Lipid Deposition In Liver Research Articles

Radix Polygoni Multiflori and Its Main Component Emodin Attenuate Non-Alcoholic Fatty Liver Disease in Zebrafish by Regulation of AMPK Signaling Pathway.

PurposeNonalcoholic fatty liver disease (NAFLD) has become a predictor of death in many diseases. This study was carried out to investigate the therapeutic effect of Radix Polygoni Multiflori Preparata (RPMP) and its main component emodin on egg yolk powder-induced NAFLD in zebrafish. Further investigation was performed to explore whether emodin was the main component of RPMP for the treatment of NAFLD as well as the underlying therapeutic mechanism of RPMP and emodin.MethodsZebrafish were divided into control group, egg yolk powder group, RPMP group and emodin group. The obesity of zebrafish was evaluated by body weight, body length and BMI. The content of lipid was detected by triglyceride (TG), total cholesterol (TC) reagent kit and the fatty acid was detected by nonesterified free fatty acids (NEFA) reagent kit. HE staining was used to detect the histological structure of liver. Whole-mount Oil red O staining and Frozen oil red O staining were carried out to investigate the lipid accumulation in liver. KEGG and STRING databases were performed to analyze the potential role of AMPK between insulin resistance (IR) and fatty acid oxidation. Western blot and RT-qPCR were carried out for mechanism research.ResultsRPMP and emodin significantly reduced zebrafish weight, body length and BMI. Both RPMP and emodin treatment could reduce the lipid deposition in zebrafish liver. RPMP significantly reduced the content of TG. However, emodin significantly reduced the contents of TG, TC and NEFA in zebrafish with NAFLD. The protein interaction network indicated that AMPK participated in both IR and fatty acid oxidation. Further investigation indicated that RPMP and emodin reduced hepatic lipogenesis via up-regulating the expressions of phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT2), amp-activated protein kinase alpha (AMPKα), proliferator-activated receptor alpha (PPARα), carnitine palmitoyl transferase 1a (CPT-1a) and acyl-coenzyme A oxidase 1 (ACOX1).ConclusionThese findings suggest that emodin is the main component of RPMP for the treatment of NAFLD, which is closely related to the regulation of AMPK signaling pathway which increases IR and fatty acid oxidation.

Aggravated Endothelial Inflammasome Activation and Neointima Proliferation by Hepatic Steotosis‐intensified Exosome Release in Liver‐Specific Asah1 Gene Knockout Mice

Nonalcoholic fatty liver disease (NAFLD) has been reported to be independently associated with increased traditional risk for cardiovascular diseases (CVDs). However, the mechanism underpinning the association between NAFLD and CVD remains poorly understood. The present study hypothesized that circulatory exosomes released from liver during NAFLD contribute to vascular endothelial inflammasome activation and neointima proliferation. In mice with liver‐specific deletion of Asah1 gene, a gene coding mouse acid ceramidase (Asah1fl/fl/Albcre), we found significantly enhanced liver lipid deposition when they were fed the high fat diet (HFD), as shown by HE staining and Oil Red Staining. These mice also had augmented expression of exosome marker, CD63 in the liver, and exosomes in their plasma was significantly increased as measured by Nanoparticle Tracking Analysis (NTA) compared to WT/WT mice. In partial ligated carotid artery, we observed that Asah1fl/fl/Albcre mice with HFD treatment markedly increased Nod‐like receptor pyrin domain 3 (NLRP3) inflammasome formation as shown by the colacolization of NLRP3 with apoptosis‐associated speck‐like protein gene (ASC) or caspase‐1. Enhanced colocaziation of fluorescent labeled inhibitor of caspases (FLICA) with endothelial cell marker, vWF and increased IL‐1β level in the carotid intima demonstrate NLRP3 inflammasome activation. Correspondingly, more severe vascular endothelial injury was found in Asah1fl/fl/Albcre mice as shown by reduction of vWF and endothelial junction protein, ZO‐1, which was accompaned by increased neointima proliferation. These results suggest that acid ceramidase importantly controls exosome release from the liver and its decifiency intensifies NAFLD and thereby releasees more exosomes to promote endothelial NLRP3 inflammasome activation and consequent neointiam proliferation.Support or Funding InformationSupported by HL057244, HL075316

N1-Methylnicotinamide Improves Hepatic Insulin Sensitivity via Activation of SIRT1 and Inhibition of FOXO1 Acetylation.

