Establishment of a methodology for investigating protectants against ethanol-induced hepatotoxicity

Ethanol-induced liver injury has been extensively reported in clinic, but still lacks an efficient in vitro platform for investigating its hepatotoxicity and protectants. This study aimed to establish a methodology on the culture conditions regarding the sealability against evaporation of ethanol, culture medium and 2D/3D culture of hepatocytes. Based on the experimental findings, it was indicated that the ethanol evaporation from culture plates was a severe problem reducing its toxicity in hepatocyte. According to the detected ethanol toxic response marked by reduced cell viability, 3D cultured hepatocytes in gel entrapment were suggested to be better than 2D hepatocyte in monolayer, but the cultures in either William’s Medium E or DMEM exhibited comparable sensitivity to ethanol toxicity. Subsequently, 3D cultured hepatocytes with Parafilm sealing were systematically illustrated to well reflect the ethanol-induced lipid accumulation, reactive oxygen species/malondialdehyde generation, glutathione depletion and cytochrome 2E1 induction. Finally, such hepatocyte models were proposed as a platform for screening of herbal component against ethanol hepatotoxicity. Nano-silibinin, for the first time, found to perform significant protection against ethanol-induced hepatotoxicity while silibinin in normal particles could not inhibit such toxicity. This protection of nano-silibinin might relate to its improved bioavailability compared to normal insoluble silibinin and could act as an anti-oxidative and anti-steatosis agent against ethanol-induced hepatotoxicity.     

In vitro screening for antihyperlipidemic activities in foodstuffs by evaluating lipoprotein profiles secreted from human hepatoma cells

We screened the antihyperlipidemic effects of seven edible plants by evaluation of the triglyceride (TG) and cholesterol profiles secreted from HepG2 cells. We found that the water- and ethanol-extracts of Brasenia schreberi at 100 μg/ml exhibited strong inhibitory activities against TG and cholesterol secretions from HepG2 cells stimulated with sodium oleate. Real-time RT-PCR analysis demonstrated that ethanol extract of B. schreberi (BSET) attenuated the expression of the sterol regulatory element binding protein-1c and -2, fatty acid synthase and HMG CoA synthase-1 genes, which are involved in lipid synthesis in hepatocyte/hepatoma cells. Furthermore, we studied the action of BSET on adipose tissue accumulation and serum parameters in mice fed a high-fat diet (HFD). BSET suppressed mesenteric and epididymal adipose tissue accumulation and normalized serum TG and glucose, but not cholesterol levels in HFD-fed mice.     

Salvia miltiorrhiza Bunge and its active component cryptotanshinone protects primary cultured rat hepatocytes from acute ethanol-induced cytotoxicity and fatty infiltration

Alcoholic liver disease involves hepatocellular injury induced by the acute or chronic consumption of ethanol. Fatty infiltration is usually followed by inflammation and focal necrosis, which can lead to cirrhosis if not treated properly in the initial stage. There have been many attempts to develop effective therapies for the disease, using natural products derived from medicinal plants. In this study, we report that the standardized fraction of Salvia miltiorrhiza Bunge (Sm-SF) and its active component, cryptotanshinone, were able to protect hepatocytes from lipopolysaccharide- and ethanol-induced cell death. They also suppressed ethanol-induced lipid accumulation as evidenced by the Nile red binding assay. The ethanol-induced activation and nuclear translocation of sterol regulatory element-binding protein-1 and the consequent transactivation of the target genes involved in fatty acid biosynthesis were inhibited by Sm-SF and cryptotanshinone in a dose-dependent manner. Cryptotanshinone, an active component of S. miltiorrhiza, has the potential to ameliorate alcoholic liver disease by blocking hepatic cell death and fatty acid synthesis.     

Ethanol self-administration and reinstatement of ethanol-seeking behavior in Per1 Brdm1 mutant mice

