Cinnamon Polyphenol Extract and Insulin Regulate Diacylglycerol Acyltransferase Gene Expression in Mouse Adipocytes and Macrophages.

Cinnamon polyphenol extract (CPE) improves people with insulin resistance. The objective was to investigate CPE and insulin on diacylglycerol acyltransferase (DGAT) gene expression important for lipid biosynthesis and compared it to anti-inflammatory tristetraprolin/zinc finger protein 36 (TTP/ZFP36) gene expression known to be regulated by both agents. Mouse 3T3-L1 adipocytes and RAW264.7 macrophages were treated with insulin and CPE followed by qPCR evaluation of DGAT and TTP mRNA levels. Insulin decreased DGAT1 and DGAT2 mRNA levels in adipocytes but had no effect on DGAT1 and increased DGAT2 mRNA levels 3-fold in macrophages. Insulin increased TTP mRNA levels 3-fold in adipocytes but had no effect in macrophages. CPE effect on DGAT1 gene expression was minimal but increased DGAT2 mRNA levels 2-4 fold in adipocytes and macrophages. CPE increased TTP mRNA levels 2-7 fold in adipocytes and macrophages. We conclude that CPE and insulin exhibited overlapping and independent effects on DGAT and TTP gene expression and suggest that CPE and insulin have profound effects on fat biosynthesis and inflammatory responses.

Ghrelin deletion protects against age-associated hepatic steatosis by downregulating the C/EBPα-p300/DGAT1 pathway.

Nonalcoholic fatty liver disease (NAFLD) is the most common liver disease worldwide. NAFLD usually begins as low-grade hepatic steatosis which further progresses in an age-dependent manner to nonalcoholic steatohepatitis (NASH), fibrosis, cirrhosis, and hepatocellular carcinoma in some patients. Ghrelin is a hormone known to promote adiposity in rodents and humans, but its potential role in hepatic steatosis is unknown. We hypothesized that genetic ghrelin deletion will protect against the development of age-related hepatic steatosis. To examine this hypothesis, we utilized ghrelin knockout (KO) mice. Although no different in young animals (3 months old), we found that at 20 months of age, ghrelin KO mice have significantly reduced hepatic steatosis compared to aged-matched wild-type (WT) mice. Examination of molecular pathways by which deletion of ghrelin reduces steatosis showed that the increase in expression of diacylglycerol O-acyltransferase-1 (DGAT1), one of the key enzymes of triglyceride (TG) synthesis, seen with age in WT mice, is not present in KO mice. This was due to the lack of activation of CCAAT/enhancer binding protein-alpha (C/EBPα) protein and subsequent reduction of C/EBPα-p300 complexes. These complexes were abundant in livers of old WT mice and were bound to and activated the DGAT1 promoter. However, the C/EBPα-p300 complexes were not detected on the DGAT1 promoter in livers of old KO mice resulting in lower levels of the enzyme. In conclusion, these studies demonstrate the mechanism by which ghrelin deletion prevents age-associated hepatic steatosis and suggest that targeting this pathway may offer therapeutic benefit for NAFLD.

Acute Testosterone Deficiency Alters Adipose Tissue Fatty Acid Storage.

Context: Although the long-term effects of testosterone on adipose tissue lipid metabolism in men have been defined, the short-term regulation of these effects is not well understood. Objective: We examined the effects of acute testosterone withdrawal on subcutaneous abdominal and femoral adipose tissue fatty acid (FA) storage and cellular mechanisms. Design: This was a prospective, randomized trial. Setting: Mayo Clinic Clinical Research Unit. Patients or Participants: Thirty-two male volunteers ages 18 to 50 participated in these studies. Interventions: Volunteers were randomized to receive (1) no treatment (control), (2) injections (7.5 mg) of Lupron®, or (3) Lupron and testosterone (L+T) replacement for 49 days, resulting in 4 weeks of sex steroid suppression in the Lupron group. Main Outcome Measures: We measured body composition, fat cell size, adipose tissue meal FA and direct free FA storage, lipoprotein lipase (LPL), acyl coenzyme A synthetase (ACS), diacylglycerol acyltransferase activities, and CD36 content. Results: Compared with control and L+T groups, acute testosterone deficiency resulted in greater femoral adipose tissue meal FA storage rates, fasting and fed LPL activity, and ACS activity. Conclusions: These results suggest that in men, testosterone plays a tonic role in restraining FA storage in femoral adipose tissue via suppression of LPL and ACS activities. FA storage mechanisms in men appear sensitive to short-term changes in testosterone concentrations.

