Tissue remodelling is dependent on the integration of signals that control turnover of ECM (extracellular matrix). Breakdown and endocytosis of collagen, a major component of the ECM, is central to this process. Whereas controlled secretion of matrix-degrading enzymes (such as matrix metalloproteinases) has long been known to mediate ECM breakdown, it is becoming clear that uPARAP/Endo180 (where uPARAP stands for urokinase plasminogen activator receptor-associated protein) serves as a receptor that mediates endocytosis of collagen by several types of cells. In the liver, the stellate cells play a major role in turnover of ECM including collagens. These cells synthesize various collagens and also produce matrix metalloproteinases. In the present study, we investigated the capacity of rat hepatic stellate cells to endocytose and degrade 125I-labelled heat-denatured collagen I. It was found that the collagen is efficiently taken up and degraded by these cells. Degradation was inhibited by inhibitors of lysosomal proteases (leupeptin and E-64d) and the vacuolar proton pump (concanamycin A), indicating that it takes place in lysosomes. Furthermore, endocytosed FITC-labelled collagen was shown to reach late endocytic compartments in which it colocalized with LysoTracker (a marker of late endocytic compartments). Competition experiments showed that uPA and unlabelled collagen are capable of inhibiting binding and uptake of [125I]collagen in a dose-dependent manner. Moreover, Western-blot analysis of cell lysate (using a polyclonal rabbit human-Endo180 antiserum) revealed a single band at 180 kDa. In addition, the antiserum was capable of reducing [125I]collagen binding to the cell surface. Finally, using two primers designed from the human uPARAP/Endo180 mRNA sequence, the expression of uPARAP/Endo180 mRNA was detected by reverse transcriptase–PCR. These results together suggest that uPARAP/Endo180 mediates endocytosis of collagen in rat liver stellate cells.
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Research Article|
March 22 2005
Uptake of denatured collagen into hepatic stellate cells: evidence for the involvement of urokinase plasminogen activator receptor-associated protein/Endo180
Seyed Ali MOUSAVI;
Seyed Ali MOUSAVI
*Department of Molecular Biosciences, University of Oslo, Blindernveien 31, 0316 Oslo, Norway
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Mitsuru SATO;
Mitsuru SATO
†Department of Anatomy, Akita University School of Medicine, 1-1-1 Hondo, Akita 010-8543, Japan
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Marita SPORSTØL;
Marita SPORSTØL
*Department of Molecular Biosciences, University of Oslo, Blindernveien 31, 0316 Oslo, Norway
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Baard SMEDSRØD;
Baard SMEDSRØD
‡Department of Experimental Pathology, University of Tromsø, Norway
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Trond BERG;
Trond BERG
1
*Department of Molecular Biosciences, University of Oslo, Blindernveien 31, 0316 Oslo, Norway
1To whom correspondence should be addressed (email trond.berg@bio.uio.no).
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Naosuke KOJIMA;
Naosuke KOJIMA
†Department of Anatomy, Akita University School of Medicine, 1-1-1 Hondo, Akita 010-8543, Japan
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Haruki SENOO
Haruki SENOO
†Department of Anatomy, Akita University School of Medicine, 1-1-1 Hondo, Akita 010-8543, Japan
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Publisher: Portland Press Ltd
Received:
June 08 2004
Revision Received:
October 14 2004
Accepted:
October 27 2004
Accepted Manuscript online:
October 27 2004
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2005
Biochem J (2005) 387 (1): 39–46.
Article history
Received:
June 08 2004
Revision Received:
October 14 2004
Accepted:
October 27 2004
Accepted Manuscript online:
October 27 2004
Citation
Seyed Ali MOUSAVI, Mitsuru SATO, Marita SPORSTØL, Baard SMEDSRØD, Trond BERG, Naosuke KOJIMA, Haruki SENOO; Uptake of denatured collagen into hepatic stellate cells: evidence for the involvement of urokinase plasminogen activator receptor-associated protein/Endo180. Biochem J 1 April 2005; 387 (1): 39–46. doi: https://doi.org/10.1042/BJ20040966
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