Rationale Alcohol consumption shows circadian rhythmicity, i.e., alcohol preference and intake change with circadian time. Circadian rhythmicity is controlled by a biological clock, which has been shown to govern behavioral, physiological, and hormonal processes in synchronization with internal as well as external cues. Molecular components of the clock include circadian clock genes such as period (Per) 1, 2, and 3. Previously, our lab demonstrated the involvement of mouse Per1 (mPer1) and Per2 (mPer2) in modulating cocaine sensitization and reward. What is more, we investigated voluntary alcohol consumption in Per2 ^Brdm1 mice with the results suggesting a relationship between this circadian clock gene and ethanol consumption. Objective To further complement the mPer2 study, our lab proceeded to assess mPer1’s possible role on alcohol intake using operant and free choice two bottle paradigms.Methods Using operant conditions, Per1 ^Brdm1 and wild type mice were trained to self-administer ethanol (10%) under a fixed ratio 1 (FR1) paradigm. This was ensued by a progressive ratio (PR) schedule. Furthermore, extinction sessions were introduced, followed by reinstatement measures of ethanol-seeking behavior. In another set of animals, the mice were exposed to voluntary long-term alcohol consumption, ensued by a 2-month deprivation phase, after which the alcohol deprivation effect (ADE) was measured.Results Mutant mice did not display a significantly divergent number of reinforced lever presses (FR1 and PR) than wild type animals. Furthermore, no significant differences between groups were obtained regarding reinstatement of ethanol-seeking behavior. Similar results were obtained in the two bottle free choice paradigm. Specifically, no genotype differences concerning consumption and preference were observed over a broad range of different ethanol concentrations. Moreover, after the deprivation phase, both groups exhibited significant ADEs, yet no genotype differences.Conclusions Contrary to the mPer2 data, the present findings do not suggest a relationship between the circadian clock gene mPer1 and ethanol reinforcement, seeking, and relapse behavior.     

Hepatoprotective effects of chestnut (Castanea crenata) inner shell extract against chronic ethanol-induced oxidative stress in C57BL/6 mice

This study was carried out to evaluate the protective effects of chestnut inner shell extract (CISE) on chronic ethanol-induced oxidative stress in liver. Mice were fed a control liquid diet (Normal-control), liquid diet containing ethanol alone (EtOH + Vehicle), or were administered CISE and ethanol (EtOH + CISE) for 6 weeks. Administration of ethanol induced liver damage with significant increase of plasma GOT, GPT, hepatic triglyceride (TG) and thiobarbituric acid reactive substance (TBARS) levels. By contrast, co-treatment of CISE with ethanol significantly decreased the activities of GOT and GPT in the plasma, and hepatic TG and TBARS levels. Histological observations were consistent with the result obtained from hepatic lipid quantification. Moreover, CISE treatment with ethanol decreased CYP2E1 expression and increased activities of catalase and superoxide dismutase, which were significantly inhibited by treatment with ethanol alone. To determine the active compound of CISE, fractionation of CISE was conducted and scoparone and scopoletin were identified as main compounds. These compounds were also shown to inhibit the ethanol-induced reduction in antioxidant enzyme activity in an in vitro model system. These results suggest that CISE has protective effects against ethanol-induced oxidative damage, possibly by inhibition of lipid accumulation, peroxidation and increase of antioxidant defense system in the liver.     

Saponins isolated from the root of Platycodon grandiflorum protect against acute ethanol-induced hepatotoxicity in mice

The protective effects of saponins isolated from the root of Platycodon grandiflorum (Changkil saponins: CKS) against alcoholic steatosis in liver injury induced by acute ethanol administration were investigated. Pretreatment with CKS prior to ethanol administration significantly prevented the increases in serum alanine aminotransferase activity, hepatic TNF-α level, hepatic lipid peroxidation and hepatic triglyceride level. CKS prevented ethanol-induced steatosis and necrosis, as indicated by liver histopathological studies. Additionally, CKS protected against ethanol-induced depletion of hepatic glutathione levels. CYP2E1 has been suggested as a major contributor to ethanol-induced oxidative stress and liver injury. The concurrent administration of CKS efficaciously abrogated the CYP2E1 induction and CYP2E1-dependents hydroxylation of aniline as compared to the individual treatment at higher doses. These findings suggest that CKS may prevent ethanol-induced acute liver injury, possibly through its ability to block CYP2El-mediated ethanol bioactivation and its free radical scavenging effects.     