Diacylglycerol acyltransferase 1 inhibition with AZD7687 alters lipid handling and hormone secretion in the gut with intolerable side effects: a randomized clinical trial.

AIM: Inhibition of diacylglycerol acyltransferase 1 (DGAT1) is a potential treatment modality for patients with type 2 diabetes mellitus and obesity, based on preclinical data suggesting it is associated with insulin sensitization and weight loss. This randomized, placebo-controlled, phase 1 study in 62 overweight or obese men explored the effects and tolerability of AZD7687, a reversible and selective DGAT1 inhibitor. METHODS: Multiple doses of AZD7687 (1, 2.5, 5, 10 and 20 mg/day, n = 6 or n = 12 for each) or placebo (n = 20) were administered for 1 week. Postprandial serum triacylglycerol (TAG) was measured for 8 h after a standardized 45% fat meal. Glucagon-like peptide-1 (GLP-1) and peptide YY (PYY) were measured and a paracetamol challenge was performed to assess gastric emptying. RESULTS: Dose-dependent reductions in postprandial serum TAG were demonstrated with AZD7687 doses ≥5 mg compared with placebo (p < 0.01). Significant (p < 0.001) increases in plasma GLP-1 and PYY levels were seen at these doses, but no clear effect on gastric emptying was demonstrated at the end of treatment. With AZD7687 doses >5 mg/day, gastrointestinal (GI) side effects increased; 11/18 of these participants discontinued treatment owing to diarrhoea. CONCLUSIONS: Altered lipid handling and hormone secretion in the gut were demonstrated during 1-week treatment with the DGAT1 inhibitor AZD7687. However, the apparent lack of therapeutic window owing to GI side effects of AZD7687, particularly diarrhoea, makes the utility of DGAT1 inhibition as a novel treatment for diabetes and obesity questionable.

Beneficial effects of mangiferin on hyperlipidemia in high-fat-fed hamsters.

SCOPE: Mangiferin, a natural polyphenol, has been shown to have hypolipidemic effect in rat and mouse. However, the mechanism of action is not well understood. This study was conducted to determine the effect and mechanism of action of mangiferin on hyperlipidemia induced in hamsters by a high-fat diet. METHODS AND RESULTS: Forty male hamsters were randomly assigned to normal control, high-fat control, and high fat with mangiferin (50 and 150 mg/kg BW) groups. Mangiferin treatment significantly decreased final body weight, liver weight and visceral fat-pad weight, serum triglyceride (TG) and total free fatty acid (FFA) concentrations, hepatic TG levels and hepatic and muscle total FFA contents. Mangiferin upregulated mRNA expression of peroxisome proliferator-activated receptor-α (PPAR-α), fatty acid translocase (CD36) and carnitine palmitoyltransferase 1 (CPT-1), but downregulated mRNA expression of sterol regulatory element-binding protein 1c (SREBP-1c), acetyl CoA carboxylase (ACC), acyl-CoA:diacylglycerol acyltransferase 2 (DGAT-2) and microsomal triglyceride transfer protein (MTP) in liver. Mangiferin also stimulated mRNA expression of PPAR-α, CD36, CPT-1 and lipoprotein lipase (LPL) in muscle. CONCLUSIONS: The results suggest that mangiferin may ameliorate hypertriglyceridemia partly by modulating the expression levels of genes involved in lipid oxidation and lipogenesis.

Osthole improves alcohol-induced fatty liver in mice by reduction of hepatic oxidative stress.

