Hepatic ACAT2 Knock Down Increases ABCA1 and Modifies HDL Metabolism in Mice Matteo Pedrelli 1,2. , Padideh Davoodpour 1.¤a , Chiara Degirolamo 3¤b , Monica Gomaraschi 4 , Mark Graham 5 , Alice Ossoli 4 , Lilian Larsson 1 , Laura Calabresi 4 , Jan-A ˚ ke Gustafsson 2,6 , Knut R. Steffensen 2 , Mats Eriksson 2,7 , Paolo Parini 1,2 * 1 Division of Clinical Chemistry, Department of Laboratory Medicine, Karolinska Institutet, Stockholm, Sweden, 2 Molecular Nutrition Unit, Department of Bioscience and Nutrition, Karolinska Institutet, Stockholm, Sweden, 3 Division of Lipid Science, Department of Pathology, Wake Forest University School of Medicine, Winston-Salem, North Carolina, United States of America, 4 Department of Pharmacological Sciences, University of Milan, Milan, Italy, 5 Cardiovascular Group, Department of Antisense Drug Discovery, Isis Pharmaceuticals, Inc., Carlsbad, California, United States of America, 6 Center for Nuclear Receptors and Cell Signaling, University of Houston, Houston, Texas, United States of America, 7 Department of Medicine, Karolinska Institute, Stockholm, Sweden Abstract Objectives: ACAT2 is the exclusive cholesterol-esterifying enzyme in hepatocytes and enterocytes. Hepatic ABCA1 transfers unesterified cholesterol (UC) to apoAI, thus generating HDL. By changing the hepatic UC pool available for ABCA1, ACAT2 may affect HDL metabolism. The aim of this study was to reveal whether hepatic ACAT2 influences HDL metabolism. Design: WT and LXRa/b double knockout (DOKO) mice were fed a western-type diet for 8 weeks. Animals were i.p. injected with an antisense oligonucleotide targeted to hepatic ACAT2 (ASO6), or with an ASO control. Injections started 4 weeks after, or concomitantly with, the beginning of the diet. Results: ASO6 reduced liver cholesteryl esters, while not inducing UC accumulation. ASO6 increased hepatic ABCA1 protein independently of the diet conditions. ASO6 affected HDL lipids (increased UC) only in DOKO, while it increased apoE- containing HDL in both genotypes. In WT mice ASO6 led to the appearance of large HDL enriched in apoAI and apoE. Conclusions: The use of ASO6 revealed a new pathway by which the liver may contribute to HDL metabolism in mice. ACAT2 seems to be a hepatic player affecting the cholesterol fluxes fated to VLDL or to HDL, the latter via up-regulation of ABCA1. Citation: Pedrelli M, Davoodpour P, Degirolamo C, Gomaraschi M, Graham M, et al. (2014) Hepatic ACAT2 Knock Down Increases ABCA1 and Modifies HDL Metabolism in Mice. PLoS ONE 9(4): e93552. doi:10.1371/journal.pone.0093552 Editor: Christina Bursill, Heart Research Institute, Australia Received September 27, 2013; Accepted March 6, 2014; Published April 2, 2014 Copyright: ß 2014 Pedrelli et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: The studies described in this paper were supported by the Swedish Heart-Lung Foundation, the Swedish Research Council, the Karolinska Institutet, the Swedish Medical Association, the Stockholm County Council, and Novo Nordisk Research Foundation. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: Mark Graham is an employee and stock holder of Isis Pharmaceuticals Inc. (Carlsbad, CA, USA). Laura Calabresi is a member of the PLOS ONE Editorial Board. This does not alter the authors’ adherence to all the PLOS ONE policies on sharing data and materials. All the other authors have declared that no competing interests exist. * E-mail: paolo.parini@ki.se ¤a Current address: Department of Immunology, Genetics and Pathology, Rudbeck Laboratory, Uppsala University, Uppsala, Sweden ¤b Current address: Department of Translational Pharmacology, Consorzio Mario Negri Sud, S.M. Imbaro (CH), Italy . These authors contributed equally to this work. MP and PD are joint first authors on this work. Introduction For many years, the inhibition of intracellular cholesterol esterification has been considered as a potential strategy to prevent atherosclerosis [1]. Acyl-coenzyme A:cholesterol acyltransferase (ACAT) is an enzyme located in the endoplasmic reticulum that catalyses the synthesis of cholesteryl esters (CE) by conjugating cholesterol to long-chain fatty acids; mainly oleic and palmitic acids. It is now clear that the two enzymes ACAT1 and ACAT2, which are encoded by the Soat1 and Soat2 genes respectively, localize in different cell types and have separate physiological functions (for review see [2,3]). ACAT1 is ubiquitously expressed and provides essential housekeeping functions to prevent the toxicity induced by increasing amounts of unesterified cholesterol (UC) in cells. Conversely, ACAT2 is exclusively expressed in hepatocytes and enterocytes, and it synthesizes CE that can be incorporated into apoB-containing lipoproteins (VLDL and chylomicrons). In mice deletion of Soat1 or Soat2 genes revealed a diverse role for the different ACAT enzymes in atherosclerosis. In Soat1 knockout animals, ACAT1 deficiency led to a marked alteration in cholesterol metabolism resulting in massive accumu- lation of UC, which caused numerous skin and brain lesions, and worsened atherosclerosis. Conversely, the deletion of the Soat2 gene has been consistently atheroprotective [4–6]. Previous studies also suggest a clear atherogenic potential of ACAT2-derived CE also in humans. In both women and men, the Atherosclerosis Risk in Communities (ARIC) study revealed an association between the PLOS ONE | www.plosone.org 1 April 2014 | Volume 9 | Issue 4 | e93552