Objective To explore the effects of N1-methylnicotinamide (MNAM) on insulin resistance and glucose metabolism in obese type 2 diabetes mellitus (T2DM) mice and regulatory mechanisms of the NAD-dependent deacetylase sirtuin-1 (SIRT1)/forkhead box protein O1 (FOXO1) pathway. Methods Blood glucose and insulin levels were examined in mice. HE and oil red O staining were used to observe the effects of MNAM on liver lipid deposition in ob/ob mice. Real-time PCR and Western blotting were used to detect expression of gluconeogenesis, insulin signaling-related proteins, and SIRT1/FOXO1 pathway-related proteins. L-O2 cells were cultured as a model of insulin resistance, and MNAM and SIRT1 inhibitors were administered in vivo. Residual glucose and insulin signaling-related proteins were detected and the mechanisms associated with the SIRT1/FOXO1 signaling pathway in insulin resistance explored. Results MNAM can effectively reduce levels of fasting blood glucose and insulin, improve liver morphology, and reduce lipid accumulation in obese type 2 diabetes mellitus mice. MNAM also downregulates the key proteins in the gluconeogenesis pathway in the liver, upregulates Sirt1 expression, and reduces acetylation of the FOXO1 protein. In vitro, MNAM could promote the glucose uptake capacity of L-O2 cells induced by palmitic acid (PA), a saturated fatty acid that induces IR in various scenarios, including hepatocytes, improving insulin resistance. As Sirt1 expression was inhibited, the reduction of hepatocyte gluconeogenesis and the regulation of the insulin signaling pathway by MNAM were reversed. Conclusion MNAM activates SIRT1 and inhibits acetylation of FOXO1, which in turn regulates insulin sensitivity in type 2 diabetic mice, leading to a reduction of hepatic glucose output and improvement of insulin resistance.

Maternal Fat-1 Transgene Protects Offspring from Excess Weight Gain, Oxidative Stress, and Reduced Fatty Acid Oxidation in Response to High-Fat Diet.

Overweight and obesity accompanies up to 70% of pregnancies and is a strong risk factor for offspring metabolic disease. Maternal obesity-associated inflammation and lipid profile are hypothesized as important contributors to excess offspring liver and skeletal muscle lipid deposition and oxidative stress. Here, we tested whether dams expressing the fat-1 transgene, which endogenously converts omega-6 (n-6) to omega-3 (n-3) polyunsaturated fatty acid, could protect wild-type (WT) offspring against high-fat diet induced weight gain, oxidative stress, and disrupted mitochondrial fatty acid oxidation. Despite similar body mass at weaning, offspring from fat-1 high-fat-fed dams gained less weight compared with offspring from WT high-fat-fed dams. In particular, WT males from fat-1 high-fat-fed dams were protected from post-weaning high-fat diet induced weight gain, reduced fatty acid oxidation, or excess oxidative stress compared with offspring of WT high-fat-fed dams. Adult offspring of WT high-fat-fed dams exhibited greater skeletal muscle triglycerides and reduced skeletal muscle antioxidant defense and redox balance compared with offspring of WT dams on control diet. Fat-1 offspring were protected from the reduced fatty acid oxidation and excess oxidative stress observed in offspring of WT high-fat-fed dams. These results indicate that a maternal fat-1 transgene has protective effects against offspring liver and skeletal muscle lipotoxicity resulting from a maternal high-fat diet, particularly in males. Altering maternal fatty acid composition, without changing maternal dietary composition or weight gain with high-fat feeding, may highlight important strategies for n-3-based prevention of developmental programming of obesity and its complications.

Rhodiola crenulata root extract ameliorates fructose-induced hepatic steatosis in rats: Association with activating autophagy

BackgroundIncreasing evidence has shown the beneficial effects of Rhodiola species on metabolic disorders, but their mechanisms are not clear. Hepatic steatosis is closely related to metabolic disorders, we aim to investigate the therapeutic effects of Rhodiola crenulata root (RCR) on fructose-induced hepatic steatosis and explore the underlying mechanisms. PurposeTo observe the effect of Rhodiola crenulata root extract (RCR) on fructose-induced hepatic steatosis in Sprague-Dawley (SD) rats and explore its possible mechanism. MethodsMale Sprague-Dawley rats were treated with liquid fructose in their drinking water over 18 weeks. The extract of RCR was co-administered (once daily by oral gavage) during the last 5 weeks. Liver lipid deposition and morphological changes were observed by Oil red O staining. Real-time fluorescence quantitative PCR, Western blot and immunoprecipitation were used to detect gene and protein expression in liver. ResultsRCR (50 mg/kg) reversed liquid fructose-induced increase in hepatic triglyceride content in rats. Attenuation of the increased vacuolization and Oil Red O staining area was evident on histological examination of liver in RCR-treated rats. However, RCR treatment did not affect chow intake and body weight of rats. Although some genes of the pathways involved in DNL (ChREBP, SREBP-1c, FAS, ACC1, SCD1, DGAT1, DGAT2 and MGAT2), fatty acid β-oxidation (PPARα, CPT1a, ACO and FGF21), VLDL-export (MTTP) and decomposition (HSL, ATGL) in the liver of fructose-fed rats were not changed significantly after RCR administration, the decrease in PPARα and PGC-1α proteins was reversed by RCR. Notably, SIRT1 mRNA and protein expression increased significantly with RCR administration. Furthermore, RCR increased expression of ATG4B, Beclin1 and decreased expression of Bcl2-Beclin1 complex dramatically. Meanwhile, RCR decreased the acetylation of beclin1. Moreover, RCR increased expression of autophagosome markers including LC3B and ATG5-ATG12-ATG16L1, and decreased expression of autophagolysosome marker p62 in the livers of fructose-fed rats. ConclusionsRCR has a certain improvement effect on fructose-induced hepatic steatosis, which is related to the activation of autophagy.

