Triglyceride Accumulation In HepG2 Cells Research Articles

Methyl Cinnamate (MC) Alleviates Free Fatty Acids (FFAs) Induced Lipid Accumulation Through the AMPK Pathway in HepG2 Cells.

AMP-activated protein kinase (AMPK) plays a critical role in energy metabolism. Its activation leads to the phosphorylation of downstream proteins such as acetyl-CoA carboxylase (ACC) and sterol regulatory element-binding protein-1 (SREBP1), subsequently inhibiting de novo fatty acid synthesis, thereby reducing intracellular triglyceride accumulation. MC is a compound found in extracts from Zanthoxylum armatum DC plants. Research has shown that MC can inhibit the differentiation of 3T3-L1 adipocytes through the CAMKK2-AMPK pathway. However, the biological effect of MC in HepG2 cells remains unknown. In this study, we utilized HepG2 cells to establish a model of MAFLD through FFAs stimulation. We investigated the biological effects of MC on HepG2 cells and studied its impact on lipid metabolism. Small interfering RNA was employed to explore the mechanism by which MC activates AMPK. Finally, molecular docking was conducted, establishing a model of the interaction between AMPK and MC. We observed that MC can alleviate triglyceride accumulation in HepG2 cells. We observed the elevated p-AMPK/AMPK, P-ACC/ ACC, and elevated CPT1a after treatment of MC in HepG2 cells. The interference of CAMKK2 mRNA did not impact the ability of MC to phosphorylate AMPK. Compound C attenuates the ability of MC to increase p-AMPK. Molecular docking results led us to hypothesize that MC directly interacts with AMPK, resulting in AMPK phosphorylation and improved lipid accumulation in HepG2 cells.

Ursane-type triterpene oligoglycosides with anti-hepatosteatosis and anti-hyperlipidemic activity from the leaves of Ilex paraguariensis A. St.-Hil.

The methanol extract from the leaves of Ilex paraguariensis A. St.-Hil. (Aquifoliaceae), popularly known as mate, maté, or yerba maté, inhibits the intracellular triglyceride accumulation in HepG2 cells and suppresses the plasma triglyceride elevation in olive oil-treated mice. Three new triterpene saponins, termed mateosides I (1), II (2), and III (3), were isolated from the extract along with 29 known compounds. The structures of 1-3 were elucidated based on chemical and spectroscopic evidence. Among the isolates, principal saponin constituents, 2 and matesaponins 1 (7) and 2 (9), potently inhibited the triglyceride accumulation in HepG2 cells simultaneously treated with oleic acid and high glucose. In vivo assay of the methanol extract of I. paraguariensis revealed that 7 and 9 showed anti-hyperlipidemic activities in olive oil-treated mice. These results suggested that the saponin constituents of I. paraguariensis could be valuable bioactive marker for the anti-obesogenic activity.

Constituents of Morus alba var. multicaulis leaf improve lipid metabolism by activating the AMPK signaling pathway in HepG2 cells.

One new compound, 3Z-1-O-β-D-glucopyranosyl-3-hexene-1,5-diol (1), together with 26 known isolates (2-27) were obtained from the leaf of Morus alba var. multicaulis. Among the known compounds, 7, 11, 12, 14, 15, 18, 19, 23, and 24 were firstly obtained from the Morus genus; 2-5, 8, 10, 13, and 20 were firstly isolated from M. alba. var. multlcaulis. Meanwhile, the NMR data of 20 and 23 have been reported here for the first time. Moreover, compounds 1-11, 13, 21, and 23-27 showed inhibitory effects on triglyceride (TG) accumulation in HepG2 cells. In mechanism, compound 1 could activate the phosphorylation of AMP-activated protein kinase α (AMPKα) to accelerate the β-oxidation of fatty acids via promoting the phosphorylation of acetyl-CoA carboxylase 1 and up-regulating carnitine palmitoyl-transferase 1A. Besides, compound 1 exerted lipolysis effect by activating hormone-sensitive lipase. In brief, compound 1 might play a role by up-regulating phosphorylation of AMPKα, enhancing the fatty acid β-oxidation and lipolysis. 27 compounds were obtained from the leaf of Morus alba var. multicaulis. Among them, 18 showed inhibitory effects on TG accumulation in HepG2 cells. Moreover, the new compound, 3Z-1-O-β-D-glucopyranosyl-3-hexene-1,5-diol (1), was found to play a role by up-regulating phosphorylation of AMPKα, enhancing the fatty acids β-oxidation and lipolysis.

