A novel assay for extracellular matrix remodeling associated with liver fibrosis: An enzyme-linked immunosorbent assay (ELISA) for a MMP-9 proteolytically revealed neo-epitope of type III collagen

ObjectivesAccumulation of extracellular matrix (ECM) components and increased matrix-metalloprotease (MMPs) activity are hallmarks of fibrosis. We developed an ELISA for quantification of MMP-9 derived collagen type III (CO3) degradation.Design andmethodsA monoclonal antibody targeting a specific MMP-9 cleaved fragment of CO3 was used for development of a competitive ELISA. The assay was investigated in serum and tissues from bile duct ligated rats (BDL).ResultsThe ELISA showed no cross-reaction with either intact CO3, or other collagens. The intra- and inter-assay CV were below 10%. Liver fibrosis was demonstrated in BDL animals by semi quantitative scoring (P < 0.0001). Serum levels of CO3-610 increased 2.5 fold in BDL animals (P < 0.001). The CO3-610 levels were 5 fold higher in ex vivo cultures of fibrotic livers compared to controls (P < 0.001).ConclusionWe have developed a novel ELISA for measuring a specific fragment CO3 generated by MMP-9 important in pathogenesis of liver fibrosis.