Therapeutic Role of Ursolic Acid on Ameliorating Hepatic Steatosis and Improving Metabolic Disorders in High-Fat Diet-Induced Non-Alcoholic Fatty Liver Disease Rats

Background

Non-alcoholic fatty liver disease (NAFLD) is one of the most prevalent liver diseases around the world, and is closely associated with obesity, diabetes, and insulin resistance. Ursolic acid (UA), an ubiquitous triterpenoid with multifold biological roles, is distributed in various plants. This study was conducted to investigate the therapeutic effect and potential mechanisms of UA against hepatic steatosis in a high-fat diet (HFD)-induced obese non-alcoholic fatty liver disease (NAFLD) rat model.

Methodology/Principal Findings

Obese NAFLD model was established in Sprague-Dawley rats by 8-week HFD feeding. Therapeutic role of UA was evaluated using 0.125%, 0.25%, 0.5% UA-supplemented diet for another 6 weeks. The results from both morphologic and histological detections indicated that UA significantly reversed HFD-induced hepatic steatosis and liver injury. Besides, hepatic peroxisome proliferator-activated receptor (PPAR)-α was markedly up-regulated at both mRNA and protein levels by UA. Knocking down PPAR-α significantly inhibited the anti-steatosis role of UA in vitro. HFD-induced adverse changes in the key genes, which participated in hepatic lipid metabolism, were also alleviated by UA treatment. Furthermore, UA significantly ameliorated HFD-induced metabolic disorders, including insulin resistance, inflammation and oxidative stress.

Conclusions/Significance

These results demonstrated that UA effectively ameliorated HFD-induced hepatic steatosis through a PPAR-α involved pathway, via improving key enzymes in the controlling of lipids metabolism. The metabolic disorders were accordingly improved with the decrease of hepatic steatosis. Thereby, UA could be a promising candidate for the treatment of NAFLD.     

GCN2 deficiency protects against high fat diet induced hepatic steatosis and insulin resistance in mice

Nonalcoholic fatty liver disease (NAFLD) is characterized by hepatic lipid deposition and oxidative stress. It has been demonstrated that general control nonderepressible 2 (GCN2) is required to maintain hepatic fatty acid homeostasis under conditions of amino acid deprivation. However, the impact of GCN2 on the development of NAFLD has not been investigated. In this study, we used Gcn2^?/? mice to investigate the effect of GCN2 on high fat diet (HFD)-induced hepatic steatosis. After HFD feeding for 12?weeks, Gcn2^?/? mice were less obese than wild-type (WT) mice, and Gcn2^?/? significantly attenuated HFD-induced liver dysfunction, hepatic steatosis and insulin resistance. In the livers of the HFD-fed mice, GCN2 deficiency resulted in higher levels of lipolysis genes, lower expression of genes related to FA synthesis, transport and lipogenesis, and less induction of oxidative stress. Furthermore, we found that knockdown of GCN2 attenuated, whereas overexpression of GCN2 exacerbated, palmitic acid-induced steatosis, oxidative & ER stress, and changes of peroxisome proliferator-activated receptor gamma (PPARγ), fatty acid synthase (FAS) and metallothionein (MT) expression in HepG2 cells. Collectively, our data provide evidences that GCN2 deficiency protects against HFD-induced hepatic steatosis by inhibiting lipogenesis and reducing oxidative stress. Our findings suggest that strategies to inhibit GCN2 activity in the liver may provide a novel approach to attenuate NAFLD development.     

Vitamin E alleviates non-alcoholic fatty liver disease in phosphatidylethanolamine N-methyltransferase deficient mice