Resveratrol alleviates non-alcoholic fatty liver disease through epigenetic modification of the Nrf2 signaling pathway.

Recent findings have demonstrated the aberrant DNA methylation of the Nrf2-Keap1 genes in human cancers; however, the epigenetic control of this pathway in non-alcoholic fatty liver disease (NAFLD) is unknown. Resveratrol can modify epigenetic mechanisms. Our objectives in this study were to explore the correlation between promoter methylation of the Nrf2-Keap1 genes and NAFLD, and that investigate the effect of resveratrol on the epigenetic regulation Nrf2-Keap1 in vitro and in vivo models of NAFLD. Resveratrol attenuated high fat-diet (HFD)-induced methylation of the Nrf2 promoter in the liver of mice, and this effect was correlated with reduction in triglyceride level and decrease in the expression of lipogenesis-related genes such as FAS and SREBP-1c. In addition, treatment of HepG2 cells with high glucose (HG) enhanced methylation level of the Nrf2 promoter, whereas resveratrol reversed this effect. Treatment of the cells with resveratrol or 5-aza, a demethylating agent, could prevent HG-induced reactive oxygen species production and expression of Nrf2-controlled antioxidant genes. Moreover, resveratrol or 5-aza could significantly attenuate HG-induced triglyceride accumulation in HepG2 cells. These findings indicate that resveratrol attenuates NAFLD through the epigenetic modification the Nrf2 signaling.

Inhibition Effect of Triglyceride Accumulation by Large Yellow Croaker Roe DHA-PC in HepG2 Cells.

The phospholipids (PLs) of large yellow croaker (Pseudosciaena crocea, P. crocea) roe contain a high level of polyunsaturated fatty acids, especially docosahexaenoic acid (DHA), which can lower blood lipid levels. In previous research, PLs of P. crocea roe were found able to regulate the accumulation of triglycerides. However, none of these involve the function of DHA-containing phosphatidylcholine (DHA-PC), which is the main component of PLs derived from P. crocea roe. The function by which DHA-PC from P. crocea roe exerts its effects has not yet been clarified. Herein, we used purified DHA-PC and oleic acid (OA) induced HepG2 cells to establish a high-fat model, and the cell activity and intracellular lipid levels were then measured. The mRNA and protein expression of Fatty Acid Synthase (FAS), Carnitine Palmitoyl Transferase 1A (CPT1A) and Peroxisome Proliferator-Activated Receptor α (PPARα) in HepG2 cells were detected via RT-qPCR and western blot as well. It was found that DHA-PC can significantly regulate triglyceride accumulation in HepG2 cells, the effect of which was related to the activation of PPARα receptor activity, upregulation of CPT1A, and downregulation of FAS expression. These results can improve the understanding of the biofunction of hyperlipidemia mediated by DHA-PC from P. crocea roe, as well as provide a theoretical basis for the utilization of DHA-PC from P. crocea roe as a functional food additive.

Arazyme Suppresses Hepatic Steatosis and Steatohepatitis in Diet-Induced Non-Alcoholic Fatty Liver Disease-Like Mouse Model.

Arazyme, a metalloprotease from the spider Nephila clavata, exerts hepatoprotective activity in CCL4-induced acute hepatic injury. This study investigated the hepatoprotective effects in high-fat diet (HFD)-induced non-alcoholic fatty liver disease-like C57BL/6J mice. The mice were randomly divided into four groups (n = 10/group): the normal diet group, the HFD group, the arazyme group (HFD with 0.025% arazyme), and the milk thistle (MT) group (HFD with 0.1% MT). Dietary supplementation of arazyme for 13 weeks significantly lowered plasma triglyceride (TG) and non-esterified fatty acid levels. Suppression of HFD-induced hepatic steatosis in the arazyme group was caused by the reduced hepatic TG and total cholesterol (TC) contents. Arazyme supplementation decreased hepatic lipogenesis-related gene expression, sterol regulatory element-binding transcription protein 1 (Srebf1), fatty acid synthase (Fas), acetyl-CoA carboxylase 1 (Acc1), stearoyl-CoA desaturase-1 (Scd1), Scd2, glycerol-3-phosphate acyltransferase (Gpam), diacylglycerol O-acyltransferase 1 (Dgat1), and Dgat2. Arazyme directly reduced palmitic acid (PA)-induced TG accumulation in HepG2 cells. Arazyme suppressed macrophage infiltration and tumor necrosis factor α (Tnfa), interleukin-1β (Il1b), and chemokine-ligand-2 (Ccl2) expression in the liver, and inhibited secretion of TNFα and expression of inflammatory mediators, Tnfa, Il1b, Ccl2, Ccl3, Ccl4, and Ccl5, in PA-induced RAW264.7 cells. Arazyme effectively protected hepatic steatosis and steatohepatitis by inhibiting SREBP-1-mediated lipid accumulation and macrophage-mediated inflammation.

