CONTRIBUCIÓN ESPECIAL

LIVER FIBROSIS: PATHOGENESIS, PREVENTION AND TREATMENT

Detlef Schuppan Medizinische Klinik I, Universität Erlangen-Nürnberg, Germany
(6 pages, 4 figures, 3 tables)

THE DEVELOPMENT OF LIVER FIBROSIS

Fibrosis results from excessive accumulation of extracellular matrix (ECM). ECM describes the connective tissue molecules found in all multicellular organisms. These molecules are grouped into major molecular classes, mainly the collagens, noncollagenous glycoproteins, glycosaminoglycans, proteoglycans and elastin. In most organs, collagens, especially the fibril forming collagen types I and III, but also basement membrane collagen type IV, are the most abundant ECM components (1). In liver cirrhosis their relative tissue content may increase up to tenfold. This increase explains most of the complications of cirrhosis, such as an impaired exchange of metabolites between the sinusoidal blood and the hepatocytes via sinusoidal sclerosis (capillarization) and the formation of porto-venous shunts that prevent sinusoidal perfusion. The latter is also the basis for the increase in portal pressure that leads to esophageal or gastric varices and the development of ascites. Lastly, the continuous stimulus for hepatocyte proliferation in an abnormal ECM environment (regenerative nodules) predisposes for the development of hepatocellular carcinoma.

As demonstrated in Fig.1 a variety of adverse stimuli such as hepatotoxins, hepatotropic viruses, hypoxia, immune reactions to the liver, metabolic diseases, biliary stasis or simply mechanical stress can trigger liver fibrogenesis, i.e., the excess synthesis and deposition of ECM. In acute liver diseases, such as self-limited viral hepatitis, fibrogenesis is balanced by fibrolysis, i.e., the removal of excess ECM by proteolytic enzymes, the most important of which are the matrix metalloproteinases (MMPs). With repeated injury of sufficient severity, as occurs in many chronic liver diseases, fibrogenesis prevails, finally resulting in morphologically apparent fibrosis or cirrhosis. 

Fig 1.- Initiation and maintenance of fibrogenesis

Fibrogenesis is accompanied by an upregulation of collagen synthesis, a downregulation MMP secretion and activity, and by an increase of the physiological inhibitors of the MMPs, the tissue inhibitors of MMPs (TIMPs), of which the universal MMP-inhibitor TIMP-1 is the most important (1,2). Collagens, MMPs and TIMPs are mainly produced by activated hepatic stellate cells (HSC, synonymous with Ito cells) and by activated portal fibroblast (PF) (1-4).

Activated Kupffer cells or proliferating bile duct epithelia are major sources of potentially fibrogenic cytokines and growth factors that further stimulate HSC and PF to become activated myofibroblastic cells (1-4). Similar cell types are found in other organs prone to fibrosis such as the pancreas, kidney, lung, intestine, skin and arteries (1,5) (Table 1). Usually, activation to myofibroblastic cells is the key step of a protective program aimed at rapid closure of a potentially lethal wound (1,5). This program is self-limiting if the offending agent is present for a short period of time but leads to fibrosis and cirrhosis when continuously activated. It follows that the activated HSC and PF are an important target for an antifibrotic therapy in chronic liver diseases.

POTENTIAL ANTIFIBROTIC AGENTS

Since HSC and PF were identified as the the major fibrogenic cell types and since they undergo spontaneous activation in cell culture, the stage has been set for the development of specific antifibrotic agents. Such agents are currently identified and tested in numerous laboratories worldwide. Once active in the in vitro culture, all substances have to undergo ‚proof of principle' in a suitable animal model, predominantly hepatic fibrosis and cirrhosis of the rat. Models that evolve chronically and reproducibly, such as biliary cirrhosis due to bile duct occlusion, are preferable over those characterized by major hepatocyte necrosis, such as induced by carbon tetrachloride, dimethylnitrosamine or galactosamine, because the former more closely resemble human chronic liver disease and allow to identify a ‚ "true antifibrotic" instead of an anti-inflammatory, anti-necrotic or radical scavenging effect.