Tectoridin,an isoflavone glycoside from the flower of Pueraria lobata,prevents acute ethanol-induced liver steatosis in mice

In traditional Chinese medicine, the flower of Pueraria lobata (Puerariae Flos) has been used in therapy to counteract the problems associated with alcohol drinking and liver injury. In this study, we investigated the hepatoprotective effects and its mechanisms of tectoridin, an isoflavone glycoside from the flower of P. lobata (Willd.) Ohwi. Ethanol (5 g/kg) was given orally every 12 h for a total of three doses. 1 h after the last dose of ethanol, tectoridin (25, 50 and 100 mg/kg) was given intragastrically five times in three consecutive days. The mice were sacrificed at 4 h after tectoridin treatment. Peroxisome proliferators-activated receptor α (PPARα), sterol regulatory element-binding protein (SREBP)-1c and their target genes were evaluated by biochemical analysis and quantitative real-time polymerase chain reaction (qPCR). Mitochondria were isolated for the mitochondrial permeability transition (MPT) and membrane potential (ΔΨ_m) assay. Acute ethanol exposure resulted in the significant increase of the alanine aminotransferase (ALT), aspartate aminotransferase (AST) and triglyceride (TG) levels and hepatic mitochondria dysfunction shown as the increase of MPT and the decrease of ΔΨ_m. However, tectoridin treatment dramatically attenuated these effects. In addition, tectoridin remarkably alleviated the over-production of thiobarbituric acid-reactive substance. Furthermore, tectoridin inhibited the decrease of PPARα expression and its target genes, including medium-chain acyl-CoA dehydrogenase (MCAD), acyl-CoA oxidase (ACO) and cytochrome P450 4A (CYP 4A) at mRNA and enzyme activity levels. These data showed that tectoridin protected against ethanol-induced liver steatosis mainly through modulating the disturbance of PPARα pathway and ameliorating mitochondrial function.     

昆明种小鼠细胞色素氧化酶CYP1A基因表达的昼夜节律变化

目的探究昆明种小鼠细胞色素氧化酶1A1(CYP1A1)的昼夜节律、性别差异以及对肝毒物毒性变化的影响。方法昆明小鼠在环境控制的SPF饲养室内适应性饲养2周后,于06:00,10:00,14:00,18:00,22:00和次日02:00处死取肝,用逆转录PCR方法检测24h内核激素孤儿受体α(Rev-erbα),时钟基因(Per)1,Per2和CYP1A1,CYP1A2及其调控核受体基因AhR的表达。另取小鼠于6:00和18:00 ip给予对乙酰氨基酚500mg·kg-1,12h后检测血清中丙氨酸转氨酶(ALT)和天冬氨酸转氨酶(AST)活性。结果细胞色素氧化酶基因CYP1A1,CYP1A2及其调节基因AhR在18:00左右表达最高,且雌鼠的表达高于雄鼠,在6:00左右表达最低,表达峰谷差为4～7倍,其节律变化的差异与Rev-erbα,Per1,Per2的节律差异大致相符。与此节律相对应,对乙酰氨基酚引起的肝毒性在18:00给药高于6:00给药。结论昆明种小鼠细胞色素氧化酶CYP1A1基因表达存在昼夜节律及性别差异,该差异可影响肝毒物如对乙酰氨基酚的代谢和毒性。     

Amelioration by chicory seed extract of diabetes- and oleic acid-induced non-alcoholic fatty liver disease (NAFLD)/non-alcoholic steatohepatitis (NASH) via modulation of PPARα and SREBP-1

We evaluated the effect of chicory (Cichorium intybus L.) seed extract (CI) on hepatic steatosis caused by early and late stage diabetes in rats (in vivo), and induced in HepG2 cells (in vitro) by BSA-oleic acid complex (OA). Different dosages of CI (1.25, 2.5 and 5 mg/ml) were applied along with OA (1 mM) to HepG2 cells, simultaneously and non-simultaneously; and without OA to ordinary non-steatotic cells. Cellular lipid accumulation and glycerol release, and hepatic triglyceride (TG) content were measured. The expression levels of sterol regulatory element-binding protein-1c (SREBP-1c) and peroxisome proliferator-activated receptor alpha (PPARα) were determined. Liver samples were stained with hematoxylin and eosin (H&E). Significant histological damage (steatosis-inflammation-fibrosis) to the cells and tissues and down-regulation of SREBP-1c and PPARα genes that followed steatosis induction were prevented by CI in simultaneous treatment. In non-simultaneous treatment, CI up-regulated the expression of both genes and restored the normal levels of the corresponding proteins; with a greater stimulating effect on PPARα, CI acted as a PPARα agonist. CI released glycerol from HepG2 cells, and targeted the first and the second hit phases of hepatic steatosis. A preliminary attempt to characterize CI showed caffeic acid, chlorogenic acid, and chicoric acid, among the constituents.     