Metformin attenuates triglyceride accumulation in HepG2 cells through decreasing stearyl-coenzyme A desaturase 1 expression

BackgroundThe prevalence of nonalcoholic fatty liver disease (NAFLD) has increased worldwide. Metformin decreases triglyceride (TG) accumulation in hepatocytes in vivo and in vitro. Stearyl-coenzyme A desaturase 1 (SCD1) knockout mice also show decreased liver TG accumulation; however, whether SCD1 plays a role in the effect of metformin on TG accumulation is unknown. Therefore, the aim of this study was to investigate whether SCD1 mediated the effect of metformin on TG accumulation.MethodsHepG2 and AML12 cells were exposed to high glucose and high insulin with or without metformin. An adenovirus was used for the SCD1 knockdown and overexpression. The triglyceride level in cells was detected. The expression of related genes was detected by Western blot and quantitative real-time PCR. A dual-luciferase reporter assay was used to determine the effect of metformin on the transcriptional activity of the SCD1 promoter.ResultsMetformin decreased TG accumulation to normal level in HepG2 cells exposed to high glucose and high insulin. The expression of SCD1 and fatty acid synthetase (FAS) was also decreased to normal level by metformin. Knockdown of SCD1 mimicked the effect of metformin on decreasing TG levels in AML12 cells, and the overexpression of SCD1 attenuated the effect of metformin on decreasing TG accumulation in HepG2 cells. The dual-luciferase reporter assay showed that the transcriptional activity of the SCD1 promoter (− 550/+ 199) after metformin treatment was 2-fold lower compared to control group in HepG2 cells. Additionally, the phosphorylation of AMPK after metformin treatment was 2-fold higher, and the expression of sterol regulatory element-binding protein-1c (SREBP-1c) after metformin treatment was about 2-fold lower compared to high glucose and high insulin group in HepG2 cells.ConclusionsTogether, these results reveal that metformin reduces TG accumulation in HepG2 cells via inhibiting the expression of SCD1.

Epigallocatechin-3-Gallate-Rich Green Tea Extract Ameliorates Fatty Liver and Weight Gain in Mice Fed a High Fat Diet by Activating the Sirtuin 1 and AMP Activating Protein Kinase Pathway.

The prevalence of metabolic diseases has risen globally in parallel with the obesity epidemic over the past few decades. Green tea has been reported to have metabolically beneficial effects on obesity; however, the mechanism by which green tea regulates lipid metabolism is not clearly understood. Male c57BL/6 mice were fed a normal chow diet, a high-fat diet (HFD), or an HFD supplemented with various doses of epigallocatechin gallate-rich green tea extract (GTE) for 12 weeks. GTE supplementation reduced body weight gain, prevented hepatic fat accumulation, decreased hypertriglyceridemia, and improved hyperglycemia and insulin resistance in HFD-fed mice. The underlying mechanisms of these beneficial effects of GTE might involve the upregulation of sirtuin 1 and AMP activated protein kinase (AMPK) and the downregulation of enzymes related to de novo lipogenesis. Consistent with the in vivo findings, GTE increased the expression and activity of sirtuin 1, enhanced the binding of sirtuin 1 to liver kinase B1 (LKB1) and subsequent deacetylation of LKB1, and reduced triglyceride accumulation in HepG2 cells. These results suggest the possible therapeutic potential of dietary epigallocatechin gallate-rich GTE supplementation for preventing the development and progression of hepatic steatosis and obesity.

