Liver, Pancreas and Biliary Tract
Liver expression of matrix metalloproteases and their inhibitors in hepatocellular carcinoma

https://doi.org/10.1016/j.dld.2009.01.016Get rights and content

Abstract

Background

Several studies have suggested the significance of some metalloproteases in the malignant behaviour of hepatocellular carcinoma.

Aims

To evaluate the liver expression of MMPs and their tissular inhibitors in patients with HCC.

Methods

An immunohistochemical study using tissue microarrays on samples obtained from 30 HCC patients, with antibodies against MMPs (1, 2, 7, 9, 11, 13 and 14) and TIMPs (1, 2 and 3) was performed. Results were correlated with various clinico-pathological findings and with overall survival.

Results

MMP-1 is mainly expressed by stromal cells, and MMP-13, TIMP-1 and TIMP-2 by inflammatory cells. A positive correlation between MMP-1 expression and larger size tumours (p < 0.01) was found. Increased TIMP-2 expression was associated with higher preoperative serum levels of α-fetoprotein (p < 0.01). Unsupervised hierarchical clustering for total score values designated two groups, one of them characterised by high MMPs and TIMPs expressions, including 21 cases (70%) for tumour cell clustering, 5 cases for fibroblasts (16.6%) and 6 cases for inflammatory cells (20%). All patients showing elevated MMPs and TIMPs expression in stromal cells presented a poor prognosis (p < 0.05).

Conclusions

High liver MMPs and TIMPs expressions in peritumour stromal cells are related to a poorer prognosis in HCC.

Introduction

The majority of hepatocellular carcinoma (HCC) patients show at the time of diagnosis advanced or unresectable disease. The prognosis is poor even for those patients who undergo resection. This is due to the recurrence rates, which are among the highest of any solid tumour, reaching 50% 2 years after resection [1], [2], [3]. In addition, HCCs are inherently chemotherapy-resistant tumours [4]. Metastatic recurrence is the main obstacle towards the improvement of treatment efficacy. Proteolytic enzymes of various classes participate in the degradation of the stromal connective tissue and basement membrane components, therefore facilitating tumour invasion and metastasis. However, some matrix components, particularly the interstitial collagens, are very resistant to proteolytic attacks, being degraded only by matrix metalloproteases (MMPs) [5].

The human MMP family currently consists of 28 members of homologous zinc-dependent endopeptidases, which can be divided into eight structural classes or, based on their substrate specificity and primary structure into the more familiar subgroups of collagenases (MMP-1, -8 and -13), gelatinases (MMP-2 and -9), stromelysins (MMP-3, -10 and -11), membrane-associated MMPs (MMP-14, -15, -16, -17, -23, -24 and -25) and other novel MMPs [6], [7], [8], [9]. MMPs are synthesized as inactive zymogens, being then activated predominantly pericellularly by other MMPs or by serine proteases. MMPs’ activity is specifically inhibited by the so-called tissue inhibitors of metalloproteases (TIMPs). Currently, four different TIMPs are known to exist: TIMPs-1, -2, -3 and -4. In addition, this enzymatic system has two additional aspects of relevance in cancer biology: MMPs are able to impact on tumour cell behaviour in vivo as a consequence of their ability to cleave growth factors, cell surface receptors, cell adhesion molecules, and chemokines/cytokines [10], [11], [12], [13], [14], [15]. MMPs, by cleaving proapoptotic factors produce a more aggressive phenotype via generation of apoptotic resistant cells [16]. MMPs also regulate angiogenesis in cancer, positively through their ability to mobilize or activate proangiogenic factors [17], [18], and negatively via generation of angiogenesis inhibitors, such as angiostatin and endostatin, cleaved from large protein precursors [19], [20], [21]. On the other hand, it is now assumed that TIMPs are multifactorial proteins also involved in the induction of proliferation and in the inhibition of apoptosis [22], [23].