PPARβ/δ modulates ethanol-induced hepatic effects by decreasing pyridoxal kinase activity

Because of the significant morbidity and lethality caused by alcoholic liver disease (ALD), there remains a need to elucidate the regulatory mechanisms that can be targeted to prevent and treat ALD. Toward this goal, minimally invasive biomarker discovery represents an outstanding approach for these purposes. The mechanisms underlying ALD include hepatic lipid accumulation. As the peroxisome proliferator-activated receptor-β/δ (PPARβ/δ) has been shown to inhibit steatosis, the present study examined the role of PPARβ/δ in ALD coupling metabolomic, biochemical and molecular biological analyses. Wild-type and Pparβ/δ-null mice were fed either a control or 4% ethanol diet and examined after 4–7 months of treatment. Ethanol fed Pparβ/δ-null mice exhibited steatosis after short-term treatment compared to controls, the latter effect appeared to be due to increased activity of sterol regulatory element binding protein 1c (SREBP1c). The wild-type and Pparβ/δ-null mice fed the control diet showed clear differences in their urinary metabolomic profiles. In particular, metabolites associated with arginine and proline metabolism, and glycerolipid metabolism, were markedly different between genotypes suggesting a constitutive role for PPARβ/δ in the metabolism of these amino acids. Interestingly, urinary excretion of taurine was present in ethanol-fed wild-type mice but markedly lower in similarly treated Pparβ/δ-null mice. Evidence suggests that PPARβ/δ modulates pyridoxal kinase activity by altering K_m, consistent with the observed decreased in urinary taurine excretion. These data collectively suggest that PPARβ/δ prevents ethanol-induced hepatic effects by inhibiting hepatic lipogenesis, modulation of amino acid metabolism, and altering pyridoxal kinase activity.     

Role of Nrf2 in preventing ethanol-induced oxidative stress and lipid accumulation

Oxidative stress and lipid accumulation play important roles in alcohol-induced liver injury. Previous reports showed that, in livers of nuclear factor erythroid 2-related factor 2 (Nrf2)-activated mice, genes involved in antioxidant defense are induced, whereas genes involved in lipid biosynthesis are suppressed. To investigate the role of Nrf2 in ethanol-induced hepatic alterations, Nrf2-null mice, wild-type mice, kelch-like ECH-associated protein 1-knockdown (Keap1-KD) mice with enhanced Nrf2, and Keap1-hepatocyte knockout (Keap1-HKO) mice with maximum Nrf2 activation, were treated with ethanol (5 g/kg, po). Blood and liver samples were collected 6 h thereafter. Ethanol increased alanine aminotransferase and lactate dehydrogenase activities as well as thiobarbituric acid reactive substances in serum of Nrf2-null and wild-type mice, but not in Nrf2-enhanced mice. After ethanol administration, mitochondrial glutathione concentrations decreased markedly in Nrf2-null mice but not in Nrf2-enhanced mice. H₂DCFDA staining of primary hepatocytes isolated from the four genotypes of mice indicates that oxidative stress was higher in Nrf2-null cells, and lower in Nrf2-enhanced cells than in wild-type cells. Ethanol increased serum triglycerides and hepatic free fatty acids in Nrf2-null mice, and these increases were blunted in Nrf2-enhanced mice. In addition, the basal mRNA and nuclear protein levels of sterol regulatory element-binding protein 1(Srebp-1) were decreased with graded Nrf2 activation. Ethanol further induced Srebp-1 mRNA in Nrf2-null mice but not in Nrf2-enhanced mice. In conclusion, Nrf2 activation prevented alcohol-induced oxidative stress and accumulation of free fatty acids in liver by increasing genes involved in antioxidant defense and decreasing genes involved in lipogenesis.     

Chronic exposure of mice to environmental endocrine-disrupting chemicals disturbs their energy metabolism

We evaluated the effects of a 20-week chronic exposure of mice to a low dose of cypermethrin (CYP), atrazine (ATZ) and 17α-ethynyestradiol (EE2) on energy metabolism. Here, male mice were exposed to 50 μg/kg BW/day CYP, 100 μg/kg BW/day ATZ or 1 μg/kg BW/day EE2 supplied in their drinking water for 20 weeks. During the exposure, mice were fed a high energy diet (HD). The bodyweights were not significantly affected by chronic exposure to EDCs, while the serum-free fatty acids (FFA) levels, hepatic lipid accumulation and triacylglycerol (TG) contents increased significantly in the ATZ- and CYP-HD groups. To determine the mechanism involved, we determined the expression levels of the genes in the glucose and fat metabolism pathways in the liver and adipose tissue. The results showed that chronic exposure to ATZ and CYP increased the mRNA levels of a number of key genes involved in both the de novo FFA synthesis pathway and the transport of FFA from blood. The increased amount of FFA was partially consumed as energy through β-oxidation in the mitochondria. Some of the FFA was used to synthesize TG in the liver by up-regulating primary genes, which resulted in increased TG levels and lipid accumulation. The results indicate that chronic exposure to EDCs has the potential to cause energy metabolic dysregulation and hepatotoxicity in mice.     

