Measurement of CO3-610, a Potential Liver Biomarker Derived from Matrix Metalloproteinase-9 Degradation of Collagen Type III, in a Rat Model of Reversible Carbon-Tetrachloride-Induced Fibrosis

Abstract

Background and aim:The current study utilized a carbon tetrachloride (CCl4)-induced liver fibrosis model to measure levels of the MMP9-mediated collagen type III degradation fragment CO3-610 (site of cleavage: KNGETGPQGP), during disease progression and regression, and to investigate a potential prognostic role of the biomarker.Materials and methods:72 female Sprague-Dawley rats aged 6 months old were injected with CCl4 twice a week over different periods of time to induce varying degrees of liver fibrosis. After 4, 6 and 8 weeks of treatment, administration of CCl4 was stopped. The 6- and 8-week treatment groups were left to regress for a further 6 or 12 weeks at which point they were sacrificed and livers removed and sectioned. Liver fibrosis was quantified using Visiopharm software to analyse Sirius red-stained sections. Serum levels of CO3-610 were measured in all animals using an ELISA assay as described by Barascuk et al.1Results:Quantitative histology revealed total collagen deposition in the liver increased as fibrosis progressed. In animals treated with CCl4 for 4 weeks, collagen comprised on average 4.94% of the total tissue in liver sections, while after 6 weeks the mean was 8.25%, and after 8 weeks, 9.11%. During the regression phase, the total collagen deposition gradually decreased to a mean of 6.9% and 5.09% for animals regressing 6 and 12 weeks respectively after 6 weeks treatment, and 6.27% for animals regressed 12 weeks after 8 weeks treatment. CO3-610 values increased progressively in rats treated for 4 weeks (by a mean of 55.0 ng/ml), 6 weeks (mean 61.1 ng/ml) and 8 weeks (mean 70.2 ng/ml). During the regression phase, CO3-610 values rapidly decreased by a mean of 28.9 ng/ml at 6 weeks and 21.6 ng/ml at 12 weeks in animals previously treated for 6 weeks, and by a mean of 19.52 ng/ml in animals treated for 8 weeks and regressed for 12 weeks. CO3-610 levels were statistically significantly correlated with total collagen during disease progression (r = 0.5701, P < 0.0001). No statistically significant correlation was observed during regression (r = 0.2081, P = 0.1138).Conclusion:Levels of the MMP-9 generated fragment of collagen type III, CO3-610, correlated with the degree of liver fibrosis in rats during the progression phase, but were not correlated with total collagen levels during regression. CO3-610 seems to be produced only under the CCL4 stimulus, and signifies CO3-610 as a potential marker of progression rather than regression. The corresponding steep elevations in levels of CO3-610 total collagen and collagen type III during liver fibrosis progression underline a potential prognostic capacity of the biomarker.

Serum markers of the extracellular matrix remodeling reflect antifibrotic therapy in bile-duct ligated rats

“…They may carry unique disease “fingerprints,” why this approach is coined Protein Fingerprint Technology (Karsdal et al, 2009, 2012; Barascuk et al, 2010; Vassiliadis et al, 2011; Veidal et al, 2011a,b; Leeming et al, 2012, 2013). These markers have been validated in different models and severities of fibrosis in previous studies and compared to control non-fibrotic rats (Karsdal et al, 2009, 2012; Barascuk et al, 2010; Vassiliadis et al, 2011; Veidal et al, 2011a,b; Leeming et al, 2012, 2013). …”

Novel insights into the function and dynamics of extracellular matrix in liver fibrosis

“…Fibrosis is also a dynamic condition with accelerated ECM turnover in which both tissue formation and tissue degradation are highly upregulated (15,16,297,350), resulting in pathological collagen deposition and altered MMP expression profiles (353). Moreover, modifications to amino acids or proteolytic cleavage at specific locations by specific PTMs result in both immunologically and functionally different proteins (161).…”

Extracellular Matrix Remodeling: The Common Denominator in Connective Tissue DiseasesPossibilities for Evaluation and Current Understanding of the Matrix as More Than a Passive Architecture, but a Key Player in Tissue Failure

“…57 Type III collagen has been mostly assiociated with various fibrotic diseases. [58][59][60][61] Type IV collagen is the main component of the BM, a specialized type of ECM that separates the epithelium from the stroma in all tissues in the body. It consists of three domains: N-terminal 7S domain, a central triple helix, and a large C-terminal NC1 globular domain.…”

“…atherosclerosis (type I and III collagens, titin and versican) 217 . various fibrotic diseases including liver (type I, III, IV, V, VI collagens and biglycan), [59][60][61]144,196,197,199,200,218 lung (elastin, type I, III, and V collagen), 74,[220][221][222][223][224][225][226][227][228][229][230][231][232] and kidney. 233 Key lessons on the importance of the structural components of the matrix may be harvested from the genetic mutations that lead to pathologies.…”