Endothelin stimulates transforming growth factor-β1 and collagen synthesis in stellate cells from control but not cirrhotic rat liver

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Abstract

Interactions between hepatic stellate cells and endothelin-1 are implicated in liver fibrosis. We determined endothelin-1, its receptors and its effects on the synthesis of a fibrogenic agent transforming growth factor (TGF)-β1 and collagen in stellate cells from control and CCl4-induced cirrhotic rats. The basal synthesis of endothelin-1, TGF-β1 and collagen was much higher in cirrhotic stellate cells than in control cells. Endothelin-1 stimulated TGF-β1 and collagen synthesis via endothelin ETA and endothelin ETB receptors, respectively, in control stellate cells, but did not elicit these effects in the cirrhotic cells despite increased density of the respective receptor subtypes in them. These results indicate that the actions of endothelin-1 on stellate cells may be an important physiological mechanism in maintenance of hepatic architecture. However, inability of endothelin-1 to stimulate TGF-β1 and collagen synthesis in cirrhotic stellate cells suggests that it does not influence fibrogenic activity by direct action on them probably because the processes are already maximally activated.

Introduction

Physiologically, perisinusoidal stellate cells maintain the architecture of the liver by synthesizing components of extracellular matrix and the sinusoidal blood flow by contractile activity. During chronic liver injury, stellate cells lose their storage of retinoids, express α-smooth muscle actin, and transform into proliferating myofibroblast-like cells that are highly contractile and excessively fibrogenic. Thus, stellate cells play an important role in fibrogenesis during the development of liver cirrhosis Blomhoff and Wake, 1991, Geerts et al., 1994, Nakata and Shibayama, 1987.

Hepatic concentrations of endothelin-1, a powerful constrictor of the hepatic vasculature Gandhi et al., 1990, Tran-Thi et al., 1993, and its receptors increase progressively during the development of experimental cirrhosis (Gandhi et al., 1998). Furthermore, hepatic concentrations of endothelin-1 and its receptors increase in human cirrhosis Gandhi et al., 1996a, Pinzani et al., 1996 and there is a direct relationship between endothelin-1 receptor gene expression and portal pressure in cirrhotic patients (Leivas et al., 1998). Endothelin-1 causes contraction of quiescent, (Zhang et al., 1995) as well as transformed stellate cells Housset et al., 1993, Pinzani et al., 1996. Antagonism of endothelin-1 receptors has been reported to reduce the concentration of type I collagen and its mRNA in rat models of liver fibrosis (Rockey and Chung, 1996). However, whether endothelin-1 exerts this effect by direct actions on stellate cells is not known. Therefore, we investigated endothelin-1, its receptors and its effects on the fibrogenic activity of stellate cells isolated from the livers of carbon tetrachloride (CCl4)-induced cirrhotic rats.

Section snippets

Materials and methods

The following chemicals and reagents were purchased from the indicated sources: protease type XIV, 5-[N-2,3-dihydroxypropylacetamido-2,4,6-troiodo-N,N′-bis(2,3-dihydroxypropyl)isophthalamide] (Nycodenz), CCl4 (99.9% purity) and BQ-788 (N-cis-2,6-dimethylpiperidinecarbonyl-l-γMeLeu-d-Trp(COOMe)-d-Nle-ONa) (Sigma, St. Louis, MO); collagenase type IV (Worthington Biochemical, Freehold, NJ); transforming growth factor β1 (TGF-β1) (Life Technologies, Grand Island, NY); endothelin-1, sarafotoxin S6c

Results

We use phenobarbital to enhance the toxicity of CCl4 and establishment of uniform cirrhosis Gandhi et al., 1996b, Gandhi et al., 1998. However, considering the effects of phenobarbital alone on the liver tissue, we compared various parameters described in this paper between the cells isolated from untreated rats (these cells are routinely used in our laboratory) and rats treated with phenobarbital. No difference was observed in the morphological and immunohistochemical characteristics, as well

Discussion

The present investigation was performed considering the evidence that interactions between endothelin-1 and stellate cells can be of significant importance in physiology and pathology of the liver. Although all of the hepatic cell types synthesize endothelin-1 Gandhi et al., 1996b, Housset et al., 1993, Kuddus et al., 2000, Laura et al., 1998, Pinzani et al., 1996 and react to its agonistic activities in cell culture Caligiuri et al., 1998, Gandhi et al., 1992a, Gandhi et al., 1992b, Housset et

Acknowledgements

This work was supported by VA merit award and NIH grant DK 54411.

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