Several studies have suggested the significance of some MMPs in the malignant behaviour of HCC, such as MMP-2 [24], [25], [26], [27], [28], [29], [30], [31], MMP-7 [26], [32], MMP-9 [29], [31], [33], [34], [35], and MMP-14 (MT1-MMP) [29], [36], [37], [38]. These MMPs were associated with several indicative parameters of tumour aggressiveness and poor prognosis. However, the cellular type expressing each factor (tumour cell and/or peritumour stromal cell) was not specifically considered in HCCs. Likewise, there are few available data referring to the integrated expression of these factors in HCC.

The objectives of the present work are: (i) to evaluate the expression of several MMPs and TIMPs in HCCs by using the tissue microarray (TMA) technique, which allows us to integrate the biological aspects of this complex enzymatic expression in the morphological context of HCCs; (ii) to investigate their relationship with clinico-pathological and biological characteristics of the tumours; and (iii) to determine the correlation between the expressions of these proteins and overall survival.

Section snippets

Patient selection

The study population included 30 patients diagnosed with HCC at the University Hospital Central of Asturias in Oviedo, Spain, during a 13-year period (1989–2002) and before a program of liver transplantation was developed. The patients included in the study have been followed up at the same hospital since then.

All the cases were diagnosed on a clinical basis with imaging and pathological confirmation of the tumour. Patients showed limited nodular disease and associated cirrhosis, stage A in 24

Results

More than 600 determinations of cancer specimens from 30 patients with HCC and controls were performed on TMAs. Minimal internal variance of score data between duplicate tissue cores from the same patients was detected in the tissue arrays, showing a high agreement for each protein (r > 0.95 and p < 0.0001, for each protein). In the validation study there was a total concordance in the global expression, as well as in the intensity of immunostaining, for each MMP and TIMP between TMA cases and the

Discussion

In the present study we have described new findings on the expressions of MMPs and TIMPs in HCC, together with an important expression by tumour stromal cells, as well as their clinical relevance. It is known that the expression of MMPs in neoplastic tissues is high due to a paracrine regulation by growth factors and cytokines secreted by either tumour or stromal cells [44]. Nevertheless, the high expressions of MMPs and TIMPs by HCCs tissues may be also because of the interplay between

Conflict of interest statement

None declared.

References (59)

  • Z.H. Gao et al.

    Association of E-cadherin, matrix metalloproteinases, and tissue inhibitors of metalloproteinases with the progression and metastasis of hepatocellular carcinoma

    Mod Pathol

    (2006)
  • S. Arii et al.

    Overexpression of matrix metalloproteinase 9 gene in hepatocellular carcinoma with invasive potential

    Hepatology

    (1996)
  • A. Hayasaka et al.

    Elevated plasma levels of matrix metalloproteinase-9 (92-kD type IV collagenase/gelatinase B) in hepatocellular carcinoma

    Hepatology

    (1996)
  • T. Harada et al.

    Membrane-type matrix metalloproteinase-1 (MT1-MTP) gene is overexpressed in highly invasive hepatocellular carcinomas

    J Hepatol

    (1998)
  • I. Stamenkovic

    Matrix metalloproteinases in tumor invasion and metastasis

    Semin Cancer Biol

    (2000)
  • R.C. Benyon et al.

    Expression of tissue inhibitor of metalloproteinases 1 and 2 is increased in fibrotic human liver

    Gastroenterology

    (1996)
  • T. Takahara et al.

    Dual expression of matrix metalloproteinase-2 and membrane-type 1-matrix metalloproteinase in fibrotic human livers

    Hepatology

    (1997)
  • Y. Kang et al.

    A multigenic program mediating breast cancer metastasis to bone

    Cancer Cell

    (2003)
  • J.M. Freije et al.

    Molecular cloning and expression of collagenase-3, a novel human matrix metalloproteinase produced by breast carcinomas

    J Biol Chem

    (1994)
  • V. Knauper et al.

    The role of the C-terminal domain of human collagenase-3 (MMP-13) in the activation of procollagenase-3, substrate specificity, and tissue inhibitor of metalloproteinase interaction

    J Biol Chem

    (1997)
  • R. Ala-aho et al.

    Collagenases in cancer

    Biochimie

    (2005)
  • H. Nakatsukasa et al.

    Cellular distribution of transcripts for tissue inhibitor of metalloproteinases 1 and 2 in human hepatocellular carcinomas

    Hepatology

    (1996)
  • J. Yamamoto et al.