The aim of our study was to examine the therapeutic effect of osthole, an active constituent isolated from the fruit of Cnidium monnieri (L.) Cusson, on alcohol-induced fatty liver in mice and investigate its potential mechanisms of treatment. A mouse alcoholic fatty liver model was established by feeding 52% alcohol for 4 weeks. These experimental mice were then treated with osthole 10, 20 and 40 mg/kg for 6 weeks. The levels of serum total cholesterol (TC), triglyceride (TG), low density lipoprotein-cholesterol (LDL-C) and hepatic tissue contents of TC, TG and malondialdehyde (MDA) in osthole-treated groups were significantly decreased, while the level of superoxide dismutase (SOD) was significantly increased compared with the model group. Moreover, the cytochrome P450 (CYP) 2E1 and diacylglycerol acyltransferase (DGAT) mRNA expressions in mouse liver were significantly decreased, and the carnitine palmitoyltransferase (CPT) 1A mRNA expression was increased by osthole treatment. Importantly, the histological evaluation of liver demonstrated that osthole dramatically decreased lipid accumulation. It was concluded that osthole was effective in treating mouse alcoholic fatty liver, and its main mechanisms might be related to reduction of hepatic oxidative stress, including the inhibition of reactive oxygen species (ROS) production, enhancement of antioxidative enzyme activity, and reduction of lipid accumulation and peroxidation.

Fenofibrate ameliorates diabetic and dyslipidemic profiles in KKAy mice partly via down-regulation of 11beta-HSD1, PEPCK and DGAT2. Comparison of PPARalpha, PPARgamma, and liver x receptor agonists.

Fenofibrate and rosiglitazone are prescribed to treat hypertriglyceridemia and diabetes, respectively. Since fenofibrate improves lipid profile in diabetic patients and improves insulin resistance in animal models, we examined the mechanism of antidiabetic effects of fenofibrate in KKAy mouse, an animal model of diabetes and dyslipidemia. KKAy mice were treated with fenofibrate, rosiglitazone, liver x receptor agonist, N-(2,2,2-trifluoroethyl)-N-[4-[2,2,2-trifluoro-1-hydroxy-1-(trifluoromethyl)ethyl]phenyl]-benzenesulfonamide (T0901317), and a combination of fenofibrate and T090317 for 2 weeks. Fenofibrate lowered serum triglycerides by 90% and free fatty acid (FFA) by 50% via inhibition of hepatic fatty acid synthesis. Fenofibrate also prevented T0901317-induced increases of triglycerides by dampening T090317-mediated sterol response element binding protein 1c (SREBP1c) up-regulation. Glucose lowering was comparable (approximately 40%) in fenofibrate and rosiglitazone treated mice. T090317 also showed mild reduction in serum glucose, in part, via down-regulation of phosphoenol pyruvate carboxykinase (PEPCK). Combining fenofibrate with T0901317 caused greater reduction in serum glucose, suggesting an additive effect. The mechanism of lipid and glucose lowering in KKAy mice was examined. Liver PEPCK showed down-regulation in all treatment groups with fenofibrate showing greater effects. Combination of fenofibrate with T090317 showed additive effects on PEPCK down-regulation. Fenofibrate decreased hepatic diacyl glycerol acyl transferase 2 (DGAT2) mRNA leading to reduction in triglyceride synthesis. Most importantly, fenofibrate down regulated expression of hepatic and adipose 11beta hydroxysteroid dehydrogenase (11beta-HSD1) gene, which contributed in attenuating diabetic state. Thus, amelioration of antidiabetic and hyperlipidemic state by fenofibrate in KKAy mice occurred via down-regulation of DGAT2, PEPCK and 11beta-HSD1. It is also shown that the undesirable lipogenic effects of T090317 could be dampened by fenofibrate.

Diacylglycerol acyltransferases: Potential roles as pharmacological targets.

