Selenium supplementation attenuates procollagen-1 and interleukin-8 production in fat-loaded human C3A hepatoblastoma cells treated with TGFβ1 Catriona Clarke a , Hussam Baghdadi b , Alexander F. Howie b , J. Ian Mason b , Simon W. Walker c , Geoffrey J. Beckett c, ⁎ a Clinical Biochemistry, Western General Hospital, Edinburgh, UK b Clinical Biochemistry Section, University of Edinburgh, The Queen's Medical Research Institute, Edinburgh, UK c Clinical Biochemistry, University of Edinburgh, The Royal Infirmary of Edinburgh, Edinburgh, UK abstract article info Article history: Received 9 December 2009 Received in revised form 10 February 2010 Accepted 17 February 2010 Available online 23 February 2010 Keywords: Selenium Non-alcoholic steatohepatitis Cytokine Epithelial mesenchymal transition Fatty acid Transforming growth factor β1 Background: Non-alcoholic fatty liver disease (NAFLD) is associated with obesity, insulin resistance and hepatic steatosis. Non-alcoholic steatohepatitis (NASH) is a serious consequence of NAFLD where chronic tissue damage and inflammation result in fibrosis which may progress to cirrhosis. Transforming growth factor β1 (TGFβ1), proinflammatory cytokines and oxidative stress are thought to play crucial roles in the pathogenesis of these conditions. The contributions of individual liver cell types to fibrogenesis remain controversial and the influence of selenium status has not been investigated. Methods: In this study we have used a cell culture model of fat-loading using oleate-treated human hepatoblastoma (C3A) cells to investigate how fat-loading and selenium status might influence the production of collagen in response to TGFβ1. The secretion of inflammatory cytokines was also investigated, together with the epithelial character of the treated cells. Results: We found that in response to treatment with TGFβ1, C3A cells produced mRNA encoding the pro-αI chain of procollagen I, secreted procollagen I peptide, up-regulated production of the proinflammatory cytokine interleukin-8 (IL-8) and the mesenchymal marker vimentin, and down-regulated albumin production. Most of these responses were considerably enhanced when cells were fat-loaded with oleate and were attenuated by selenium addition at a dose that optimised the expression of thioredoxin reductase and glutathione peroxidase. Conclusions: Our data establish that both fat-loading and suboptimal selenium status enhance collagen and IL-8 production by C3A hepatocytes in response to TGFβ1, possibly as part of an epithelial to mesenchymal transition. General significance: These findings suggest that the hepatocyte may be an important contributor to the pathogenesis of fibrosis associated with NAFLD. © 2010 Elsevier B.V. All rights reserved. 1. Introduction Non-alcoholic steatohepatitis (NASH) is a clinically significant consequence of non-alcoholic fatty liver disease (NAFLD), seen in a subset of patients and characterised by hepatic steatosis, chronic inflammation, extensive fibrosis with hepatocyte damage and impaired function [1]. NAFLD, associated with obesity, insulin resistance and dyslipidaemia, is defined as excessive accumulation of fat in the liver (steatosis) to 5–10% (by weight) [2]. The conventional model of hepatic fibrosis focuses on hepatic stellate cells (HSC), senescent vitamin A-storing cells that, in response to hepatocyte injury and apoptosis, are activated to myofibroblasts. Activated stellate cells modify the microenvironment of the liver, restructuring the extracellular matrix (ECM) and recruiting circulat- ing immune cells. They proliferate and, with the increased volume of ECM, are thought to fill the space left by dying hepatocytes [3], whilst progenitor stem cells proliferate and differentiate into new hepato- cytes [4]. A critical cytokine in this process is TGFβ1 [5], released predominantly by macrophages and thought to orchestrate healing, including the production of collagen by stellate cells. There is a growing body of evidence to indicate that hepatocytes themselves respond to TGFβ1 by secreting collagen [6]. This has been shown in primary mouse hepatocytes [7,8], a mouse hepatocyte cell line [7] and in mouse models [9], and occurs at the level of both the mRNA [7,9] and protein [7,8]. It has been proposed that, under the influence of Biochimica et Biophysica Acta 1800 (2010) 611–618 ⁎ Corresponding author. Clinical Biochemistry, University of Edinburgh, The Royal Infirmary of Edinburgh, 51 Little France Crescent, Little France, Edinburgh, EH16 4SA UK. Tel.: +44 131 242 6868; fax: +44 131 242 6812. E-mail addresses: catriona.clarke@nhs.net (C. Clarke), S0460067@sms.ed.ac.uk (H. Baghdadi), F.Howie@ed.ac.uk (A.F. Howie), J.I.Mason@ed.ac.uk (J.I. Mason), Simon.Walker@ed.ac.uk (S.W. Walker), G.J.Beckett@ed.ac.uk (G.J. Beckett). 0304-4165/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.bbagen.2010.02.007 Contents lists available at ScienceDirect Biochimica et Biophysica Acta journal homepage: www.elsevier.com/locate/bbagen