Phosphatidylethanolamine N-methyltransferase (PEMT) converts phosphatidylethanolamine (PE) to phosphatidylcholine (PC), mainly in the liver. Pemt^?/? mice are protected from high-fat diet (HFD)-induced obesity and insulin resistance, but develop severe non-alcoholic fatty liver disease (NAFLD) when fed a HFD, mostly due to impaired VLDL secretion. Oxidative stress is thought to be an essential factor in the progression from simple steatosis to steatohepatitis. Vitamin E is an antioxidant that has been clinically used to improve NAFLD pathology. Our aim was to determine whether supplementation of the diet with vitamin E could attenuate HFD-induced hepatic steatosis and its progression to NASH in Pemt^?/? mice. Treatment with vitamin E (0.5?g/kg) for 3?weeks improved VLDL-TG secretion and normalized cholesterol metabolism, but failed to reduce hepatic TG content. Moreover, vitamin E treatment was able to reduce hepatic oxidative stress, inflammation and fibrosis. We also observed abnormal ceramide metabolism in Pemt^?/? mice fed a HFD, with elevation of ceramides and other sphingolipids and higher expression of mRNAs for acid ceramidase (Asah1) and ceramide kinase (Cerk). Interestingly, vitamin E supplementation restored Asah1 and Cerk mRNA and sphingolipid levels. Together this study shows that vitamin E treatment efficiently prevented the progression from simple steatosis to steatohepatitis in mice lacking PEMT.     

Hepatic HuR protects against the pathogenesis of non-alcoholic fatty liver disease by targeting PTEN

The liver plays an important role in lipid and glucose metabolism. Here, we show the role of human antigen R (HuR), an RNA regulator protein, in hepatocyte steatosis and glucose metabolism. We investigated the level of HuR in the liver of mice fed a normal chow diet (NCD) and a high-fat diet (HFD). HuR was downregulated in the livers of HFD-fed mice. Liver-specific HuR knockout (HuR^LKO) mice showed exacerbated HFD-induced hepatic steatosis along with enhanced glucose tolerance as compared with control mice. Mechanistically, HuR could bind to the adenylate uridylate-rich elements of phosphatase and tensin homolog deleted on the chromosome 10 (PTEN) mRNA 3′ untranslated region, resulting in the increased stability of Pten mRNA; genetic knockdown of HuR decreased the expression of PTEN. Finally, lentiviral overexpression of PTEN alleviated the development of hepatic steatosis in HuR^LKO mice in vivo. Overall, HuR regulates lipid and glucose metabolism by targeting PTEN.Subject terms: Type 2 diabetes, Dyslipidaemias     

Hesperidin methyl chalcone ameliorates lipid metabolic disorders by activating lipase activity and increasing energy metabolism

Obesity has become an increasingly serious health issue with the continuous improvement in living standards. Its prevalence has become an economic burden on health care systems worldwide. Flavonoids have been shown to be beneficial in the prevention and treatment of obesity. Here, we evaluated the therapeutic potential of the flavonoid hesperidin methyl chalcone (HMC) on mice with high-fat diet (HFD)-induced hepatic steatosis in vivo and in vitro. Treatment with HMC reduced oleic and palmitic acid-induced increases in intracellular triglyceride accumulation in HepG2, AML12 and LMH cells. HMC also enhanced energy metabolism and lowered oxidative stress. We used Discovery studio to dock key proteins associated with lipid metabolism disorders to HMC, and found that HMC interacted with lipase. Furthermore, we demonstrated that HMC improved lipase activity and lipolysis. In addition, we found that HMC promoted glucose absorption, alleviated lipid metabolic disorders, improved HFD-induced liver injury, and regulated HFD-induced changes in energy metabolism. In conclusion, our study demonstrated that HMC ameliorated HFD-induced obesity and its complications by promoting lipase activity, and provides a novel approach for the prevention and treatment of obesity and related diseases.     

Reversal of high-fat diet-induced hepatic steatosis by n-3 LCPUFA: role of PPAR-α and SREBP-1c

