Nonalcoholic steatohepatitis (NASH) involves liver lipid accumulation (steatosis) combined with hepatic inflammation. The transition towards hepatic inflammation represents a key step in pathogenesis, because it will set the stage for further liver damage, culminating in hepatic fibrosis, cirrhosis, and liver cancer. The actual risk factors that drive hepatic inflammation during the progression to NASH remain largely unknown. The role of steatosis and dietary cholesterol in the etiology of diet-induced NASH was investigated using hyperlipidemic mouse models fed a Western diet. Livers of male and female hyperlipidemic (low-density lipoprotein receptor–deficient [ldlr^−/−] and apolipoprotein E2 knock-in [APOE2ki]) mouse models were compared with livers of normolipidemic wild-type (WT) C57BL/6J mice after short-term feeding with a high-fat diet with cholesterol (HFC) and without cholesterol. Whereas WT mice displayed only steatosis after a short-term HFC diet, female ldlr^−/− and APOE2ki mice showed steatosis with severe inflammation characterized by infiltration of macrophages and increased nuclear factor κB (NF-κB) signaling. Remarkably, male ldlr^−/− and APOE2ki mice developed severe hepatic inflammation in the absence of steatosis after 7 days on an HFC diet compared with WT animals. An HFC diet induced bloated, “foamy” Kupffer cells in male and female ldlr^−/− and APOE2ki mice. Hepatic inflammation was found to be linked to increased plasma very low-density lipoprotein (VLDL) cholesterol levels. Omitting cholesterol from the HFC diet lowered plasma VLDL cholesterol and prevented the development of inflammation and hepatic foam cells. Conclusion: These findings indicate that dietary cholesterol, possibly in the form of modified plasma lipoproteins, is an important risk factor for the progression to hepatic inflammation in diet-induced NASH. (HEPATOLOGY 2008;48:474–486.)

Non-alcoholic steatohepatitis (NASH) involves steatosis combined with inflammation, which can progress into fibrosis and cirrhosis. Exploring the molecular mechanisms of NASH is highly dependent on the availability of animal models. Currently, the most commonly used animal models for NASH imitate particularly late stages of human disease. Thus, there is a need for an animal model that can be used for investigating the factors that potentiate the inflammatory response within NASH. We have previously shown that 7-day high-fat-high-cholesterol (HFC) feeding induces steatosis and inflammation in both APOE2ki and Ldlr^−/− mice. However, it is not known whether the early inflammatory response observed in these mice will sustain over time and lead to liver damage. We hypothesized that the inflammatory response in both models is sufficient to induce liver damage over time. APOE2ki and Ldlr^−/− mice were fed a chow or HFC diet for 3 months. C57Bl6/J mice were used as control. Surprisingly, hepatic inflammation was abolished in APOE2ki mice, while it was sustained in Ldlr^−/− mice. In addition, increased apoptosis and hepatic fibrosis was only demonstrated in Ldlr^−/− mice. Finally, bone-marrow-derived-macrophages of Ldlr^−/− mice showed an increased inflammatory response after oxidized LDL (oxLDL) loading compared to APOE2ki mice. Ldlr^−/− mice, but not APOE2ki mice, developed sustained hepatic inflammation and liver damage upon long term HFC feeding due to increased sensitivity for oxLDL uptake. Therefore, the Ldlr^−/− mice are a promising physiological model particularly vulnerable for investigating the onset of hepatic inflammation in non-alcoholic steatohepatitis.

