Matrix metalloproteinase-9-mediated type III collagen degradation as a novel serological biochemical marker for liver fibrogenesis

Background: During fibrogenesis in the liver, in which excessive remodelling of the extracellular matrix (ECM) occurs, both the quantity of type III collagen (CO3) and levels of matrix metalloproteinases (MMPs), including MMP-9, increase significantly. MMPs play major roles in ECM remodelling, via their activity in the proteolytic degradation of extracellular macromolecules such as collagens, resulting in the generation of specific cleavage fragments. These neo-epitopes may be used as markers of fibrosis. Aims: The current study investigated whether a novel enzyme-linked immunosorbent assay (ELISA) assay specifically measuring an MMP-9-cleaved sequence of type III collagen located at position 610 (CO3-610C) may be used as a marker of liver fibrosis. Material and methods: Bile duct ligation (BDL) was performed in 20 rats, with sham operations performed on another 20 rats. Serum levels of the neoepitope CO3-610C (MMP-mediated type III collagen degradation) were determined with an ELISA at 14 and 28 days post-surgery. Liver fibrosis was evaluated by quantitative digital image analysis of Sirius red-stained formalin-fixed and paraffin-embedded sections. Western blot and densitometry were performed to confirm the CO3-610C ELISA data. Results: CO3-610C levels in serum increased significantly in BDL rats compared with those undergoing sham operations (% increase: 14 days = 153%, P < 0.0001; 28 days = 134%, P = 0.0014). This increase was confirmed by Western blot and densitometry of the identified bands. The CO3-610C levels correlated to liver fibrosis (R-2 = 0.23 and P = 0.01), as evaluated by quantitative digital histology. Discussion and conclusion: The data suggest that MMP-9-mediated CO3 turnover is a central event in the pathogenesis of fibrosis, and that the neo-epitope generated may be a novel biochemical marker.