Metabolomics of the Protective Effect of Ampelopsis grossedentata and Its Major Active Compound Dihydromyricetin on the Liver of High‐Fat Diet Hamster

The flavonoid dihydromyricetin (DMY) is the main component of Ampelopsis grossedentata (Hand-Mazz) W. T. Wang (AG), a daily beverage and folk medicine used in Southern China to treat jaundice hepatitis, cold fever, and sore throat. Recently, DMY and AG were shown to have a beneficial effect on lipid metabolism disorder. However, the mechanisms of how DMY and AG protect the liver during lipid metabolism disorder remain unclear. In this study, we first analyzed the chemical compounds of AG by HPLC-DAD-ESI-IT-TOF-MSn. Of the 31 compounds detected, 29 were identified based on previous results. Then, the effects of DMY and AG on high-fat diet hamster livers were studied and the metabolite levels and metabolic pathway activity of the liver were explored by 1H NMR metabolomics. Compared to the high-fat diet group, supplementation of AG and DMY attenuated the high-fat-induced increase in body weight, liver lipid deposition, serum triglycerides and total cholesterol levels, and normalized endogenous metabolite concentrations. PCA and PLS-DA score plots demonstrated that while the metabolic profiles of hamsters fed a high-fat diet supplemented with DMY or AG were both far from those of hamsters fed a normal diet or a high-fat diet alone, they were similar to each other. Our data suggest that the underlying mechanism of the protective effect of DMY and AG might be related to an attenuation of the deleterious effect of high-fat diet-induced hyperlipidemia on multiple metabolic pathways including amino acid metabolism, ketone body metabolism, energy metabolism, tricarboxylic acid cycle, and enhanced fatty acid oxidation.

Improvement in Menopause‐Associated Hepatic Lipid Metabolic Disorders by Herbal Formula HPC03 on Ovariectomized Rats

Postmenopausal women have an increased risk of developing nonalcoholic fatty liver disease (NAFLD). We formulated a combination of three herb mixtures (HPC03) and observed lipid-lowering efficacy. HepG2 cells were treated with oleic acid to induce an NAFLD model (in vitro). Also, we investigated potential of HPC03 in an ovariectomize- (OVX-) induced NAFLD model (in vivo). We separated the mice into six groups, as follows: SHAM, OVX, OVX + β-estradiol, and OVX + HPC03 (50, 100, and 200 mg/kg). Rats were administered with/without HPC03 for 12 weeks. HPC03 dose dependently inhibited the lipid accumulation involved in lipogenesis in HepG2 cells. The body weight, fat mass, and weights of the liver were decreased in the OVX group than that in the other groups. HPC03 had decreased adiposity that was induced by OVX. HPC03 treatment reduced liver lipid deposition and prevented the increase in serum and liver triglyceride export when there was a deficiency in estradiol. HPC03 improves OVX-induced fatty liver and lipid metabolism. These findings suggest that HPC03 from postmenopausal women has a protective effect during NAFLD conditions.

Polyphenol-rich Trapa quadrispinosa pericarp extract ameliorates high-fat diet induced non-alcoholic fatty liver disease by regulating lipid metabolism and insulin resistance in mice.