Nonalcoholic fatty liver disease is characterized by an abnormal accumulation of triacylglycerides in the liver in absence of significant alcohol consumption. Under these conditions, it has been observed an impaired bioavailability of hepatic n-3 long-chain polyunsaturated fatty acids (LCPUFAs). The aim of this study was to test the reversion of the prosteatotic and proinflammatory effects of high-fat diet (HFD) in the mouse liver by changing to normocaloric diet and n-3 LCPUFA supplementation. Male C57BL/6J mice were given either control diet (CD) or HFD for 12 weeks. Control and HFD groups were divided into subgroups that continue with CD or subjected to CD plus n-3 LCPUFA for 8 additional weeks. After this time, blood and liver samples were taken and metabolic, morphologic, oxidative stress, inflammatory and signaling parameters were analyzed. The dietary change from HFD to a normocaloric diet with n-3 LCPUFA supplementation significantly reduced insulin resistance and liver steatosis when compared to switching HFD to normocaloric diet alone. In addition, HFD-induced increases in adiposity, adipocyte enlargement and liver oxidative stress and inflammatory cytokine expression were suppressed by n-3 LCPUFA to control values. Importantly, n-3 LCPUFA supplementation abolish HFD-induced enhancement in hepatic SREBP-1c/PPAR-α ratios, suggesting a change in the metabolic status of the liver from a lipogenic condition to one favoring fatty acid oxidation and steatosis attenuation. These findings may provide the rational basis for the use of normocaloric diets supplemented with n-3 LCPUFA in patients with liver steatosis.     

Targeting the gut microbiota with resveratrol: a demonstration of novel evidence for the management of hepatic steatosis

Resveratrol is a natural polyphenol that has been reported to reduce the risk of obesity and nonalcoholic fatty liver disease (NAFLD). Recent evidence has demonstrated that the gut microbiota plays an important role in the protection against NAFLD and other metabolic diseases. The present study aimed to investigate the relationship between the gut microbiota and the beneficial effects of resveratrol on the amelioration of NAFLD in mice. We observed marked decreases in body weight and liver steatosis and improved insulin resistance in high-fat diet (HFD)-fed mice treated with resveratrol. Furthermore, we found that resveratrol treatment alleviated NAFLD in HFD-fed mice by improving the intestinal microenvironment, including gut barrier function and gut microbiota composition. On the one hand, resveratrol improved gut intestinal barrier integrity through the repair of intestinal mucosal morphology and increased the expression of physical barrier- and physiochemical barrier-related factors in HFD-fed mice. On the other hand, in HFD-fed mice, resveratrol supplementation modulated the gut bacterial composition. The resveratrol-induced gut microbiota was characterized by a decreased abundance of harmful bacteria, including Desulfovibrio, Lachnospiraceae_NK4A316_group and Alistipes, as well as an increased abundance of short-chain fatty acid (SCFA)-producing bacteria, such as Allobaculum, Bacteroides and Blautia. Moreover, transplantation of the HFDR-microbiota into HFD-fed mice sufficiently decreased body weight, liver steatosis and low-grade inflammation and improved hepatic lipid metabolism. Collectively, resveratrol would provide a potentially dietary intervention strategy against NAFLD through modulating the intestinal microenvironment.     

Involvement of G protein-coupled receptor kinase 2 (GRK2) in the development of non-alcoholic steatosis and steatohepatitis in mice and humans

Insulin resistance (IR) and obesity are important risk factors for non-alcoholic fatty liver disease (NAFLD). G protein-coupled receptor kinase 2 (GRK2) is involved in the development of IR and obesity in vivo. However, its possible contribution to NAFLD and/or non-alcoholic steatohepatitis (NASH) independently of its role on IR or fat mass accretion has not been explored. Here, we used wild-type (WT) or GRK2 hemizygous (GRK2±) mice fed a high-fat diet (HFD) or a methionine and choline-deficient diet (MCD) as a model of NASH independent of adiposity and IR. GRK2± mice were protected from HFD-induced NAFLD. Moreover, MCD feeding caused an increased in triglyceride content and liver-to-body weight ratio in WT mice, features that were attenuated in GRK2± mice. According to their NAFLD activity score, MCD-fed GRK2± mice were diagnosed with simple steatosis and not overt NASH. They also showed reduced expression of lipogenic and lipid-uptake markers and less signs of inflammation in the liver. GRK2± mice preserved hepatic protective mechanisms as enhanced autophagy and mitochondrial fusion and biogenesis, together with reduced endoplasmic reticulum stress. GRK2 protein was increased in MCD-fed WT but not in GRK2± mice, and enhanced GRK2 expression potentiated palmitic acid-triggered lipid accumulation in human hepatocytes directly relating GRK2 levels to steatosis. GRK2 protein and mRNA levels were increased in human liver biopsies from simple steatosis or NASH patients in two different human cohorts. Our results describe a functional relationship between GRK2 levels and hepatic lipid accumulation and implicate GRK2 in the establishment and/or development of NASH.     