Pyrazinamide induced hepatic injury in rats through inhibiting the PPARα pathway

Pyrazinamide (PZA) causes serious hepatotoxicity, but little is known about the exact mechanism by which PZA induced liver injury. The peroxisome proliferator‐activated receptors alpha (PPARα) is highly expressed in the liver and modulates the intracellular lipidmetabolism. So far, the role of PPARα in the hepatotoxicity of PZA is unknown. In the present study, we described the hepatotoxic effects of PZA and the role of PPARα and its target genes in the downstream pathway including L‐Fabp, Lpl, Cpt‐1b, Acaa1, Apo‐A1 and Me1 in this process. We found PZA induced the liver lipid metabolism disorder and PPARα expressionwas down‐regulated which had a significant inverse correlation with liver injury degree. These changeswere ameliorated by fenofibrate, the co‐treatment that acts as a PPARα agonist. In contrast, short‐termstarvation significantly aggravated the severity of PZA‐induced liver injury. In conclusion, this study demonstrated the critical role played by PPARα in PZA‐induced hepatotoxicity and provided a better understanding of the molecular mechanisms underlying PZA‐induced liver injury. Copyright © 2016 John Wiley & Sons, Ltd.     

Involvement of 5-HT3 and 5-HT4 Receptors in the Regulation of Circadian Clock Gene Expression in Mouse Small Intestine

Several lines of evidence suggest that 5-HT receptors play a critical role in the expression of clock genes in the suprachiasmatic nucleus, the main circadian oscillator in hamsters. The contributions of 5-HT-receptor subtypes in the intestine, where they are expressed at high concentrations, are however not yet clarified. The 5-HT synthesis inhibitor, p-chlorophenylalanine, attenuated the daily rhythm of Per1 and Per2 gene expression in the intestine. Injection of 5-HT and agonists of the 5-HT₃ and 5-HT₄ receptors increased Per1/Per2 expression and decreased Bmal1 expression in a dose-dependent manner. Although treatment with antagonists of 5-HT₃ and 5-HT₄ alone did not affect clock gene expression, co-injection of these antagonists with 5-HT blocked the 5-HT-induced changes in clock gene expression. Increased tissue levels of 5-HT due to treatment with the antidepressants clomipramine and fluvoxamine did not affect clock gene expression. The present results suggest that the 5-HT system in the small intestine may play a critical role in regulating circadian rhythms through 5-HT₃/5-HT₄-receptor activation.     

In vitro and in vivo hepatoprotective effects of the aqueous extract from Taraxacum officinale (dandelion) root against alcohol-induced oxidative stress

The protective effects of Taraxacum officinale (dandelion) root against alcoholic liver damage were investigated in HepG2/2E1 cells and ICR mice. When an increase in the production of reactive oxygen species was induced by 300 mM ethanol in vitro, cell viability was drastically decreased by 39%. However, in the presence of hot water extract (TOH) from T. officinale root, no hepatocytic damage was observed in the cells treated with ethanol, while ethanol-extract (TOE) did not show potent hepatoprotective activity. Mice, which received TOH (1 g/kg bw/day) with ethanol revealed complete prevention of alcohol-induced hepatotoxicity as evidenced by the significant reductions of serum aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and lactate dehydrogenase activities compared to ethanol-alone administered mice. When compared to the ethanol-alone treated group, the mice receiving ethanol plus TOH exhibited significant increases in hepatic antioxidant activities, including catalase, glutathione-S-transferase, glutathione peroxidase, glutathione reductase, and glutathione. Furthermore, the amelioration of malondialdehyde levels indicated TOH’s protective effects against liver damage mediated by alcohol in vivo. These results suggest that the aqueous extract of T. officinale root has protective action against alcohol-induced toxicity in the liver by elevating antioxidative potentials and decreasing lipid peroxidation.     