In China, Trapa quadrispinosa (also called water caltrop) has long been used as a function food and folk medicine to treat diabetes mellitus for years. In the present study, the extract of T. quadrispinosa pericarp (TQPE) which mainly contains hydrolysable tannins was prepared to investigate the potential therapeutic action in non-alcoholic fatty liver disease (NAFLD) mice induced by high fat-diet (HFD). After the administration of TQPE (15, 30 mg/kg/day) for 8 weeks, the increased weight of body and liver were significantly suppressed. TQPE also ameliorated liver lipid deposition and reduced lipids parameters of blood in mice. Moreover, TQPE attenuated oxidative stress and showed a hepatoprotective effect in mice. TQPE was also found to decrease the value of homeostatic model assessment for insulin resistance. In addition, TQPE administration increased the phosphorylation of AMP-activated protein kinase (AMPK) and Acetyl-CoA carboxylase (ACC) and inhibited sterol regulatory element-binding protein (SREBP) in the liver tissue. Meanwhile, TQPE elevated insulin receptor substrate-1 (IRs-1) and protein kinase B (Akt) phosphorylation. These results reflected that, as a nature product, TQPE is a potential agent for suppressing the process of NAFLD via regulation of the AMPK/SREBP/ACC and IRs-1/Akt pathways.

Liraglutide improves lipid metabolism by enhancing cholesterol efflux associated with ABCA1 and ERK1/2 pathway

BackgroundReverse cholesterol transport (RCT) is an important cardioprotective mechanism and the decrease in cholesterol efflux can result in the dyslipidemia. Although liraglutide, a glucagon like peptide-1 analogue, has mainly impacted blood glucose, recent data has also suggested a beneficial effect on blood lipid. However, the exact mechanism by which liraglutide modulates lipid metabolism, especially its effect on RCT, remain undetermined. Hence, the aim of the present study was to investigate the potential impacts and potential underlying mechanisms of liraglutide on the cholesterol efflux in both db/db mice and HepG2 cells.MethodsSix-week old db/db mice with high fat diet (HFD) and wild type mice were administered either liraglutide (200 μg/kg) or equivoluminal saline subcutaneously, twice daily for 8 weeks and body weight was measured every week. After the 8-week treatment, the blood was collected for lipid evaluation and liver was obtained from the mice for hematoxylin–eosin (HE) staining, red O staining and Western blotting. Cholesterol efflux was assessed by measuring the radioactivity in the plasma and feces after intraperitoneal injection of 3H-labeled cholesterol. HepG2 Cells were treated with different concentrations of glucose (0, 5, 25, and 50 mmol/L) with or without liraglutide (1000 nmol/L) for 24 h. The intracellular cholesterol efflux was detected by BODIPY-cholesterol fluorescence labeling. Real-time PCR or Western blotting was used to examine the expression levels of ABCA1, ABCG1 and SR-B1.ResultsLiraglutide significantly decreased blood glucose, serum total cholesterol (TC), triglyceride (TG) and low-density lipoprotein cholesterol (LDL-C). It also reduced liver lipid deposition in db/db mice fed with HFD. Moreover, the movement of 3H-cholesterol from macrophages to plasma and feces was significantly enhanced in db/db mice fed with HFD after liraglutide adminstration. In vitro study, liraglutide could promote the cholesterol efflux of HepG2 cells under high glucose, and also increase the expression of ABCA1 by activating the ERK1/2 pathway.ConclusionsLiraglutide could improve lipid metabolism and hepatic lipid accumulation in db/db mice fed with HFD by promoting reversal of cholesterol transport, which was associated with the up-regulation of ABCA1 mediated by the ERK1/2 phosphorylation.

FoxO1 is a critical regulator of hepatocyte lipid deposition in chronic stress mice.

Forkhead box O1 (FoxO1) is involved in lipid metabolisms. However, its role in chronic stress-related nonalcoholic fatty liver disease (NAFLD) is unclear. The scientific premise of our study was based on the finding that FoxO1 expression is increased in the liver of mice after chronic stress. It is important to understand the mechanisms involved in the activation of FoxO1 and how its function affects the liver lipid deposition. We employed a murine chronic stress model, in which mice were treated by plantar electrical stimulation and restraint for 6 weeks, and a cellular model, in which Hepa1–6 cells were treated with corticosterone. We also used a pharmacologic approach as1842856, a highly specific FoxO1 inhibitor. Lipid metabolism related genes levels were measured by qRT-PCR and the lipid levels by biochemical detection. We show that the level of FoxO1 is significantly elevated in the liver of chronic stress mice. Transcription factor FoxO1 regulates a lipid synthesis phenotype of hepatocyte that is involved in the development and progression of NAFLD. We have shown that inhibition of FoxO1 induced phenotypic conversion of hepatocytes and down-regulates lipid synthesis genes expression by hepatocytes, which contribute to lipid deposition in NAFLD. At the cellular level, the inhibitor of FoxO1 as1842856 can also attenuate the lipid deposition of Hepa1–6 cells induced by corticosterone. Targeting FoxO1 is a novel therapeutic target for chronic stress-related NAFLD.

Alisol A 24-Acetate Isolated from the Alismatis Rhizoma Improves Hepatic Lipid Deposition in Hyperlipidemic Mice by ABCA1/ABCG1 Pathway.