Table 2 lists substances which have been tested in suitable animal models (5). Part of these drugs is currently undergoing phase 2 or 3 clinical testing with pre- and post-treatment biopsy for exact morphometrical determination of the area of connective tissue and with a spectrum of surrogate markers of liver fibrogenesis (see below). Promising drugs are an oral endothelin A receptor (ETAR) antagonist (6), silymarin (7), interferon alpha (8,9), derivatives of pentoxifyllin (10) and antagonists to the fibrogenic cytokine transforming growth factor beta (TGF-b) or connective tissue growth factor (CTGF) (11,12). Antagonizing the endothelin A receptor offers the additional potential to lower portal hypertension which is in part mediated by endothelin 1 (ET-1) induced contraction of activated HSC (Fig.2). Reports on the antifibrotic acvtivity of hepatocyte growth factor (HGF) (13) have to be interpreted with care, since this cytokine rather causes hypertrophy and hyperplasia of hepatocytes, thus reducing the relative and not the absolute collagen content in the liver, with the additional danger of promoting hepatic malignancy. A further potental antifibrotic is rapamycin, an immunosuppressant used in organ transplantation (14).

ET-1/ET AR system and stellate cell activation

A promising target is the induction of stress relaxation of fibrogenic cells, a matrix (integrin) receptor-mediated process that is associated with a decrease in collagen synthesis and an increase in collagenase activity. This stress relaxation occurs once mesenchymal cells are placed from a ‚ "stressed", two-dimensional environment (mimicking a situation of wounding) into a ‚"relaxed", three-dimensional environment (5,15). Stress relaxation mitigates or even abrogates signals transferred via certain mitogenic growth factors and can revert the same cell that caused fibrogenesis into a fibrolytic cell that preferably releases MMPs instead of collagens. Thus, the receptors for platelet-derived growth factor and endothelin-1 (via the ETA receptor) transmit potent stress signals that trigger proliferation and ECM synthesis in activated HSC and PF. Interestingly, also soluble proteolytic fragments of collagen VI which are released from the liver matrix during remodeling serve as a potent growth stimulator and anti-apoptotic factor for fibrogenic cells, an effect that is mediated via a non-integrin collagen VI receptor (16-18). Apart from direct inhibition of these receptors by peptides or peptide analogues, coupling specific receptor recognizing cyclic peptides or other ligands to a drug carrier allows highly specific targeting of the activated fibrogenic cells in the liver (Fig.3). This has been shown both in vitro and in vivo with cyclic peptides recognizing the receptors for PDGF, collagen VI and mannose-6-phosphate (19-21). With these ligands specific uptake of the targeted carrier in activated HSC of fibrotic rat livers can reach up to 50% in vivo.

Receptor-targeted antifibrotic therapy

Circulating matrix proteins related to fibrogenesis and fibrolysis

SERUM MARKERS OF LIVER FIBROSIS

Table 3 shows some of the serum fibrosis markers that may be useful in future studies of antifibrotic drug effects in the liver. Most of these markers appear to reflect fibrogenesis rather than fibrolysis (3,5,22). They open the possibility to assess the future evolution of fibrosis and the effect of potential antifibrotic treatment in an individual patient on a frequent basis. However, these markers still await validation in large prospective follow-up studies of patients with liver diseases. Several such studies are currently underway. They involve more than 1000 patients with sequential liver biopsies 18-24 months apart. From these biopsies the increase of the connective tissue area and volume will be determined morphometrically. In addition, quantitative RT-PCR, to quantitate hepatic expression of several collagens, of MMPs and TIMP-1 will be performed from fractions of diagnostic biopsies, allowing a direct comparison with the serum fibrosis markers.

Bibliografía

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* Medizinische Klinik I, University Erlangen-Nürnberg, Germany

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