Fibronectin Protects from Excessive Liver Fibrosis by Modulating the Availability of and Responsiveness of Stellate Cells to Active TGF-β

Nina Kawelke,Anja von Au,Matthaeus Vasel,Carla Sens,Steven Dooley,Inaam A. Nakchbandi,Carol Feghali-Bostwick

doi:10.1371/journal.pone.0028181

Nina Kawelke, Anja von Au + Show 5 more

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https://doi.org/10.1371/journal.pone.0028181

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Abstract

Fibrotic tissue in the liver is mainly composed of collagen. Fibronectin, which is also present in fibrotic matrices, is required for collagen matrix assembly in vitro. It also modulates the amount of growth factors and their release from the matrix. We therefore examined the effects of the absence of fibronectin on the development of fibrosis in mice.Conditional deletion of fibronectin in the liver using the Mx promoter to drive cre expression resulted in increased collagen production and hence a more pronounced fibrosis in response to dimethylnitrosamine in mice. Exclusive deletion of fibronectin in hepatocytes or normalization of circulating fibronectin in Mx-cKO mice did not affect the development of fibrosis suggesting a role for fibronectin production by other liver cell types. The boosted fibrosis in fibronectin-deficient mice was associated with enhanced stellate cell activation and proliferation, elevated concentrations of active TGF-β, and increased TGF-β-mediated signaling.In vitro experiments revealed that collagen-type-I production by fibronectin-deficient hepatic stellate cells stimulated with TGF-β was more pronounced, and was associated with augmented Smad3-mediated signaling. Interfering with TGF-β signaling using SB431542 normalized collagen-type-I production in fibronectin-deficient hepatic stellate cells. Furthermore, precoating culture plates with fibronectin, but not collagen, or providing fibronectin fibrils unable to interact with RGD binding integrins via the RGD domain significantly diminished the amount of active TGF-β in fibronectin-deficient stellate cells and normalized collagen-type-I production in response to TGF-β stimulation. Thus, excessive stellate cell activation and production of collagen results from increased active TGF-β and TGF-β signaling in the absence of fibronectin.In conclusion, our data indicate that fibronectin controls the availability of active TGF-β in the injured liver, which impacts the severity of the resulting fibrosis. We therefore propose a novel role for locally produced fibronectin in protecting the liver from an excessive TGF-β-mediated response.

Highlights

The development of liver fibrosis is one of the early steps in the pathogenesis of advanced liver disease and liver failure
We have shown that two isoforms of fibronectin reflect the severity of liver fibrosis in patients with chronic hepatitis C raising the possibility that fibronectin itself may play a role in the pathogenesis of fibrosis [9]
Hepatic stellate cells were isolated from control mice (CT) and conditional knockout mice carrying the Mx promoter attached to cre recombinase (Mx-cKO)

Summary

Introduction

The development of liver fibrosis is one of the early steps in the pathogenesis of advanced liver disease and liver failure. Hepatic stellate cells play a key role in this process by producing extracellular matrix molecules that become incorporated in a distorted fibrogenous network hindering normal function in hepatocytes [1]. Fibronectin is one of the molecules produced by hepatic stellate cells [3]. It is part of the extracellular matrix [4]. Its continuous presence supports matrix integrity, both in vitro and in vivo [5,7]. It further regulates cell proliferation and cell cycle progression [8]. We have shown that two isoforms of fibronectin reflect the severity of liver fibrosis in patients with chronic hepatitis C raising the possibility that fibronectin itself may play a role in the pathogenesis of fibrosis [9]

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