Structural changes of gut microbiota in a rat non-alcoholic fatty liver disease model treated with a Chinese herbal formula

Accumulating evidence indicates that disruption of the gut microbiota by a high-fat diet (HFD) may play a pivotal role in the progression of metabolic disorders such as non-alcoholic fatty liver disease (NAFLD). In this study, the structural changes of gut microbiota were analyzed in an HFD-induced NAFLD rat model during treatment with an ancient Chinese herbal formula (CHF) used in clinical practice – Qushi Huayu Fang. CHF treatment significantly reduced body weight, alleviated hepatic steatosis, and decreased the content of triglycerides and free fatty acids in the livers of the rats. Gut microbiota of treated and control rats were profiled with polymerase chain reaction-denaturing gradient gel electrophoresis and bar-coded pyrosequencing of the V3 region of 16S rRNA genes. Both analyses indicated that the CHF-treated group harbored significantly different gut microbiota from that of model rats. Partial least squares discriminant analysis and taxonomy-based analysis were further employed to identify key phylotypes responding to HFD and CHF treatment. Most notably, the genera Escherichia/Shigella, containing opportunistic pathogens, were significantly enriched in HFD-fed rats compared to controls fed normal chow (P < 0.05) but they decreased to control levels after CHF treatment. Collinsella, a genus with short chain fatty acid producers, was significantly elevated in CHF-treated rats compared to HFD-fed rats (P < 0.05). The results revealed that the bacterial profiles of HFD-induced rats could be modulated by the CHF. Elucidation of these differences in microbiota composition provided a basis for further understanding the pharmacological mechanism of the CHF.     

Immune response gene 1 deficiency aggravates high fat diet-induced nonalcoholic fatty liver disease via promotion of redox-sensitive AKT suppression

Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disorder worldwide. Immune response gene 1 (IRG1) catalyzes the production of bio-active itaconate, which is actively involved in the regulation of signal transduction. A recent study has found that the expression of IRG1 was significantly down-regulated in obesity-associated fatty liver, but the potential roles of IRG1 in the development NAFLD remain unclear. The present study found that genetic deletion of IRG1 aggravated high fat diet (HFD)-induced metabolic disturbance, including obesity, dyslipidemia and insulin resistance. In addition, HFD induced more severe liver steatosis and higher serum ALT and AST level in IRG1 KO mice, which were accompanied with altered expression of genes involved in lipid uptake, synthesis and catabolism. RNA-seq and immunoblot analysis indicated that deficiency of IRG1 is associated with suppressed activation of AKT, a master metabolic regulator. Mechanistically, IRG1/itaconate enhanced the antioxidative NRF2 pathway and prevented redox-sensitive suppression of AKT. Interestingly, supplementation with 4-octyl itaconate (4-OI), a cell-permeable derivate of itaconate, alleviated HFD-induced oxidative stress, AKT suppression and liver steatosis. Therefore, IRG1 probably functions as a protective regulator in the development of NAFLD and the cell-permeable 4-OI might have potential value for the pharmacological intervention of NAFLD.     

N-3 PUFAs inhibited hepatic ER stress induced by feeding of a high-saturated fat diet accompanied by the expression LOX-1