The Alisol A 24-acetate is an effective component of the Alismatis Rhizoma (AR) extract, which is often used in the treatment of hyperlipidemia. This study explored the effect and mechanism of the Alisol A 24-acetate from AR on lipid deposition in the liver of hyperlipidemic mice. After establishing hyperlipidemic mouse model (Model) by oral high-fat diet (HFD), the animals were treated with Alisol A 24-acetate for 4 weeks. The changes of blood lipid in mice were detected by ELISA. Hematoxylin and eosin (H&E) staining was used to evaluate the degree of liver lipid deposition in hyperlipidemic mice. Quantitative reverse transcriptase PCR (RT-qPCR) was used to detect the expression of ABCG1 and ABCA1 mRNA in the liver. The expression of ABCG1 and ABCA1 protein was detected by Western blotting (WB). After 4 weeks of high-fat diet, the levels of TC, TG, and LDL-C in the mouse were increased, and the HDL-C level was decreased. After treatment with Alisol A 24-acetate, the levels of TC, TG, and LDL-C in the blood of hyperlipidemic mouse were significantly reduced, and the level of HDL-C was increased. The results of H&E staining showed that the lipid deposition in the liver of hyperlipidemic mouse was improved after treatment with Alisol A 24-acetate. RT-qPCR and WB analysis documented that ABCG1 and ABCA1 mRNA and protein expression in hyperlipidemic mice were promoted after the Alisol A 24-acetate treatment. In conclusion, Alisol A 24-acetate effectively alleviates the liver lipid deposition in hyperlipidemic mice, and this effect is achieved mostly by promoting the expression of ABCG1 and ABCA1 at the mRNA and protein levels.

High level of dietary olive oil decreased growth, increased liver lipid deposition and induced inflammation by activating the p38 MAPK and JNK pathways in large yellow croaker (Larimichthys crocea)

A feeding experiment was conducted to determine the effects of fish oil replaced by olive oil (OO) on growth performance, serum biochemical, antioxidant capacity and inflammatory response in large yellow croaker (Larimichthys crocea). Four iso-nitrogenous and iso-lipidic diets were formulated by replacing fish oil (FO) with 0% (the control group), 33.3%, 66.7% and 100% OO. Fish fed the diet with 100% OO had the lowest growth performance among dietary treatments. However, there were no significant differences in SGR and FI among fish fed diets with 0% (the control group), 33.3% and 66.7% OO (P > 0.05). As to morphological parameters, HSI was significantly increased in fish fed the diet with 100% OO than the control group (P < 0.05). Furthermore, the lipid content of the liver in fish fed the diet with 100% OO was significantly higher than the control group (P < 0.05). Fish fed the diet with 100% OO had the highest content of C18:1n-9 among dietary treatments. Serum total triglyceride (TG), low-density lipoprotein-cholesterol (LDL-C) levels and activity of serum alanine transaminase (ALT) were significantly increased in fish fed the diet with 100% OO compared with the control group (P < 0.05). Meanwhile, dietary OO decreased the activity of superoxide dismutase (SOD) and the total antioxidant capacity (T-AOC) in fish fed diets with increasing dietary OO levels. However, the content of malondialdehyde (MDA) was significantly increased in fish fed the diet with 100% OO compared with the control group (P < 0.05). The expression of pro-inflammatory genes, COX-2, IL-1β and TNFα, were significantly increased in the liver of fish fed the diet with 100% OO compared with the control group (P < 0.05), which was probably due to the activation of p38 mitogen-activated protein kinase (p38 MAPK) pathways and Jun N-terminal kinase (JNK) as the increased protein ratio of p-p38 MAPK to p38 MAPK and p-JNK to JNK. These results suggested that high level of dietary OO decreased the growth performance and antioxidant capacity but induced inflammation via the activation of p38 MAPK and JNK pathways in large yellow croaker.

Higher dietary protein intake preserves lean body mass, lowers liver lipid deposition, and maintains metabolic control in participants with long-chain fatty acid oxidation disorders.