Citreoviridin induces triglyceride accumulation in hepatocytes through inhibiting PPAR-α in vivo and in vitro

Citreoviridin (CIT) is a mycotoxin produced by Penicillum citreonigrum, Aspergillus terreus and Eupenicillium ochrosalmoneum. CIT occurs naturally in moldy rice and corn. CIT is associated with the development of atherosclerosis in the general population. Alteration in hepatic lipid metabolism is a pathogenic factor in atherosclerosis. However the effect and the underlying mechanism of CIT on hepatic lipid metabolism are largely unknown. In this study, we reported that CIT induced triglyceride accumulation in mice liver and human liver HepG2 cells as shown in oil red O staining. CIT (0.1 mg/kg–0.3 mg/kg) for 6 weeks elevated liver triglyceride contents in mice. CIT inhibited the transactivation activity of peroxisome proliferator-activated receptor-α (PPAR-α) in hepatocyte in vivo and in vitro, as shown by the reduced mRNA levels of PPAR-α target genes which play key roles in lipid metabolism in various aspects. PPAR-α agonist fenofibrate attenuated CIT-induced triglyceride accumulation in HepG2 cells. Furthermore, CIT increased serum total cholesterol/high-density lipoprotein cholesterol ratio, a strong risk factor for cardiovascular disease. In summary, we reported that CIT induced PPAR-α−dependent hepatic triglyceride accumulation and dyslipidemia. Our data will provide new mechanistic insights into CIT-induced lipid alterations.

Up-regulated HOTAIR induced by fatty acids inhibits PTEN expression and increases triglycerides accumulation in HepG2 cells

ABSTRACTNon-alcoholic fatty liver disease (NAFLD) is characterized by hepatic lipid accumulation unrelated to excess alcohol intake, in which the hepatic expression of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is inhibited. Long non-coding RNA HOTAIR suppresses PTEN expression in hepatic stellate cells during liver fibrosis, and it is involved in liver lipid dysregulation. In this study, we evaluated whether the PTEN down-regulation and lipid accumulation in hepatic cells might be mediated by HOTAIR during the development of NAFLD. Free fatty acids (FFAs) treatment promoted triglyceride accumulation in HepG2 cells, significantly increasing HOTAIR expression and inhibiting PTEN expression (both at mRNA and protein levels).The effects on HOTAIR and PTEN expressions disappeared after withdrawal of the FFAs treatment. SiRNA-mediated HOTAIR knockdown prevented PTEN down-regulation and triglyceride accumulation in HepG2 cells treated with FFAs; and CAPE (an NF-κBp65 inhibitor) treatment prevented the HOTAIR up-regulation and PTEN down-regulation. FFAs could induce the up-regulation of HOTAIR in HepG2 cells probably dependent on the NF-κB signaling, and consequently suppress PTEN expression and promote triglyceride accumulation. Aberrant up-regulation of HOTAIR mediated by excessive circulating FFAs levels may be a crucial mechanism associated with liver steatosis.

Glial cell line–derived neurotrophic factor–induced mice liver defatting: A novel strategy to enable transplantation of steatotic livers

Moderate macrovesicular steatosis (>30%), which is present in almost 50% of livers considered for transplantation, increases the risk of primary graft dysfunction. Our previously published data showed that glial cell line-derived neurotrophic factor (GDNF) is protective against high-fat diet (HFD)-induced hepatic steatosis in mice. Hence, we hypothesized that perfusion of steatotic livers with GDNF may reduce liver fat content before transplantation. Livers from 8 weeks of regular diet (RD) and of HFD-fed mice were perfused ex vivo for 4 hours with either vehicle, GDNF, or a previously described defatting cocktail. The liver's residual fat was quantified colorimetrically using a triglyceride (TG) assay kit and by Oil Red O (ORO) and Nile red/Hoechst staining. Liver tissue injury was assessed by using a lactate dehydrogenase (LDH) activity assay. In vitro induction of lipolysis in HepG2 cells was assessed by measuring glycerol and free fatty acid release. ORO staining showed significantly more steatosis in livers from HFD-fed mice compared with RD-fed mice (P < 0.001). HFD livers perfused with GDNF had significantly less steatosis than those not perfused (P = 0.001) or perfused with vehicle (P < 0.05). GDNF is equally effective in steatotic liver defatting compared to the defatting cocktail; however, GDNF induces less liver damage than the defatting cocktail. These observations were consistent with data obtained from assessment of liver TG content. Assessment of liver injury revealed significant hepatocyte injury in livers perfused with the control defatting cocktail but no evidence of injury in livers perfused with either GDNF or vehicle. In vitro, GDNF reduced TG accumulation in HepG2 cells and stimulated increased TG lipolysis. In conclusion, GDNF can decrease mice liver fat content to an acceptable range and could be a potential defatting agent before liver transplantation.

