Figure 2.

rMMP-3 proteolytic domain induces EMT in NRK52e cells and GM6001 blocks this induction. A–C: Phase-contrast images of NRK52e cells untreated (ctr) (A: Scale bar = 25 μm) or treated with rMMP-3 (2 μg ml⁻¹) for 72 hours in absence (B) or presence (C) of GM6001 (25 μmol/L). D–O: Immunofluorescence images showing E-cadherin (D–F: Scale bar = 15 μm), α-SMA (G–I), β-catenin (J–L) and HSP47 (a collagen-binding stress protein that always coexists with collagen) (M–O) of NRK52e cells with corresponding treatments. P–R: Phase contrast images of confluent NRK52e cells untreated (Ctr) (P: Scale bar = 25 μm), or treated with rMMP-3 2 μg ml⁻¹ (Q) and 5 μg ml⁻¹ (R) for 72 hours. S: Western blot using E-cadherin N-terminal Ab (N-20) of 80-kd proteolytic E-cadherin ectodomain present in the medium of NRK52e cells treated with rMMP-3 in absence or presence of GM6001 (25 μmol/L). Equal amounts of protein were loaded in each lane and checked by Ponceau S staining. T and U: Western blot analysis of E-cadherin and α-SMA versus β-actin in lysates of NRK52e cells treated with rMMP-3 in absence or presence of GM6001. V and W: Time course of E-cadherin and α-SMA protein levels in NRK52e cells during treatment with rMMP-3. *P < 0.05.