Nrf2 protects stellate cells from Smad-dependent cell activation

• Published in: PloS one 2018-07, Vol.13 (7), p.e0201044-e0201044
• Main Authors: Prestigiacomo, Vincenzo, Suter-Dick, Laura
• Format: Article
• Language: English
• Subjects: Antioxidants; Biology and life sciences; Cell activation; Cell culture; Cell growth; Cell Movement - drug effects; Cell Movement - genetics; Cell Proliferation - drug effects; Cell Proliferation - genetics; Collagen; Extracellular matrix; Fibronectin; Fibronectins; Fibrosis; Gene expression; Gene Knockdown Techniques; Genes; Growth factors; Hepatic Stellate Cells - cytology; Hepatic Stellate Cells - drug effects; Hepatic Stellate Cells - metabolism; Humans; Hypoxia; Inhibition; Inhibitors; Kinases; Life sciences; Liver; Liver diseases; Markers; Medicine and Health Sciences; NF-E2-Related Factor 2 - deficiency; NF-E2-Related Factor 2 - genetics; NF-E2-Related Factor 2 - metabolism; Oxidative stress; Penicillin; Phosphatase; Platelet-derived growth factor; Platelet-Derived Growth Factor - pharmacology; Proteins; Research and Analysis Methods; Rodents; siRNA; Smad protein; Smad Proteins - metabolism; Stellate cells; Transcription factors; Transforming growth factor-b1; Transforming growth factors
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0201044

Hepatic stellate cells (HSC) orchestrate the deposition of extracellular matrix (ECM) and are the primary effector of liver fibrosis. Several factors, including TGF-β1, PDGF and oxidative stress, have been shown to trigger HSC activation. However, the involvement of cellular defence mechanisms, such as the activation of antioxidant response by Nrf2/Keap1 in the modulation of HSC activation is not known. The aim of this work was to elucidate the role of Nrf2 pathway in HSC trans-differentiation involved in the development of fibrosis. To this end, we repressed Nrf2 and Keap1 expression in HSC with specific siRNAs. We then assessed activation markers, as well as proliferation and migration, in both primary and immortalised human HSCs exposed to Smad inhibitors (SB-431542 hydrate and SB-525334), TGF-β1 and/or PDGF. Our results indicate that knocking down Nrf2 induces HSC activation, as shown by an increase in αSMA-positive cells and by gene expression induction of ECM components (collagens and fibronectin). HSC with reduced Nrf2-levels also showed an increase in migration and a decrease in proliferation. We could also demonstrate that the activation of Nrf2-deficient HSC involves the TGF-β1/Smad pathway, as the activation was successfully inhibited with the two tested Smad inhibitors. Moreover, TGF-β1 elicited a stronger induction of HSC activation markers in Nrf2 deficient cells than in wild type cells. Thus, our data suggest that Nrf2 limits HSCs activation, through the inhibition of the TGF-β1/Smad pathway in HSCs.