Bioactive cyclolanstane-type saponins from the stems of Astragalus membranaceus (Fisch.) Bge. var. mongholicus (Bge.) Hsiao.

Twelve cyclolanstane-type saponins including six new ones, astrolanosaponins A1 (1), A2 (2), B (3), C (4), D (5), and E (6) were obtained from the stem of Astragalus membranaceus (Fisch.) Bge. var. mongholicus (Bge.) Hsiao, and their structures were elucidated by chemical and spectroscopic methods. Of the known ones, cycloastragenol-3-O-β-D-glucopyranoside (7), astraverrucin II (8), cycloaraloside E (9), huangqiyenin A (10), and huangqiyenin B (11) were isolated from the species first. Meanwhile, compounds 1-3, 5-9, and aleksandroside I (12) showed inhibitory effects on triglyceride accumulation in HepG2 cells.

9,10-Dihydrophenanthrene derivatives and one 1,4-anthraquinone firstly isolated from Dioscorea zingiberensis C. H. Wright and their biological activities

Two new phenanthrene derivatives, 2,5,7-trimethoxy-9,10-dihydrophenanthrene-1,4-dione (1) and 2,5,6-trihydroxy-3,4-dimethoxy-9,10-dihydrophenanthrene (3), one new anthracenedione, 2,5,7-trimethoxyanthracene-1,4-dione (2), together with two known 9,10-dihydrophenanthrenes (4–5) were isolated from the rhizomes of Dioscorea zingiberensis C. H. Wright. The structures of these new compounds were established based on extensive NMR spectroscopy. Several isolated compounds were evaluated for the inhibition against nitric oxide (NO) production in the lipopolysaccharide (LPS)-activated RAW 264.7 macrophage cell line, DPPH radical scavenging, and inhibitory activity on Free Fatty Acids (FFAs) induced triglyceride accumulation in HepG2 cells. Compound 2 exhibited moderate anti-inflammatory activity, compound 3 possessed comparable DPPH radical scavenging activity as Vitamin C, compounds 2 and 4 showed potent inhibitory activities on triglyceride accumulation.

Anthocyanin-rich extracts from blackberry, wild blueberry, strawberry, and chokeberry: antioxidant activity and inhibitory effect on oleic acid-induced hepatic steatosis in vitro.

Limited information is available regarding the relationship between the chemical structures and inhibitory effects of anthocyanin (ACN) on triglyceride (TG) overaccumulation. Thus this study investigated the antioxidant activity and inhibitory effect of blackberry, wild blueberry, strawberry, and chokeberry ACN-rich extracts, with different structural characteristics, on oleic acid-induced hepatic steatosis in vitro. Four major ACNs from these berries, with different aglycones, namely cyanidin-3-glucoside (Cy-3-glu), delphinidin-3-glucoside, pelargonidin-3-glucoside, and malvidin-3-glucoside, were also investigated. Blackberry ACN-rich extract exhibited the most significant inhibitory effect on TG clearance (30.5% ± 3.4%) and reactive oxygen species generation. TG clearance was significantly correlated with total phenolic content (r = 0.991, P < 0.05) and oxygen radical absorbance capacity value (r = 0.961, P < 0.05). Furthermore, Cy-3-glu showed the highest inhibitory effect on intracellular TG overaccumulation, with a maximum TG clearance of 61.3% at 40 µg mL(-1) . Our findings suggest that the inhibitory effects of different ACNs on oleic acid-induced hepatic steatosis significantly vary. Cy-3-glu, which contains the ortho hydroxyl group in its B ring, possibly confers the protective effects of antioxidants and inhibits TG accumulation in HepG2 cells. © 2015 Society of Chemical Industry.

Flavonoid ingredients of Ginkgo biloba leaf extract regulate lipid metabolism through Sp1-mediated carnitine palmitoyltranferase 1A up-regulation.