Effect of lacosamide on ethanol induced conditioned place preference and withdrawal associated behavior in mice: Possible contribution of hippocampal CRMP-2

BackgroundExcessive consumption of ethanol is known to activate the mTORC1 pathway and to enhance the Collapsin Response Mediator Protein-2 (CRMP-2) levels in the limbic region of brain. The latter helps in forming microtubule assembly that is linked to drug taking or addiction-like behavior in rodents. Therefore, in this study, we investigated the effect of lacosamide, an antiepileptic drug and a known CRMP-2 inhibitor, which binds to CRMP-2 and inhibits the formation of microtubule assembly, on ethanol-induced conditioned place preference (CPP) in mice.MethodsThe behavior of mice following ethanol addiction and withdrawal was assessed by performing different behavioral paradigms. Mice underwent ethanol-induced CPP training with alternate dose of ethanol (2 g/kg, po) and saline (10 ml/kg, po). The effect of lacosamide on the expression of ethanol-induced CPP and on ethanol withdrawal associated anxiety and depression-like behavior was evaluated. The effect of drug on locomotor activity was also assessed and hippocampal CRMP-2 levels were measured.ResultsEthanol-induced CPP was associated with enhanced CRMP-2 levels in the hippocampus. Lacosamide significantly reduced the expression of ethanol-induced CPP and alleviated the levels of hippocampal CRMP-2 but aggravated withdrawal-associated anxiety and depression in mice.ConclusionThe present study demonstrated the beneficial effect of lacosamide in attenuation of expression of ethanol induced conditioned place preference via reduction of hippocampal CRMP-2 level. These findings suggest that lacosamide may be investigated further for ethanol addiction but not for managing withdrawal.     

Hepatoprotective effect of Angelica archangelica in chronically ethanol-treated mice

Angelica archangelica (AAA) has been effectively used in folk medicines as a remedy against stomachal and intestinal disturbances, arthritic disease, etc. However, there is still lack of scientific proof about its antioxidant capability. This study aimed at investigating the effects of total AAA against chronic ethanol-induced hepatotoxicity. ICR mice were divided into five groups, each consisting of 10 animals. A single dose of ethanol (70%, 0.1 ml, p.o.) was used to induce hepatotoxicity in these mice which resulted in a significant elevation of the activities of serum GOT and GPT. Treatment of mice with AAA (10, 25, and 50 mg/kg p.o.) after 2 weeks ameliorated the ethanol-induced hepatotoxicity effects. Hepatotoxicity was evidenced by a significant increase in hepatic lipid peroxidation manifested as the presence of malondialdehyde. It was found that AAA inhibits the malondialdehyde formation in mouse liver homogenates both in vitro and in vivo. AAA is cytoprotective agent effective against chronic ethanol-induced hepatotoxicity, possibly through inhibition of the production of oxygen free radicals that cause lipid peroxidation, and hence indirectly protects the liver from oxidative stress.     

The herbal composition GGEx18 from Laminaria japonica,Rheum palmatum,and Ephedra sinica inhibits high-fat diet-induced hepatic steatosis via hepatic PPARα activation

Context: The activation of peroxisome proliferator-activated receptor α (PPARα) target genes promotes hepatic oxidation of fatty acids. We hypothesized that Gyeongshingangjeehwan 18 (GGEx18), a mixture of three herbs, Laminaria japonica Aresch (Laminariaceae), Rheum palmatum L. (Polygonaceae), and Ephedra sinica Stapf (Ephedraceae), can regulate high-fat diet-induced hepatic steatosis through PPARα activation in the liver.

Objective: To investigate the effects of GGEx18 on obesity-related hepatic steatosis and the responsible mechanism.

Materials and methods: The effects of GGEx18 on hepatic lipid accumulation, serum lipid profiles, and the expression of PPARα target genes were studied in high-fat diet-induced obese mice. The effects of GGEx18 on the expression of the PPARα targets and PPARα reporter gene activation were measured in NMu2Li liver cells.

Results: GGEx18 administration to obese mice for 9 weeks markedly (p?<?0.05) decreased hepatic lipid accumulation compared with that in obese control mice. Serum triglyceride and total cholesterol levels were significantly (p <0.05) decreased by GGEx18. GGEx18 treatment increased the messenger RNA levels of PPARα target genes, which are responsible for fatty acid oxidation, in liver tissues. Consistent with the in vivo data, similar activation of genes was observed in GGEx18-treated NMu2Li liver cells. GGEx18 also elevated PPARα reporter gene expression in NMu2Li cells.

Discussion and conclusion: These results suggest that GGEx18 prevents hepatic steatosis and hyperlipidemia in high-fat diet-induced obese mice, and this process may be mediated through PPARα activation in the liver.