Triglyceride (TG) synthesis occurs in many cell-types, but only the adipocyte is specialised for TG storage. The increased incidence of obesity and its attendant pathologies have increased interest in pharmacological strategies aimed at inhibition of triglyceride synthesis. In the liver this would also appear to offer the advantages of the prevention of steatosis and/or dyslipidaemia. The two major enzymes that have DGAT activity appear to have specialised functions, that are most evident in triglyceride-secreting tissues. The presence of triglyceride in non-adipose cells can lead to (through lipolysis), or be a marker for, undesirable complications such as insulin resistance, or can be indicative of simultaneously high capacities for triglyceride synthesis, lipolysis and oxidation of fatty acids as in highly aerobic, trained muscle. Consequently, inhibition of triglyceride synthesis may not be a straightforward strategy, either in terms of its achievement pharmacologically or in its anticipated outcomes. The metabolic complexities of triglyceride synthesis, with particular reference to the diacylglycerol acyltransferases (DGATs) are considered in this short review.

Inhibition of diacylglycerol acyltransferase by betulinic acid from Alnus hirsuta.

During the screening for diacylglycerol acyltransferase (DGAT) inhibitors from natural products, the lupane-type triterpenoid betulinic acid was isolated from the methanol extract of Alnus hirsuta. It potently inhibited DGAT in the rat liver microsomes with an IC (50) value of 9.6 microM. Enzyme kinetic studies showed apparent Km and Ki values of 13.3 microM and 8.1 microM using [(14)C]oleoyl-CoA as a substrate. A decrease in the apparent Vmax was observed with betulinic acid, whereas the apparent Km remained constant. Therefore, a Lineweaver-Burk plot of DGAT inhibition by betulinic acid showed a non-competitive type of inhibition. In the cell-based assay, betulinic acid inhibited triglyceride (TG) formation by human HepG2 cells. These findings suggest that betulinic acid may be a potential lead compound in the treatment of obesity.

Hepatic lipid and carbohydrate metabolism in rats fed a commercial mixture of conjugated linoleic acids (Clarinol G-80).

BACKGROUND: Conjugated linoleic acids (CLAs) exert numerous effects in animal models as well as in humans. Among other things, CLAs decrease plasma lipid levels and bring about hepatic steatosis. The latter effects are attributed to an agonistic action of CLAs on the peroxisome-proliferator-activated receptor family primarily responsible for activating genes involved in lipid metabolism and are related to changes in mRNA levels. Such changes are not necessarily reflected in changes in activity of controlling enzymes. AIM OF THE STUDY: To investigate the effects of CLAs treatment on lipid metabolism, we determined lipid concentrations in plasma, lipoproteins and liver and measured the activity of a number of key enzymes in hepatic lipid metabolism as differences in lipid concentrations should be related to changes in enzyme activities. These variables were determined with the rat as a model. METHODS: Rats were fed a control diet or a diet containing 1.15% trans-10, cis-12 isomer and 1.11% cis-9, trans-11 isomer as part of a commercial mixture of CLAs. After 2 w the animals were killed, and plasma and liver fractions isolated. Subsequently, lipid concentrations of cholesterol, triacylglycerols and phospholipids were determined in the isolated lipoproteins. In livers homogenates, the concentrations of glycogen, cholesterol, triacylglycerol and phospholipids and the activities of enzymes catalyzing pacesetting steps of metabolism were determined, i. e. acetyl-CoA carboxylase, fatty acid synthase, diacylglycerol acyltransferase, 3-hydroxy- acyl-CoA dehydrogenase, citrate synthase and phosphofructokinase. RESULTS: CLAs induced a lowering of the cholesterol levels in total plasma and in LDL and HDL lipoproteins and of phospholipid concentrations in LDL and HDL. CLAs treatment decreased the hepatic activity of diacylglycerol acyltransferase and had no effect on any of the other enzyme activities. CONCLUSIONS: In other studies enhanced specific activities of ACC and FAS were found in livers of mice using the same or similar methods and experimental protocol as in the present study. The lack of effect of CLAs treatment on hepatic key enzymes of fatty acid synthesis and oxidation in Wistar rats questions the use of this strain for studying the mechanism(s) underlying CLA's effects on these parameters. However, in the rat model we observed reduced levels of cholesterol in total plasma and in LDL and HDL. Therefore, some aspects like loss of body fat are better studied in mice; for other aspects like reduction in serum cholesterol level the rat may be the model of choice.