BackgroundLipid accumulation is the primary evidence of non-alcoholic fatty liver disease (NAFLD). Ginkgo biloba extract (GBE) and its flavonoid ingredients (quercetin, kaempferol, and isorhamnetin) could lessen the lipid accumulation associated with up-regulation of the rate-limiting enzyme, carnitine palmitoyltransferase 1A (CPT1A), in the β-oxidation of long-chain fatty acids. In this study, we investigated the mechanisms by which GBE and its flavonoids induced expression of CPT1A.ResultsCPT1A inhibition with RNAi resulted in triglyceride accumulation in HepG2 cells. Through deletion and mutation analysis of CPT1A’s promoter combined with electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) experiments, the CPT1A promoter region (−50 to −5 nt) was determined to contain two putative Sp1 binding sites, namely Sp1a and Sp1b, which might act as the GBE regulation response DNA element. Sp1 might be induced to transfer from cytoplasma to nucleus to bind the promoter region of −50 to −5 nt by GBE. The regulatory effects of GBE on CPT1A were also verified on the flavonoid ingredients quercetin, kaempferol, and isorhamnetin.ConclusionSp1 was crucial in regulating CPT1A expression with GBE and its flavonoid ingredients, and the −50 to −5 nt region of CPT1A promoter played important roles in Sp1 binding.Electronic supplementary materialThe online version of this article (doi:10.1186/s12929-014-0087-x) contains supplementary material, which is available to authorized users.

Fucosterol, a liver X receptors agonist, stimulates RCT and regulates the expression of key genes in cholesterol homeostasis in vitro

We investigated whether fucosterol,sterol riches in marine algae, could activate liver X receptors (LXRs) activity, the critical transcription factors in the reverse cholesterol transport. Fucosterol dose‐dependently stimulated the transcriptional activity of both LXR‐α and LXR‐β in a reporter gene assay, responses that were attenuated by the LXR antagonist As2O3. Fucosterol also activated coactivator recruitment in cell‐free time‐resolved fluorescence resonance energy transfer analysis. In THP‐1‐derived macrophages, it induced the transcriptional activation of ABCA1, ABCG1, and apoE, key genes in reverse cholesterol transport, and thereby significantly increased the efflux of cholesterol. Fucosterol also regulated intestinal NPC1L1 and ABCA1 in Caco‐2 cells. Notably, fucosterol did not induce cellular triglyceride accumulation in HepG2 cells, primarily due to its upregulation of Insig‐2a, which delays nuclear translocation of SREBP‐1c, a key hepatic lipogenic transcription factor. These results suggest that fucosterol is a dual‐LXR agonist that regulates the expression of key genes in cholesterol homeostasis in multiple cell lines without inducing hepatic triglyceride accumulation.

Licochalcone A regulates hepatic lipid metabolism through activation of AMP-activated protein kinase

Licochalcone A (LA) is a major phenolic ingredient of Glycyrrhiza plant. Although multiple pharmacological activities of LA have been reported, effect on hepatic lipid metabolism is unknown yet. The present study showed LA to suppress the hepatic triglyceride accumulation in HepG2 cells and ICR mice fed on a high fat diet (HFD). LA inhibited lipogenesis via suppression of sterol regulatory element-binding protein 1c (SREBP1c) and its target enzymes (stearoyl-CoA desaturase 1, fatty acid synthase and glycerol-3-phosphate acyltransferase) transcription. In addition, LA up-regulated gene expression of proteins such as peroxisome proliferator-activated receptor α (PPARα) and fatty acid transporter (FAT/CD36), which are responsible for lipolysis and fatty acid transport, respectively. These effects were mediated through activation of AMP-activated protein kinase (AMPK), and were abrogated when HepG2 cells were treated with an AMPK inhibitor, compound C. To explore how LA activates AMPK, oxygen consumption rate and ATP levels were measured in HepG2 cells. LA significantly inhibited the mitochondrial respiration and ATP levels, suggesting that LA activated AMPK indirectly. In animal study, LA (5 and 10mg/kg) was orally administered to six-week-old mice once a day for 3weeks. In vitro results were likely to hold true in vivo experiment, as LA markedly lowered the triglyceride levels and activated AMPK signaling pathway in the liver of ICR mice fed on a HFD. In conclusion, the current study suggests that LA suppressed hepatic triglyceride accumulation through modulation of AMPK-SREBP signaling pathway and thus LA may be a potential therapeutic agent for treating fatty liver disease.