Excessive consumption of saturated fat leads to non-alcoholic fatty liver disease (NAFLD), which is attenuated by supplementation of n-3 polyunsaturated fatty acids (PUFAs). Endoplasmic reticulum (ER) stress is crucial in the development of NAFLD, but how high-saturated fat diet (HFD) causes ER stress and NAFLD remains unclear. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is involved in hepatic ER stress. We aimed to explore the roles of LOX-1 in HFD-induced ER stress. Male Sprague–Dawley rats were fed an HFD without or with supplementation of fish oil for 16 weeks. The effects of n-3 PUFAs on hepatic ER stress degrees and the expression levels of LOX-1 were examined. Then human L02 hepatoma cells were treated with palmitate or palmitate and DHA to determine the ER stress and LOX-1 expression levels in vitro. After that the expression of LOX-1 in L02 cells was either knocked-down or overexpressed to analyze the roles of LOX-1 in palmitate-induced ER stress. The feeding of HFD induced NAFLD development and ER stress in the liver, and LOX-1 expressing level, which were all reversed by fish oil supplementation. In vitro, DHA treatment reduced the expression of LOX-1, and palmitate-induced ER stress. SiRNA-mediated knock-down of LOX-1 inhibited palmitate-induced ER stress, whereas overexpression of LOX-1 dramatically induced ER stress in L02 cells.LOX-1 is critical for HFD-induced ER stress, and inhibition of its expression under the treatment of n-3 PUFAs could ameliorate HFD-induced NAFLD.     

Betaine attenuates hepatic steatosis by reducing methylation of the MTTP promoter and elevating genomic methylation in mice fed a high-fat diet

Aberrant DNA methylation contributes to the abnormality of hepatic gene expression, one of the main factors in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). Betaine is a methyl donor and has been considered to be a lipotropic agent. However, whether betaine supplementation improves NAFLD via its effect on the DNA methylation of specific genes and the genome has not been explored. Male C57BL/6 mice were fed either a control diet or high-fat diet (HFD) supplemented with 0%, 1% and 2% betaine in water (wt/vol) for 12 weeks. Betaine supplementation ameliorated HFD-induced hepatic steatosis in a dose-dependent manner. HFD up-regulated FAS and ACOX messenger RNA (mRNA) expression and down-regulated PPARα, ApoB and MTTP mRNA expression; however, these alterations were reversed by betaine supplementation, except ApoB. MTTP mRNA expression was negatively correlated with the DNA methylation of its CpG sites at −184, −156, −63 and −60. Methylation of these CpG sites was lower in both the 1% and 2% betaine-supplemented groups than in the HFD group (averages; 25.55% and 14.33% vs. 30.13%). In addition, both 1% and 2% betaine supplementation significantly restored the methylation capacity [S-adenosylmethionine (SAM) concentration and SAM/S-adenosylhomocysteine ratios] and genomic methylation level, which had been decreased by HFD (0.37% and 0.47% vs. 0.25%). These results suggest that the regulation of aberrant DNA methylation by betaine might be a possible mechanism of the improvements in NAFLD upon betaine supplementation.     

Reduction of high-fat diet-induced liver proinflammatory state by eicosapentaenoic acid plus hydroxytyrosol supplementation: involvement of resolvins RvE1/2 and RvD1/2

High-fat diet (HFD)-fed mice show obesity with development of liver steatosis and a proinflammatory state without establishing an inflammatory reaction. The aim of this work was to assess the hypothesis that eicosapentaenoic acid (EPA) plus hydroxytyrosol (HT) supplementation prevents the inflammatory reaction through enhancement in the hepatic resolvin content in HFD-fed mice. Male C57BL/6J mice were fed an HFD or a control diet and supplemented with EPA (50 mg/kg/day) and HT (5 mg/kg/day) or their respective vehicles for 12 weeks. Measurements include liver levels of EPA, DHA and palmitate (gas chromatography), liver resolvins and triglyceride (TG) and serum aspartate transaminase (AST) (specific kits) and hepatic and serum inflammatory markers (quantitative polymerase chain reaction and enzyme-linked immunosorbent assay). Compared to CD, HFD induced body weight gain, liver steatosis and TG accumulation, with up-regulation of proinflammatory markers in the absence of histological inflammation or serum AST changes; these results were accompanied by higher hepatic levels of resolvins RvE1, RvE2, RvD1 and RvD2, with decreases in EPA and DHA contents. EPA+HT supplementation in HFD feeding synergistically reduced the steatosis score over individual treatments and increased the hepatic levels of EPA, DHA and resolvins, with attenuation of proinflammatory markers. Lack of progression of HFD-induced proinflammatory state into overt inflammation is associated with resolvin up-regulation, which is further increased by EPA+HT supplementation eliciting steatosis attenuation. These findings point to the importance of combined protocols in hepatoprotection due to the involvement of cross-talk mechanisms, which increase effectiveness and diminish dosages, avoiding undesirable effects.     

