Elevated serum aminoterminal procollagen type-III-peptide parallels collagen accumulation in rats with secondary biliary fibrosis

doi:10.1016/S0168-8278(96)80331-X

Journal of Hepatology

Volume 25, Issue 1, July 1996, Pages 79-84

https://doi.org/10.1016/S0168-8278(96)80331-X Get rights and content

Abstract

Background/Aims: The aminoterminal procollagen type-III-peptide, which can be released during collagen type III deposition, has been suggested as a serum marker of fibrogenesis in patients with chronic liver disease. However, longitudinal studies correlating procollagen type-III-peptide concentrations in serum with the evolution of liver fibrosis are still needed. The purpose of the present study was to prove the significance of procollagen type-III-peptide concentrations in serum as a non-invasive marker of liver fibrogenesis in an animal model that best resembles progressive human liver fibrosis.

Methods: In 88 female Wistar rats the biliary system was occluded by double ligation of the choledochal duct followed by retrograde injection of a mixture of prolamine/ethanole (Ethibloc®). Sixteen rats served as controls. Groups of 8–10 rats were sacrificed at days 2, 7, 14, 21, 30, 32, 35, 60 and 90 after bile duct occlusion. In the groups histological staging (fibrosis score), determination of total liver hydroxyproline, measurement of serum procollagen type-III-peptide and routine liver function tests were performed.

Results: First histological signs of liver fibrosis were seen as early as 7 days after bile duct occlusion. Progressive fibrosis was paralleled by an increase of serum procollagen type-III-peptide. There was a significant correlation between serum procollagen type-III-peptide and histological stages of fibrosis (r=0.80; p<0.0001) as well as between serum procollagen type-III-peptide and hydroxyproline in total liver tissue (r=0.85; p<0.0001)

Conclusions: These results indicate that: (1) complete bile duct occlusion in rats produces progressive hepatic fibrosis resembling human secondary biliary fibrosis, and (2) procollagen type-III-peptide concentrations in serum reflect ongoing collagen formation in the liver unrelated to serum markers of cholestasis and inflammation.

