng/ml) and cell culture supernatants were assessed for fibronectin. (C) IMR-90 fibroblasts were cultured to 60% confluence in a 96-well plate, growth arrested in serum-free media for 24 hours, and treated with plasmin for 16 hours. Apoptosis was assessed by enzyme-linked immunosorbent assay (ELISA) for ssDNA. *P < 0.001 compared with control (n = 4). (D) IMR-90 fibroblasts cultured as above were treated with plasmin (0.1 U/ml) for the times noted and whole cell lysates were assessed for cleaved caspase-3 by SDS-PAGE and Western immunoblotting. Membranes were stripped and probed for GAPDH. (E) IMR-90 fibroblasts were treated with plasmin ± TGF-β1 (2.0 ng/ml) for 16 hours, and lysates were assessed for cleaved caspase-3 by SDS-PAGE and Western immunoblotting. Membranes were stripped and probed for GAPDH. Data are representative of at least three independent experiments.