Mesenchymal stromal cells protect hepatocytes from lipotoxicity through alleviation of endoplasmic reticulum stress by restoring SERCA activity

The aim of this study was to investigate how mesenchymal stromal cells (MSCs) modulate metabolic balance and attenuate hepatic lipotoxicity in the context of non-alcoholic fatty liver disease (NAFLD). In vivo, male SD rats were fed with high-fat diet (HFD) to develop NAFLD; then, they were treated twice by intravenous injections of rat bone marrow MSCs. In vitro, HepG2 cells were cocultured with MSCs by transwell and exposed to palmitic acid (PA) for 24 hours. The endoplasmic reticulum (ER) stressor thapsigargin and sarco/ER Ca²⁺-ATPase (SERCA2)–specific siRNA were used to explore the regulation of ER stress by MSCs. We found that MSC administration improved hepatic steatosis, restored systemic hepatic lipid and glucose homeostasis, and inhibited hepatic ER stress in HFD-fed rats. In hepatocytes, MSCs effectively alleviated the cellular lipotoxicity. Particularly, MSCs remarkably ameliorated the ER stress and intracellular calcium homeostasis induced by either PA or thapsigargin in HepG2 cells. Additionally, long-term HFD or PA stimulation would activate pyroptosis in hepatocytes, which may contribute to the cell death and liver dysfunction during the process of NAFLD, and MSC treatment effectively ameliorates these deleterious effects. SERCA2 silencing obviously abolished the ability of MSCs against the PA-induced lipotoxicity. Conclusively, our study demonstrated that MSCs were able to ameliorate liver lipotoxicity and metabolic disturbance in the context of NAFLD, in which the regulation of ER stress and the calcium homeostasis via SERCA has played a key role.     

Hepatocyte-specific deletion of Janus kinase 2 (JAK2) protects against diet-induced steatohepatitis and glucose intolerance

Non-alcoholic fatty liver disease (NAFLD) is becoming the leading cause of chronic liver disease and is now considered to be the hepatic manifestation of the metabolic syndrome. However, the role of steatosis per se and the precise factors required in the progression to steatohepatitis or insulin resistance remain elusive. The JAK-STAT pathway is critical in mediating signaling of a wide variety of cytokines and growth factors. Mice with hepatocyte-specific deletion of Janus kinase 2 (L-JAK2 KO mice) develop spontaneous steatosis as early as 2 weeks of age. In this study, we investigated the metabolic consequences of jak2 deletion in response to diet-induced metabolic stress. To our surprise, despite the profound hepatosteatosis, deletion of hepatic jak2 did not sensitize the liver to accelerated inflammatory injury on a prolonged high fat diet (HFD). This was accompanied by complete protection against HFD-induced whole-body insulin resistance and glucose intolerance. Improved glucose-stimulated insulin secretion and an increase in β-cell mass were also present in these mice. Moreover, L-JAK2 KO mice had progressively reduced adiposity in association with blunted hepatic growth hormone signaling. These mice also exhibited increased resting energy expenditure on both chow and high fat diet. In conclusion, our findings indicate a key role of hepatic JAK2 in metabolism such that its absence completely arrests steatohepatitis development and confers protection against diet-induced systemic insulin resistance and glucose intolerance.     