Medical nutrition therapy for long-chain fatty acid oxidation disorders (LC-FAODs) currently emphasizes fasting avoidance, restricted dietary long-chain fatty acid intake, supplementation with medium chain triglycerides, and increased carbohydrate intake. We hypothesize that increasing dietary protein intake relative to carbohydrate intake would preserve metabolic control yet induce physical benefits including reduced hepatic lipogenesis. Therefore, we compared two dietary approaches with similar fat intake but different carbohydrate to protein ratios in participants diagnosed with LC-FAODs. Thirteen participants were enrolled and randomized into either a high-protein (PRO) or a high-carbohydrate (CHO) diet for 4 months. Baseline and 4-month assessments included body composition, ectopic lipid deposition, and resting energy expenditure. End of study assessments also included total energy expenditure, metabolic responses to oral feedings, and whole-body fatty acid oxidation capacity. At the end of the dietary intervention, both groups had similar energy expenditure, fat and glucose oxidation rates, and glucolipid responses to mixed meal and oral glucose loads. Neither dietary group experienced worsening symptoms related to their LC-FAOD. Compared to the CHO group, the PRO group exhibited increased blood levels of short-chain acylcarnitines, reduced intrahepatic lipid content, and maintained lean body mass while the CHO group lost lean mass. In patients with LC-FAODs, increasing protein intake maintained metabolic control, reduced liver fat without risk of metabolic decompensation, and helped preserve lean body mass. We propose that a modest increase in dietary protein along with fasting avoidance and fat restriction may improve body composition and energy expenditure in patients with LC-FAODs.

Maternal betaine protects rat offspring from glucocorticoid-induced activation of lipolytic genes in adipose tissue through modification of DNA methylation.

Excessive exposure of glucocorticoids activates adipose lipolysis, increases circulating free fatty acids, and contributes to ectopic lipid deposition in liver and skeletal muscle. Our previous study demonstrated that maternal betaine supplementation attenuates glucocorticoid-induced hepatic lipid accumulation in rat offspring. However, it is unclear whether maternal betaine supplementation is effective in preventing glucocorticoid-induced lipolysis in the adipose tissue of offspring. In this study, 20 pregnant rats were fed with basal or betaine-supplemented (10g/kg) diets throughout gestation and lactation, and the offspring rats were raised on the basal diet from weaning till 3months of age followed by daily intraperitoneal injection of saline or 0.1mg/kg dexamethasone (DEX) for 3weeks. Chronic DEX treatment significantly (P < 0.05) decreased serum corticosterone level and increased proinflammatory cytokines, such as TNFα, IL-1β, and IL-6. Meanwhile, GR protein content in adipose tissue was increased in response to DEX treatment, which was associated with a significant (P < 0.05) up-regulation of ATGL and HSL expression at both mRNA and protein levels. All these DEX-induced changes were significantly (P < 0.05) attenuated in progeny rats derived from betaine-supplemented dams. Furthermore, DEX-induced hypomethylation of ATGL and HSL gene promoters was reversed by maternal betaine supplementation. Taken together, these results suggest that maternal betaine supplementation is effective in alleviating glucocorticoid-induced lipolysis in adipose tissue with modification of DNA methylation on the promoter of lipolytic genes.

MiR-122 promotes hepatic lipogenesis via inhibiting the LKB1/AMPK pathway by targeting Sirt1 in non-alcoholic fatty liver disease

BackgroundNon-alcoholic fatty liver disease (NAFLD) is a common hepatic disease with an increasing prevalence but an unclear aetiology. This study aimed to investigate the functional implications of microRNA-122 (miR-122) in the pathogenesis of NAFLD and the possible molecular mechanisms.MethodsBoth in vitro and in vivo models of NAFLD were generated by treating HepG2 and Huh-7 cells with free fatty acids (FFA) and by feeding mice a high-fat diet (HFD), respectively. HE and Oil Red O staining were used to examine liver tissue morphology and lipid deposition, respectively. Immunohistochemical (IHC) staining was used to examine Sirt1 expression in liver tissues. qRT-PCR and Western blotting were employed to measure the expression of miR-122, Sirt1, and proteins involved in lipogenesis and the AMPK pathway. Enzyme-linked immunosorbent assay (ELISA) was used to quantify triglyceride (TG) levels in HepG2 and Huh-7 cells and in liver tissues. The interaction between miR-122 and the Sirt1 gene was further examined by a dual luciferase reporter assay and RNA-immunoprecipitation (RIP).ResultsNAFLD hepatic tissues and FFA-treated HepG2 and Huh-7 cells presented excess lipid production and TG secretion, accompanied by miR-122 upregulation, Sirt1 downregulation, and potentiated lipogenesis-related genes. miR-122 suppressed Sirt1 expression via binding to its 3′-untranslated region (UTR). Knockdown of miR-122 effectively mitigated excessive lipid production and suppressed the expression of lipogenic genes in FFA-treated HepG2 and Huh-7 cells via upregulating Sirt1. Furthermore, miR-122 knockdown activated the LKB1/AMPK signalling pathway.ConclusionThe inhibition of miR-122 protects hepatocytes from lipid metabolic disorders such as NAFLD and suppresses lipogenesis via elevating Sirt1 and activating the AMPK pathway. These data support miR-122 as a promising biomarker and drug target for NAFLD.

