ORIGINAL ARTICLE Macrophage LXRa gene therapy ameliorates atherosclerosis as well as hypertriglyceridemia in LDLR / mice G Li 1 , KC Biju 1 , X Xu 1 , Q Zhou 1 , C Chen 1 , AJ Valente 1 , W He 1 , RL Reddick 2 , GL Freeman 1 , SS Ahuja 1 , RA Clark 1 and S Li 1,3 Liver X receptors (LXRs) are implicated in the regulation of cholesterol homeostasis, inflammatory response and atherogenesis. Administration of LXR agonists inhibits the progress of atherosclerosis, and also increases plasma triglyceride levels, representing an obstacle to their use in treating this disease. The objective of this study was to develop an alternative approach that could overcome this obstacle. Eight-week-old low-density lipoprotein receptor-deficient (LDLR / ) mice were transplanted with hematopoietic stem cell (HSC)-enriched bone marrow cells transduced with lentivectors expressing either green fluorescent protein (GFP) (Lenti-SP-GFP, control) or LXRa (Lenti-SP-LXRa) driven by a synthetic macrophage promoter. At 4 weeks post-transplant, the mice were fed with a Western diet for 8 weeks and then killed. Compared with Lenti-SP-GFP mice, the Lenti-SP-LXRa mice had a 30% reduction in atherosclerotic lesions, which was accompanied by increases in levels of macrophage expression of cholesterol efflux genes apolipoprotein E and ATP-binding cassette A1, as well as decreases in plasma inflammatory cytokines interleukin-6 and tumor necrosis factor-a. Intriguingly, a 50% reduction of plasma triglyceride level was also observed. We conclude that HSC-based macrophage LXRa gene therapy ameliorates the development of atherosclerosis along with an unexpected concomitant reduction of plasma triglyceride levels in LDLR / mice. These findings highlight the potential value of macrophage LXR expression as an avenue for therapeutic intervention against atherosclerosis. Gene Therapy advance online publication, 10 March 2011; doi:10.1038/gt.2011.29 Keywords: LXRa; stem cell gene therapy; macrophage; atherosclerosis; triglyceride INTRODUCTION Liver X receptors a and b (LXRa and LXRb), members of the nuclear receptor transcription factor superfamily, are master regulators of whole-body cholesterol homeostasis, as well as metabolic and inflam- matory signaling integration. 1,2 The LXR transcription factors, form- ing obligate heterodimers with the retinoid X receptor (RXR), regulate target gene expression through interaction with LXR response elements in the regulatory regions of these genes. The internal ligands for LXRs are oxidized derivatives of cholesterol (oxysterols). LXRa is highly expressed in the liver and selectively at lower levels in other tissues involved in lipid homeostasis, such as intestine, adipocytes and macrophages, whereas LXRb is expressed ubiquitously. 1 It is generally believed that the two isoforms can compensate for each other in biological activity. 3,4 Ligand activation of the LXRs decreases intestinal cholesterol absorption efficiency, induces cholesterol efflux from lipid- laden macrophages and negatively regulates inflammatory gene expression, suggesting a beneficial effect on the development of cardiovascular disease. Direct evidence for a role of the LXR pathway in atherosclerosis susceptibility has been obtained from studies using genetically modified mice. Apolipoprotein E-deficient (apoE / ) or low-density lipoprotein receptor-deficient (LDLR / ) mice trans- planted with bone marrow cells from LXRa/b double-knockout (LXRa/b / ) animals exhibited significantly greater atherosclerotic lesion development than their counterparts receiving bone marrow cells from wild-type mice. 5 In addition, LXRa/b / mice exhibit increased cholesterol accumulation in arterial wall macrophages even on normal chow diets. Consistently, activation of LXRs by synthetic agonists provides protection against lesion development and leads to regression of established lesions in mouse models. 6,7 However, administration of synthetic LXR agonists triggers induction of the hepatic lipogenic pathway and causes hypertriglyceridemia. This undesirable effect is a significant obstacle in the development of LXR agonists as potential drugs. 8,9 Atherosclerosis is a disorder of lipid metabolism, as well as a chronic inflammatory disease. 10 Macrophages, participating in both lipid metabolism and inflammation, are intimately involved in all phases of atherosclerosis, from development of the fatty streak to processes that ultimately contribute to plaque rupture and myocardial infarction. 11 Macrophage expression of certain genes is atheroprotec- tive, whereas inadequate expression or lack of expression of these same genes in macrophages increases atherosclerosis. 12 It has been reported that macrophage LXRa is required for antiatherogenic activity of LXRa agonists. 7 We hypothesized that macrophage-specific over- expression of LXRa would lead to the reduction of atherosclerosis without elevation of liver-derived plasma triglycerides, an adverse effect associated with synthetic LXR agonist treatment. Using the macrophage-specific promoter/lentiviral system we developed previously, 13 bone marrow stem cells were manipulated to express Received 15 October 2009; revised 2 August 2010; accepted 4 August 2010 1 Department of Medicine, University of Texas Health Science Center, and the South Texas Veterans Health Care System, Audie L Murphy Division, San Antonio, TX, USA; 2 Department of Pathology, University of Texas Health Science Center, and the South TexasVeterans Health Care System, Audie L Murphy Division, San Antonio, TX, USA and 3 Department of Pharmacology, University of Texas Health Science Center, and the South Texas Veterans Health Care System, Audie L Murphy Division, San Antonio, TX, USA Correspondence: Dr S Li, Department of Medicine, University of Texas Health Science Center, and the South Texas Veterans Health Care System, Audie L Murphy Division, 7703 Floyd Curl Drive, San Antonio, TX 78229-3900. E-mail: lis1@uthscsa.edu Gene Therapy (2011), 1–7 & 2011 Macmillan Publishers Limited All rights reserved 0969-7128/11 www.nature.com/gt