Betulinic acid alleviates non-alcoholic fatty liver by inhibiting SREBP1 activity via the AMPK–mTOR–SREBP signaling pathway

Non-alcoholic fatty liver disease (NAFLD) is emerging as the most common liver disease in industrialized countries. The discovery of food components that can ameliorate NAFLD is therefore of interest. Betulinic acid (BA) is a triterpenoid with many pharmacological activities, but the effect of BA on fatty liver is as yet unknown. To explore the possible anti-fatty liver effects and their underlying mechanisms, we used insulin-resistant HepG2 cells, primary rat hepatocytes and liver tissue from ICR mice fed a high-fat diet (HFD). Oil Red O staining revealed that BA significantly suppressed excessive triglyceride accumulation in HepG2 cells and in the livers of mice fed a HFD. Ca+2-calmodulin dependent protein kinase kinase (CAMKK) and AMP-activated protein kinase (AMPK) were both activated by BA treatment. In contrast, the protein levels of sterol regulatory element-binding protein 1 (SREBP1), mammalian target of rapamycin (mTOR) and S6 kinase (S6K) were all reduced when hepatocytes were treated with BA for up to 24h. We found that BA activates AMPK via phosphorylation, suppresses SREBP1 mRNA expression, nuclear translocation and repressed SREBP1 target gene expression in HepG2 cells and primary hepatocytes, leading to reduced lipogenesis and lipid accumulation. These effects were completely abolished in the presence of STO-609 (a CAMKK inhibitor) or compound C (an AMPK inhibitor), indicating that the BA-induced reduction in hepatic steatosis was mediated via the CAMKK–AMPK–SREBP1 signaling pathway. Taken together, our results suggest that BA effectively ameliorates intracellular lipid accumulation in liver cells and thus is a potential therapeutic agent for the prevention of fatty liver disease.

Fucosterol Is a Selective Liver X Receptor Modulator That Regulates the Expression of Key Genes in Cholesterol Homeostasis in Macrophages, Hepatocytes, and Intestinal Cells

Fucosterol, a sterol that is abundant in marine algae, has hypocholesterolemic activity, but the mechanism underlying its effect is not clearly understood. Because data suggest that fucosterol can increase plasma high-density lipoprotein concentrations, we investigated whether it could activate liver X receptors (LXRs), critical transcription factors in reverse cholesterol transport. Fucosterol dose-dependently stimulated the transcriptional activity of both LXR-α and -β in a reporter gene assay, responses that were attenuated by the LXR antagonist As(2)O(3). Fucosterol also activated co-activator recruitment in cell-free time-resolved fluorescence resonance energy transfer analysis. In THP-1-derived macrophages, it induced the transcriptional activation of ABCA1, ABCG1, and ApoE, key genes in reverse cholesterol transport, and thereby significantly increased the efflux of cholesterol. Fucosterol also regulated intestinal NPC1L1 and ABCA1 in Caco-2 cells. Notably, fucosterol did not induce cellular triglyceride accumulation in HepG2 cells, primarily because of its upregulation of Insig-2a, which delays nuclear translocation of SREBP-1c, a key hepatic lipogenic transcription factor. These results suggest that fucosterol is a dual-LXR agonist that regulates the expression of key genes in cholesterol homeostasis in multiple cell lines without inducing hepatic triglyceride accumulation.

Cassia tora (Leguminosae) seed extract alleviates high-fat diet-induced nonalcoholic fatty liver

The aim of this study was to examine the effects of Cassia tora seeds on high-fat diet (HFD)-induced hepatic steatosis, and elucidate the molecular mechanisms behind its effects. After being fed a HFD for two weeks, rats were orally dosed with Cassia seed ethanol extract (CSEE) (100, 200, or 300mg/kg) once daily for 8weeks. CSEE induced dose-dependent reductions in plasma lipid levels, as well as decreased the over hepatic lipid accumulation. Furthermore, CSEE treatment improved HFD-induced hepatic histological lesions. CSEE enhanced the phosphorylation of AMP-activated protein kinase (AMPK) and its primary downstream targeting enzyme, acetyl-CoA carboxylase, up-regulated the gene expression of carnitine palmitoyl transferase 1, and down-regulated sterol regulatory element binding protein 1 and fatty acid synthase protein levels in the livers of HFD-fed rats. AMPK inhibition by compound C retarded CSEE-induced reduction in triglyceride accumulation in HepG2 cells stimulated by insulin. Our findings suggest that CSEE may regulate hepatic lipid homeostasis related with an AMPK-dependent signaling pathway. Targeting AMPK activation with CSEE may represent a promising approach for the prevention and treatment of obesity-related non-alcoholic fatty liver disease.