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2008, Current Therapeutic Research - Clinical and Experimental
Background: Many therapeutic strategies have been proposed to treat liver fibrosis, but no drugs have been proved effective. Matrix metalloproteinases (MMPs) have been reported to play a role in some cellular cascades of hepatic inflammation and fibrosis.
Objective: The purpose of this study was to investigate whether silymarin and pentoxifylline (PTX) have hepatoprotective and antifibrotic effects in experimental hepatic fibrosis.
Methods: Sprague-Dawley rats were divided into 4 groups: silymarin group (silymarin 4 mg/kg · d⁻¹ orally, common bile duct ligation [CBDL]); PTX group (PTX 2 mg/kg · d⁻¹ intraperitoneally, CBDL); sham group (common bile duct [CBD] exploration only); and control group (saline 1 mL/d orally, CBDL). The CBD was explored and dissected sufficiently to allow passage of a 3/0 silk suture via midline laparotomy. On day 10, all animals were euthanized via cervical dislocation. Then, 5-cm³ liver samples from the right lobe were removed for histomorphologic evaluation and 3-mL blood samples were taken via cardiac puncture for biochemical analyses. Apoptosis was determined using the terminal deoxynucleotidyltransferase-biotin nick end-label (TUNEL) staining method. Plasma levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), and γ-glutamyltransferase; total and indirect bilirubin concentration; hepatic MMP-1 and -2 and tissue inhibitor of MMP (TIMP)-l and -2 activity; and transforming-growth factor (TGF)-β₁ concentration were measured. Collagen content was determined by measuring hydroxyproline in liver samples. Malondialdehyde (MDA) was used to estimate lipid peroxidation.
Results: Thirty-two adult male Sprague-Dawley rats were divided into 4 groups: silymarin group (n = 7), PTX group (n = 7), sham group (n = 9), and control group (n = 9). Compared with the control group (14.6 [2.44]), mean (SD) hepatocyte apoptosis (as measured by the ratio of TUNEL-positive cells) was significantly suppressed in the silymarin group (1.2 [0.13]; P = 0.001) and the PTX group (3.8 [0.34]; P = 0.001). Mean (SD) MMP-2 activity in the silymarin group (57.35 [9.89] μg/mL; P = 0.04) and the PTX group (46.88 [9.56] μg/mL; P = 0.04) was significantly lower than that observed in the control group (232.32 [79.76] μg/mL). Compared with the control group (1.37 [0.38] μg/mL), TIMP-2 activity was significantly lower in the silymarin group (0.55 [0.13] μg/mL; P = 0.04) and the PTX group (0.42 [0.09] μg/mL; P = 0.01). Compared with the control group (909.17 [117.35] μg/mL), TGF-β₁ was significantly lower in the silymarin group (518.24 [30.34] μg/mL; P = 0.01) and the PTX group (519.57 [47.27] μg/mL; P = 0.01). Histomorphologic changes were significantly greater in the sham group than in the silymarin and PTX groups: hemorrhage (2.44 [0.29] vs 1.29 [0.18] and 1.57 [0.20], respectively; both, P = 0.04); sinusoidal dilatation (2.22 [0.22] vs 1.57 [0.20] and 1.71 [0.18]; both, P = 0.04); presinusoidal polymorphonuclear cell infiltration (3-44 [0.24] vs 2.57 [0.20] and 2.14 [0.26]; P = 0.03 and P = 0.008, respectively); and inflammation (3.44 [0.24] vs 2.57 [0.20] and 2.14 [0.26]; P = 0.03 and P = 0.008, respectively). In the control group, all biochemical markers were elevated, supporting the presence of liver injury. Compared with the control group (630.00 [46.80] U/L), plasma AST activity was significantly lower in the silymarin group (443.11 [78.73]; P = 0.04) and the PTX group (349.42 [34.00]; P = 0.03). Compared with the control group (191.12 [32.93] U/L), plasma ALT activity was significantly lower in the silymarin group (86.14 [4.97]; P = 0.04) and the PTX group (84.14 [11.21]; P = 0.04). MDA concentration was significantly lower in the silymarin group compared with the control group (0.08 [0.01] vs 0.22 [0.03] nmol/mL; P = 0.004); MDA was also significantly lower in the silymarin group than in the PTX group (0.11 [0.02]; P = 0.03).
Conclusions: Silymarin and PTX were associated with lower histopathologic liver damage, hepatocyte apoptosis, and regulation of extracellular matrix proteins. Lipid peroxidation in hepatocytes was significantly lower in the silymarin group compared with the PTX group. Silymarin and PTX appeared to have hepatoprotective effects in this experimental liver fibrosis model, but further clinical and experimental studies are needed.
Hepatoprotective and antifibrotic effect of a new silybin-phosphatidylcholine-Vitamin E complex in rats
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Silybin, the main active component of silymarin, has been reported to reduce hepatic fibrosis by 30% in bile duct ligated rats, whereas Vitamin E alone does not significantly modify liver damage and collagen deposition in chronic liver injury.
The aim of the present study was to evaluate the hepatoprotective and the antifibrotic properties of a new silybin–phosphatidylcholine–Vitamin E complex, characterised by elevated oral bioavailability and lipophilicity, on rat hepatic fibrosis induced by dimethylnitrosamine administration and by bile duct ligation.
The complex was administered by gastric gavage at a dose of 250 and 75 mg/kg (as silybin and Vitamin E, respectively). Treatment with the complex was able to prevent the dimethylnitrosamine-induced loss in body and liver weight, as well as to reduce the degree of liver injury, as determined by alanine aminotransferase values and necroinflammatory score. This was associated with reduced hepatic stellate cells proliferation both after 1 and 5 weeks of treatment. Treatment with the complex reduced also hepatic stellate cells activation and collagen deposition. Treatment with dimethylnitrosamine induced an increase in α1(I) procollagen, TGF_β1, tissue inhibitor of metalloproteinase 1 and metalloproteinase 2 mRNA expression, which were significantly reduced by administration of the complex. In the bile duct ligation model, the administration of the complex was able to reduce hepatic stellate cells proliferation and activation, as well as collagen deposition and α1(I) procollagen mRNA expression.
These results suggest that this new silybin–phosphatidylcholine–Vitamin E complex could be an interesting drug to be tested in patients with chronic liver disease.
