Fibronectin Stimulates Endothelin-1 Synthesis in Rat Hepatic Myofibroblasts via a Src/ERK-Regulated Signaling Pathway

Abstract

Liver injury serves as an excellent model of wound healing, characterized by increased synthesis of various cytokines and peptides, including the vasoactive peptide endothelin-1. In the liver, wound healing is mediated by effector cells such as hepatic stellate cells, which cause tissue contraction. Endothelin-1 has autocrine effects on stellate cells and induces their contractile activities. We explored the role of various extracellular matrix components, particularly of fibronectin, in regulating endothelin-1 production during liver injury. Hepatic stellate cells were isolated from normal and injured rats. Real-time polymerase chain reaction immunoblot and enzyme-linked immunosorbent assay analyses were used to measure specific variables, including endothelin-1 production. Preproendothelin-1 promoter activity was determined by a luciferase assay. Stellate cell contraction was measured by a gel contraction assay. Fibronectin stimulated transcription of preproendothelin-1 messenger RNA and expression of endothelin through an integrin-dependent pathway in activated hepatic stellate cells. In these cells, fibronectin induced phosphorylation/activation of extracellular signal-regulated kinase (ERK) through a Shc- and Src-dependent mechanism; ERK activation was required for fibronectin-induced endothelin-1 expression. Fibronectin stimulation by stellate cells induced expression of smooth muscle alpha-actin and endothelin-1-mediated autocrine stellate cell contraction. Stellate cells isolated from injured livers of rats exhibited increased basal phosphorylation levels of Src, Shc, and ERK, as well as increased endothelin-1 synthesis. Fibronectin stimulates activated stellate cells to produce endothelin-1 and contract, via an ERK-dependent signaling pathway. The resulting autocrine functional effects of endothelin-1 are likely to be important in the wound-healing process in injured liver.