2036-P: Gut Microbes Regulate Insulin Clearance in Mice

Obesity and insulin resistance are associated with hyperinsulinemia, which is caused by increased insulin secretion and/or reduced insulin clearance. Impaired insulin clearance may be caused by ectopic lipid deposition in liver and skeletal muscle, the key sites of insulin clearance and degradation. Insulin is the key hormone regulating blood glucose. The composition of the intestinal microbiota can regulate aspects of diet-induced obesity and glucose metabolism. Here we found that the microbiota also regulates insulin clearance during prolonged diet-induced obesity. Male C57Bl/6J mice fed 45% high fat diet (HFD) for a prolonged period had impaired insulin clearance relative to chow fed mice as measured by insulin/C-peptide ratio during an oral glucose tolerance test. High fat fed mice also showed impaired insulin clearance measured after injection of insulin - mice fed chow or 45% HFD had plasma insulin concentration measured at 0, 5, 30, and 60 minutes following injection of human insulin and area under the curve calculated. Treatment of 45% HFD-fed mice with an antibiotic cocktail of Neomycin/Ampicillin for 2 weeks improved insulin clearance after insulin injection. Next, we colonized previously germ-free mice with the microbiota from chow or 45% HFD-fed “donor” mice for 6 weeks. The previously germ-free “recipient” mice colonized with feces-resident microbes from chow or HFD fed donors did not differ in insulin clearance when recipient mice were fed a chow diet. However, when colonized recipient mice were switched from chow to 45% HFD for 2 weeks the recipient mice colonized with microbes from 45% HFD fed donors showed impaired insulin clearance relative to recipient mice colonized with microbes from chow fed donors. Our results demonstrate that diet-induced changes in fecal microbes are an independent factor regulating insulin clearance. Diet-induced changes in the gut microbiota and consequent impaired insulin clearance may participate in hyperinsulinemia leading to exacerbated insulin resistance and obesity. Disclosure K.P. Foley: None. S. Zlitni: None. B.M. Duggan: None. B.D. Henriksbo: None. N.G. Barra: None. J.F. Cavallari: None. F. Forato Anhê: None. T. Lau: None. Y. Chen: None. J.D. Schertzer: None. Funding Canadian Institutes of Health Research

352-OR: Maternal Triglycerides in Gestational Diabetes Are Strongly Associated with Increased Newborn Hepatic Fat Independent of Subcutaneous Fat

The incidence of childhood obesity continues to rise and alarmingly, 40% are already estimated to have nonalcoholic fatty liver disease (NAFLD), the leading cause of liver transplantation. An early origin exposure that contributes to NAFLD may be maternal obesity, diet, triglycerides (TG) and free fatty acids (FFA) in-utero, particularly in offspring of mothers with diabetes or gestational diabetes (GDM) who are at high risk for childhood metabolic disorders. In this pilot study, we explored the hypothesis that maternal TG in diet-controlled GDM (A1GDM) is associated with offspring intrahepatic lipid (IHL) accumulation. We randomized newly diagnosed A1GDM women (n=13, BMI 32±2 [mean±SEM]) to eucaloric diets at 32 weeks. Fasting and postprandial (PP) glucose, insulin, TG, FFA, and glycerol were measured in a breakfast test meal study (hourly x5) at baseline and 37 weeks. Newborn IHL content (Magnetic Resonance Spectroscopy; MRS) and body composition (%fat, PEAPOD) were measured at ~10 days. At 32 weeks, but not at 37 weeks, maternal fasting TG (199±17 mg/dL) and the 5-hour TG area-under-the-curve (AUC) were strongly associated with newborn IHL (r=0.69, p=0.01; r=0.75, p=0.003, respectively). The 1-hour and 2-hour PPTG (205-206±19-20 mg/dL) were highly associated with IHL (r=0.69-0.73; p≤0.01) as was the 5-hour glycerol AUC (r=0.65, p=0.01). There was no relationship between IHL and subcutaneous %fat by PEAPOD. We found no associations between maternal pre-pregnancy BMI, gestational weight gain (11±1.6 kg), glucose, insulin, or FFA and IHL. These data are the first in humans to support that maternal TG exposure at the time of GDM diagnosis, when fetal subcutaneous fat depots are early in development, may result in increased liver lipid deposition. If confirmed, MRS newborn liver fat measures and early interventions targeting maternal TG may be indicated to lower fetal liver fat deposition that could potentially be a developmental risk for pediatric NAFLD. Disclosure T.L. Hernandez: None. S.S. Farabi: None. N. Hirsch: None. E.Z. Dunn: None. E.A. Haugen: None. D. Brumbaugh: None. M.S. Brown: None. J.E. Friedman: Consultant; Self; Janssen Research &amp; Development. L.A. Barbour: None. Funding National Institutes of Health