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Liver fibrosis is an over-accumulation of extra-cellular matrix (ECM) and the hepatic stellate cell (Ito cell) play a central role in the pathogenesis of liver fibrosis. There are a lot of growth factors and cytokines involved in the activation of hepatic stellate cell, including of transforming growth factor (TGF-α, TGF-β1), platelet-derived growth factor (PDGF), interleukin (IL-1α,β, IL-6) and tumor necrosis factor (TNF-α).
Sho-saiko-to (TJ-9; Xiao-Chai-Hu-Tang in Chinese) was the most popular herbal medicine for the treatment of chronic liver disease in Chinese and Japanese. Our aim of the current study was to examine whether TJ-9 regulated the growth factors and cytokines in the fibrogenesis of bile duct ligated model. Therefore, we assessed the TJ-9's potential in regulating TGF-β1, PDGF mRNA expression, the amount of IL-1α, IL-1β, IL-6, TNF-α and the fibrotic marker “PIII NP” in the serum. Then, using the immunohistochemical stain to observe the TGF-β1 expression in the tissue.
Our results showed that TJ-9 at a dose of 0.5 g/(kg day) significantly reduced the serum level of PIII NP, the mRNA expression of TGF-β1 and PDGF. For the cytokines involved in the activation of Ito cell, TJ-9 at a dose of 0.5 g/(kg day) significantly suppressed the increasing tendency of IL-1β and enhanced the production of TNF-α. Finally, we concluded that: (1) TJ-9 at a dose of 0.5 g/(kg day) significantly reduced the serum fibrotic marker PIII NP in the bile duct ligated model, and its mechanism was partly by means of downregulating the mRNA of TGF-β1 and PDGF. These results also confirmed by the immunohistochemical staining of TGF-β1. (2) TJ-9 at a dose of 0.5 g/(kg day) suppressed the increasing tendency of IL-1β and stimulated the production of TNF-α to inhibit Ito cell proliferation and collagen formation.
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Citation Excerpt :
Considering genes that were elevated at later time points with high amplitude, we observed alterations in numerous genes indicative of fibrosis and tissue remodeling including procollagen types I and III. These genes are precursors to collagen type I and III proteins, which have been suggested as serum markers of fibrogenesis in patients with chronic liver disease and animal models of liver fibrosis [12,13]. In addition, we observed a time- and dose-dependent increase in mRNA levels of tissue inhibitor of metalloproteinase-1 (TIMP-1) relative to controls.
Molecular techniques, such as cDNA microarrays, are being used to aid in the elucidation of the mechanisms of toxicity of a variety of compounds. In this study, we evaluate the molecular effects of furan in the rat liver. Sprague–Dawley rats were exposed to 4 or 40 mg/kg furan for up to 14 days. Furan induced an initial degenerative and necrotic phenotype that was followed by inflammation and fibrosis, consistent with previous observations for this compound. RNA was harvested from each lobe of the liver at several time points to observe whether lobe-specific gene expression effects occurred. Similar gene expression changes were observed in all lobes, however the magnitude of gene expression change was more pronounced in the right lobe. Finally, to help determine the correlation between gene expression changes and liver pathology, we applied traditional microarray visualization tools to the assessment of clinical chemistry and pathology parameters.
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Modulation of collagen XVIII/endostatin expression in lobular and biliary rat liver fibrogenesis
2001, Journal of Hepatology
Background/Aims: The liver is the major source of collagen XVIII (C18), the precursor of the angiogenesis inhibitor endostatin. In human liver C18 is mainly expressed by hepatocytes. However, its quantitative and temporospatial expression patterns during liver fibrogenesis are unknown.
Methods: We used RNA quantification and in situ hybridization combined with cell-specific markers to study C18 compared to procollagen α1(I) and tissue inhibitor of metalloproteinases-1 (TIMP-1) mRNA expression in acute (single dose of CCl₄) and chronic (biliary) rat liver fibrogenesis.
Results: C18 transcripts were only found in hepatocytes and bile duct epithelia of normal and fibrotic livers, and occasionally in arterial myocytes and hepatic stellate cells. 72 h after CCl₄ injection, C18 mRNA levels remained unchanged, while procollagen α1(I) mRNA was increased at 72 h and TIMP-1 mRNA peaked at 12 h (P<0.05). In biliary fibrosis C18 mRNA levels increased 1.8-fold, contrasting with 20- and 4-fold elevated procollagen α1(I) and TIMP-1 transcript levels, respectively.
Conclusions: Hepatocytes and bile duct epithelia are the predominant sources of C18 in normal and fibrotic rat liver. Contrary to procollagen α1(I) and TIMP-1, C18 expression remains constant in acute fibrogenesis and is upregulated in biliary fibrosis. Modulation of epithelial C18 expression and its processing to endostatin could allow a liver-specific anti-cancer therapy.

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Elevated serum aminoterminal procollagen type-III-peptide parallels collagen accumulation in rats with secondary biliary fibrosis

Abstract

J Hepatol

Lancet

J Hepatol

J Hepatol

Gastroenterology

J Hepatol

J Hepatol

The integrated value of serum procollagen III peptide over time predicts hepatic hydroxyproline content and stainable collagen in a model of dietary cirrhosis in the rat

Hepatology

Serum type III procollagen peptide in alcoholic liver disease and idiopathic hemochromatosis: its relationship to hepatic fibrosis, activity of the disease and iron overload

Hepatology

The N terminal propeptide of collagen type III in serum reflects activity and degree of fibrosis in patients with chronic liver disease

Hepatology

Sampling variability in percutaneous liver biopsy

Arch Intern Med

Connective tissue polypeptides in serum as parameters to monitor antifibrotic treatment in hepatic fibrogenesis

J Hepatol

Serum assays for liver fibrosis

J Hepatol

Noninvasive methods for detection of organ fibrosis

Serum aminoterminal procollagen peptide and 7S domain of type IV collagen in patients with alcohol abuse

Liver