Rationale:: Nuclear receptor Nur77, also known as NR4A1, TR3, or NGFI-B, is expressed in human atherosclerotic lesions in macrophages, endothelial cells, T cells and smooth muscle cells. Macrophages play a critical role in atherosclerosis and the function of Nur77 in lesion macrophages has not yet been investigated. Objective:: This study aims to delineate the function of Nur77 in macrophages and to assess the effect of bone marrow–specific deficiency of Nur77 on atherosclerosis. Methods and Results:: We investigated Nur77 in macrophage polarization using bone marrow-derived macrophages (BMM) from wild-type and Nur77-knockout (Nur77 ^−/− ) mice. Nur77 ^−/− BMM exhibit changed expression of M2-specific markers and an inflammatory M1-phenotype with enhanced expression of interleukin-12, IFNγ, and SDF-1α and increased NO synthesis in (non)-stimulated Nur77 ^−/− BMM cells. SDF-1α expression in nonstimulated Nur77 ^−/− BMM is repressed by Nur77 and the chemoattractive activity of Nur77 ^−/− BMM is abolished by SDF-1α inhibiting antibodies. Furthermore, Nur77 ^−/− mice show enhanced thioglycollate-elicited migration of macrophages and B cells. The effect of bone marrow–specific deficiency of Nur77 on atherosclerosis was studied in low density lipoprotein receptor-deficient (Ldlr ^−/− ) mice. Ldlr ^−/− mice with a Nur77 ^−/− -deficient bone marrow transplant developed 2.1-fold larger atherosclerotic lesions than wild-type bone marrow–transplanted mice. These lesions contain more macrophages, T cells, smooth muscle cells and larger necrotic cores. SDF-1α expression is higher in lesions of Nur77 ^−/− -transplanted mice, which may explain the observed aggravation of lesion formation. Conclusions:: In conclusion, in bone marrow–derived cells the nuclear receptor Nur77 has an anti-inflammatory function, represses SDF-1α expression and inhibits atherosclerosis.

Background & Aims Nonalcoholic steatohepatitis (NASH) is a disorder that consists of steatosis and hepatic inflammation. It is not known why only some people with steatosis develop NASH. Recently, we identified dietary cholesterol as a factor that directly leads to hepatic inflammation and hepatic foam cell formation. We propose a mechanism by which Kupffer cells (KCs) take up modified cholesterol-rich lipoproteins via scavenger receptors (SRs). KCs thereby accumulate cholesterol, become activated, and may then trigger an inflammatory reaction. Scavenging of modified lipoproteins mainly depends on CD36 and macrophage scavenger receptor 1. Methods To evaluate the involvement of SR-mediated uptake of modified lipoproteins by KCs in the development of diet-induced NASH, female low-density lipoprotein receptor-deficient (Ldlr^−/−) mice were lethally irradiated and transplanted with bone marrow from Msr1^+/+/Cd36^+/+or Msr1^−/−/Cd36^−/− mice and fed a Western diet. Results Macrophage and neutrophil infiltration revealed that hepatic inflammation was substantially reduced by approximately 30% in Msr1^−/−/Cd36^−/−-transplanted mice compared with control mice. Consistent with this, the expression levels of well-known inflammatory mediators were reduced. Apoptotis and fibrosis were less pronounced in Msr1^−/−/Cd36^−/−-transplanted mice, in addition to the protective phenotype of natural antibodies against oxidized low-density lipoprotein in the plasma. Surprisingly, the effect on hepatic inflammation was independent of foam cell formation. Conclusions Targeted inactivation of SR pathways reduces the hepatic inflammation and tissue destruction associated with NASH, independent of hepatic foam cell formation.

Hyperlipidemia is the most important risk factor for atherosclerosis, which is the major cause of cardiovascular disease. The etiology of hyperlipidemia and atherosclerosis is complex and governed by multiple interacting genes. However, mutations in two genes have been shown to be directly involved, i.e., the low-density lipoprotein receptor (LDLR) and apolipoprotein E (ApoE). Genetically modified mouse models have been instrumental in elucidating the underlying molecular mechanisms in lipid metabolism. In this review, we focus on the use of two of the most widely used mouse models, ApoE- and LDLR-deficient mice. After almost a decade of applications, it is clear that each model has unique strengths and drawbacks when carrying out studies of the role of additional genes and environmental factors such as nutrition and lipid-lowering drugs. Importantly, we elaborate on mice expressing mutant forms of APOE, including the