Class II HDAC Inhibition Hampers Hepatic Stellate Cell Activation by Induction of MicroRNA-29

et al. (2013) Class II HDAC Inhibition Hampers Hepatic Stellate Cell Activation by Induction of MicroRNA-29. PLoS ONE 8(1): e55786. doi:10.1371/journal.pone.0055786 Class II HDAC Inhibition Hampers Hepatic Stellate Cell Activation by Induction of MicroRNA-29 Inge Mannaerts 0 Nathalie Eysackers 0 Oscar O. Onyema 0 Katrien Van Beneden 0 Sergio Valente 0 Antonello Mai 0 Margarete Odenthal 0 Leo A. van Grunsven 0 Rajasingh Johnson, University of Kansas Medical Center, United States of America 0 1 Department of Cell Biology, Liver Cell Biology Lab, Vrije Universiteit Brussel, Brussels, Belgium, 2 Department of Human Anatomy, Liver Cell Biology Lab, Vrije Universiteit Brussel, Brussels, Belgium, 3 Istituto Pasteur - Fondazione Cenci Bolognetti, Dipartimento di Chimica e Tecnologie del Farmaco, Sapienza Universita' di Roma , Roma , Italy , 4 Institute of Pathology, University Hospital of Cologne , Cologne , Germany Background: The conversion of a quiescent vitamin A storing hepatic stellate cell (HSC) to a matrix producing, contractile myofibroblast-like activated HSC is a key event in the onset of liver disease following injury of any aetiology. Previous studies have shown that class I histone deacetylases (HDACs) are involved in the phenotypical changes occurring during stellate cell activation in liver and pancreas. Aims: In the current study we investigate the role of class II HDACs during HSC activation. Methods: We characterized the expression of the class II HDACs freshly isolated mouse HSCs. We inhibited HDAC activity by selective pharmacological inhibition with MC1568, and by repressing class II HDAC gene expression using specific siRNAs. Results: Inhibition of HDAC activity leads to a strong reduction of HSC activation markers a-SMA, lysyl oxidase and collagens as well as an inhibition of cell proliferation. Knock down experiments showed that HDAC4 contributes to HSC activation by regulating lysyl oxidase expression. In addition, we observed a strong up regulation of miR-29, a well-known anti-fibrotic miR, upon treatment with MC1568. Our in vivo work suggests that a successful inhibition of class II HDACs could be promising for development of future anti-fibrotic compounds. Conclusions: In conclusion, the use of MC1568 has enabled us to identify a role for class II HDACs regulating miR-29 during HSC activation. - Funding: I. Mannaerts and L. A. van Grunsven are supported by the Vrije Universiteit Brussel (GOA48, GOA78, OZR1930) and by the Fund for Scientific Research Flanders (FWO-V, http://www.fwo.be/) (G.0260.09). K. Van Beneden is supported by the Vrije Universiteit Brussel through an Onderzoeksraad project (OZR1428 and OZR1796). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. Fibrosis is characterized by excessive scar formation due to overproduction and deposition of extracellular matrix (ECM). This process usually occurs over a long period of time and can lead to organ dysfunction or death. There is no effective therapy available at the moment; therefore organ transplantation is often the only redress for patients with fibrosis. Donor shortage however underlines the need for more research on alternative therapies [1]. The identification of the hepatic stellate cells (HSCs) as the key cellular source of ECM synthesis in the liver was an important step towards the understanding of the mechanism of liver fibrosis and the development of new therapeutic strategies [2,3]. Like liver sinusoidal endothelial cells and Kupffer cells, quiescent HSCs are non-parenchymal cells. They reside in the space of Disse and are lipid droplet containing cells that play a major role in the control and metabolism of retinol in the organism [4]. Following acute or chronic liver damage, these cells undergo a process of activation towards a myofibroblastic phenotype. This activation process is the result of a series of changes in gene expression [5]. The gene expression changes lead to a loss of retinoid containing lipid droplets, increased proliferation, motility, increased a-smooth muscle actin (a-SMA) expression, contractility and synthesis of extracellular components and matrix remodeling enzymes. This activation process is the dominant factor in liver fibrogenesis [2,3]. As a consequence, inhibition of HSC activation can be an important target to develop new therapeutic strategies to intervene in liver fibrosis and cirrhosis [6,7]. Alterations in the gene expression profile of HSCs during myofibroblastic activation are associated with changes in microRNA expression [8,9]. microRNAs are small RNA molecules, that are able to inhibit protein synthesis by interacting with the 39untranslated region of mRNA derived from certain genes [10]. During HSC activation the expression of antifibrogenic microRNAs such as miR-29 is decreased [11,12], whereas others like (...truncated)