    Recurrence of hepatocellular carcinoma after surgery

    Br J Surg

    (1996)
  • R.T. Poon et al.

    Intrahepatic recurrence after curative resection of hepatocellular carcinoma: long-term results of treatment and prognostic factors

    Ann Surg

    (1999)
  • M. Huang et al.

    The study of innate drug resistance of human hepatocellular carcinoma Bel7402 cell line

    Cancer Lett

    (1999)
  • A.R. Nelson et al.

    Matrix metalloproteinases: biologic activity and clinical implications

    J Clin Oncol

    (2000)
  • C.M. Overall et al.

    Strategies for MMP inhibition in cancer: innovations for the post-trial era

    Nat Rev Cancer

    (2002)
  • M. Demers et al.

    New roles for matrix metalloproteinases in metastasis

    Crit Rev Immunol

    (2005)
  • C.E. Brinckerhoff et al.

    Interstitial collagenases as markers of tumor progression

    Clin Cancer Res

    (2000)
  • Cited by (37)

    • Molecularly targeted anti-cancer drugs inhibit the invasion and metastasis of hepatocellular carcinoma by regulating the expression of MMP and TIMP gene families

      2018, Biochemical and Biophysical Research Communications
      Citation Excerpt :

      Our establishment of the anti-invasion and anti-metastatic properties of lenvatinib provides a reasonable basis for the clinical application of lenvatinib in the treatment of liver cancer. MMP and TIMP gene families are the most important enzymes in tumor invasion and metastasis and are related with a poor prognosis in some human cancers including liver cancer [24–26]. TIMPs, as the MMPs inhibitors, have distinct tumor-stimulatory functions [27].

    • Matrix Metalloproteinase 1 as a Novel Biomarker for Monitoring Hepatocellular Carcinoma in Liver Transplant Patients

      2018, Transplantation Proceedings
      Citation Excerpt :

      In 2001, Kim et al detected the presence of MMP-1 mRNA, together with that of other MMPs, in several liver tumor cell lines [18]. In 2009, Altadill et al observed that MMP-1 overexpression by fibroblasts correlated with poor prognosis and higher metastatic activity of HCC tumor cells [19]. A later study showed that MMP-1 expression was elevated in HCC liver tissues, but decreased or absent in normal tissues [20].

    • Hinokitiol, a tropolone derivative, inhibits mouse melanoma (B16-F10) cell migration and in vivo tumor formation

      2015, European Journal of Pharmacology
      Citation Excerpt :

      Among the 28 identified members of the human MMP family, MMP-1 is widely expressed by mesenchymal and epithelial cells; it degrades interstitial collagens, the most abundant ECM proteins in the body (Vincenti and Brinckerhoff, 2007). Increased expression of MMP-1 has been observed in the well-differentiated cancer cells of hepatocellular carcinoma, and is correlated with a poor prognosis, tumor invasion, and metastasis (Altadill et al., 2009). In addition to MMPs, members of the mitogen- activated protein kinase (MAPK) superfamily are associated with increased scattering/motility, invasion, proliferation, survival, and morphogenesis.

    • Ginsenoside Rh1 suppresses matrix metalloproteinase-1 expression through inhibition of activator protein-1 and mitogen-activated protein kinase signaling pathway in human hepatocellular carcinoma cells

      2012, European Journal of Pharmacology
      Citation Excerpt :

      Among the 28 identified members of the human MMP family, MMP-1 is widely expressed by mesenchymal and epithelial cells and degrades interstitial collagens, the most abundant ECM proteins in the body (Vincenti and Brinckerhoff, 2007). Expression of MMP-1 has been observed in the well-differentiated cancer cells of hepatocellular carcinoma, and is correlated with poor prognosis, tumor invasion, and metastasis (Altadill et al., 2009; Gao et al., 2006). As hepatocellular carcinoma is usually associated with liver fibrosis/cirrhosis, where type 1 collagen is mainly deposited, it is likely that early hepatocellular carcinoma cells invade surrounding fibrous tissue by secreting MMP-1 (Okuda, 1997; Takano et al., 1995).

    View all citing articles on Scopus
    View full text