Nonalcoholic fatty liver disease impairs the cytochrome P-450-dependent metabolism of α-tocopherol (vitamin E)

This study aims to investigate in in vivo and in vitro models of nonalcoholic fatty liver disease (NAFLD) the enzymatic metabolism of α-tocopherol (vitamin E) and its relationship to vitamin E-responsive genes with key role in the lipid metabolism and detoxification of the liver. The experimental models included mice fed a high-fat diet combined or not with fructose (HFD+F) and HepG2 human hepatocarcinoma cells treated with the lipogenic agents palmitate, oleate or fructose. CYP4F2 protein, a cytochrome P-450 isoform with proposed α-tocopherol ω-hydroxylase activity, decreased in HFD and even more in HFD+F mice liver; this finding was associated with increased hepatic levels of α-tocopherol and decreased formation of the corresponding long-chain metabolites α-13-hydroxy and α-13-carboxy chromanols. A decreased expression was also observed for PPAR-γ and SREBP-1 proteins, two vitamin E-responsive genes with key role in lipid metabolism and CYP4F2 gene regulation. A transient activation of CYP4F2 gene followed by a repression response was observed in HepG2 cells during the exposure to increasing levels of the lipogenic and cytotoxic agent palmitic acid; such gene repression effect was further exacerbated by the co-treatment with oleic acid and α-tocopherol and was also observed for PPAR-γ and the SREBP isoforms 1 and 2. Such gene response was associated with increased uptake and ω-hydroxylation of α-tocopherol, which suggests a minor role of CYP4F2 in the enzymatic metabolism of vitamin E in HepG2 cells. In conclusion, the liver metabolism and gene response of α-tocopherol are impaired in experimental NAFLD.     

Involvement of SIRT1–AMPK signaling in the protective action of indole-3-carbinol against hepatic steatosis in mice fed a high-fat diet

This study addressed the effect of indole-3-carbinol (I3C) supplementation on hepatic steatosis in mice fed a high-fat diet (HFD) and clarified the underlying mechanism. Male C57BL/6N mice were divided into three groups: those who received a normal diet, those fed with HFD and those fed with 0.1% I3C-supplemented diet (I3CD). In the present study, an HFD supplemented with 0.1% I3C significantly decreased body and liver weight as well as plasma and hepatic lipid levels. The activation of the silent mating type information regulation 2 homolog 1 (SIRT1)–AMP-activated protein kinase (AMPK) signaling system by I3C correlated with decreased mRNA levels of sterol regulatory element-binding protein-1c-regulated lipogenic enzymes. In addition, I3C significantly reversed HFD-induced up-regulation of ER stress-mediated signaling molecules in the liver, which may have contributed to the protective effects of I3C against hepatic steatosis. Furthermore, HFD-induced up-regulations of inflammatory genes such as tumor necrosis factor α and interleukin 6 were significantly reversed by dietary I3C supplementation. Our study suggests that the protective action of I3C against hepatic steatosis is mediated, at least in part, through the up-regulation of a SIRT1–AMPK signaling system in the livers of HFD-fed mice. Further investigations revealed that alleviation of the ER stress response represented a critical mechanism underlying the beneficial effects of I3C on hepatic steatosis.     

Berberine reduces methylation of the MTTP promoter and alleviates fatty liver induced by a high-fat diet in rats

High-calorie food leads to nonalcoholic fatty liver disease (NAFLD) through dysregulation of genes involved in lipid metabolism, but the precise mechanism remains unclear. DNA methylation represents one of the mechanisms that contributes to dysregulation of gene expression via interaction with environmental factors. Berberine can alleviate fatty liver in db/db and ob/ob mice. Here, we investigated whether DNA methylation is involved in the pathogenesis of NAFLD induced by a high-fat diet (HFD) and whether berberine improves NAFLD through influencing the methylation status of promoters of key genes. HFD markedly decreased the mRNA levels encoding CPT-1α, MTTP, and LDLR in the liver. In parallel, DNA methylation levels in the MTTP promoter of rats with NAFLD were elevated in the liver. Interestingly, berberine reversed the downregulated expression of these genes and selectively inhibited HFD-induced increase in the methylation of MTTP. Consistently, berberine increased hepatic triglyceride (TG) export and ameliorated HFD-induced fatty liver. Furthermore, a close negative correlation was observed between the MTTP expression and its DNA methylation (at sites −113 and −20). These data indicate that DNA methylation of the MTTP promoter likely contributes to its downregulation during HFD-induced NAFLD and, further, that berberine can partially counteract the HFD-elicited dysregulation of MTTP by reversing the methylation state of its promoter, leading to reduced hepatic fat content.