Exendin-4 alleviates oxidative stress and liver fibrosis by activating Nrf2/HO-1 in streptozotocin-induced diabetic mice

To investigate the effects of exendin-4 on hepatic lipid metabolism, fibrosis and oxidative stress in mice with streptozotocin (STZ)-induced diabetes and explore the underlying mechanisms. C57BL/6J mice were fed with high-fat diet (HFD) for 4 weeks and received intraperitoneal injections of 120 mg/kg STZ to induce diabetes. After successful modeling, the mice were randomized into diabetic control group and exendin-4 treatment group (DM+E4), and in the latter group, the mice were given a daily dose of 1 nmol/kg of exendin-4 for 8 weeks. The changes in the body weight (BW) and random blood glucose (RBG) in the mice were recorded. The mRNA expressions of the genes related with liver lipid metabolism, fibrosis and oxidative stress were analyzed using RT-PCR, and the structural changes of the liver tissues were observed with HE, Sirius red and oil red O staining; the expressions of TGF-β1, Nrf2 and HO-1 proteins in the liver tissues were detected using Western blotting. The diabetic mice showed significantly higher RBG levels and BW with obvious lipid deposition, fibrosis and oxidative stress in the liver as compared with the normal control mice (P &lt; 0.001). Exendin-4 treatment of the diabetic mice did not significantly lessened liver lipid deposition but obviously reduced the levels of RBG and TG (P &lt; 0.05), lowered the expression levels of liver fibrosis-related genes TGF-β, α-SMA and Col-Ⅰ (P &lt; 0.05), increased the expression levels of the antioxidant genes Nrf2, HO-1 and GPX4 (P &lt; 0.01), and enhanced the protein expressions of Nrf2 and HO-1 in the liver tissues (P &lt; 0.01). Exendin-4 improves liver fibrosis and oxidative stress in diabetic mice by activating Nrf2/HO-1 pathway without significantly reducing liver lipid deposition.

Phenotypic characterization of a novel type 2 diabetes animal model in a SHANXI MU colony of Chinese hamsters.

Developing animal models for human diseases is critical for studying complex diseases such as type 2 diabetes mellitus (T2DM). Since inbred colonies of Chinese hamsters tend toward spontaneous development of diabetes, we investigated them as a possible model. We regarded individuals with fasting blood glucose (FBG) higher than 6.0 mmol/L and post-prandial blood glucose (PBG) higher than 7.0 mmol/L as diabetic based on the mean and 95% frequency distribution values of FBG and PBG. Diabetic hamsters were characterized based on metabolic profiles, histopathological features, and changes in the expression of genes involved in glucose and lipid metabolism. Metabolic analyses showed that diabetic hamsters exhibited mild hyperglycemia, hypertriglyceridemia, glucose intolerance, and insulin resistance. Histopathological analysis revealed that cell nuclei migrated inward in skeletal muscle and obvious partial liver lipid deposition and focal necrosis was found. We additionally observed mild injury, atrophy, and occasional vacuolization in islet cells. Changes in the expression of several genes related to glucose and lipid metabolism were observed. Decreased expression of adiponectin and GLUT4 and increased expression of PPARγ, Akt, and leptin was observed in skeletal muscle. Decreased expression of adiponectin with increased expression of PPARγ and leptin was observed in the liver. These results indicate that we have established a spontaneous diabetic hamster line that closely mimics human T2DM, which may hold potential for further research on the pathogenesis and treatment of this disease.

Quantification of liver lipid deposition and melano-macrophages in lesser guitarfish Zapteryx brevirostris submitted to different feeding cycles

The objective of this study was to evaluate and quantify the fat storage in the liver of lesser guitarfish Zapteryx brevirostris submitted to different periods of restriction and refeeding. In a water recirculation system, 21 animals were divided into three groups: control (n=7), fed every day; W1 (n=7), with one week of restriction and two of refeeding and W2 (n=7), with two weeks of restriction and two of refeeding. After 76 days, the animals were euthanized and liver samples were collected, paraffin embedded, stained with PAS and evaluated with stereological procedures and histopatological changes were recorded. The area occupied by fat in the liver of control group was higher than that found in groups W1 and W2 (p<0.001). Melano-macrophage center areas and diameters have risen with increased fasting period while there were swollen and necrotic hepatocytes. These results pointed to physiological adaptations and liver lesions caused by free radicals during fasting periods. Similarly to teleost fish, there was a consumption of lipid reserves while enhancing the gluconeogenesis during fasting.