Adipose Tissue Protects against Hepatic Steatosis in Male Rats Fed a High-Fat Diet plus Liquid Fructose: Sex-Related Differences.

Non-alcoholic fatty liver disease is a sexual dimorphic disease, with adipose tissue playing an essential role. Our previous work showed that female rats fed a high-fat high-fructose diet devoid of cholesterol (HFHFr) developed simple hepatic steatosis dissociated from obesity. This study assessed the impact of the HFHFr diet on the male rat metabolism compared with data obtained for female rats. A total of 16 Sprague Dawley (SD) male rats were fed either a control (standard rodent chow and water) or HFHFr (high-fat diet devoid of cholesterol, plus 10% fructose in drinking water) diet for 3 months. Unlike female rats, and despite similar increases in energy consumption, HFHFr males showed increased adiposity and hyperleptinemia. The expression of hormone-sensitive lipase in the subcutaneous white adipose tissue was enhanced, leading to high free fatty acid and glycerol serum levels. HFHFr males presented hypertriglyceridemia, but not hepatic steatosis, partially due to enhanced liver PPARα-related fatty acid ß-oxidation and the VLDL-promoting effect of leptin. In conclusion, the SD rats showed a sex-related dimorphic response to the HFHFr diet. Contrary to previous results for HFHFr female rats, the male rats were able to expand the adipose tissue, increase fatty acid catabolism, or export it as VLDL, avoiding liver lipid deposition.

KHK, PNPLA3 and PPAR as Novel Targets for the Anti-Steatotic Action of Bempedoic Acid.

Bempedoic acid (BemA) is an ATP-citrate lyase (ACLY) inhibitor used to treat hypercholesterolemia. We studied the anti-steatotic effect of BemA, and the mechanisms involved, in a model of fatty liver in female rats obtained through the administration of a high-fat diet supplemented with liquid fructose (HFHFr) for three months. In the third month, a group of rats was treated with BemA (30 mg/kg/day) by gavage. Plasma analytes, liver histology, adiposity, and the expression of key genes controlling fatty acid metabolism were determined, and PPAR agonism was explored by using luciferase reporter assays. Our results showed that, compared to HFHFr, BemA-treated rats exhibited lower body weight, higher liver/body weight, and reduced hepatic steatosis. In addition to ACLY inhibition, we found three novel mechanisms that could account for the anti-steatotic effect: (1) reduction of liver ketohexokinase, leading to lower fructose intake and reduced de novo lipogenesis; (2) increased expression of patatin-like phospholipase domain-containing protein 3, a protein related to the export of liver triglycerides to blood; and (3) PPARα agonist activity, leading to increased hepatic fatty acid ß-oxidation. In conclusion, BemA may represent a novel approach to treat hepatic steatosis, and therefore to avoid progression to advanced stages of non-alcoholic fatty liver disease.

ChREBP-driven DNL and PNPLA3 Expression Induced by Liquid Fructose are Essential in the Production of Fatty Liver and Hypertriglyceridemia in a High-Fat Diet-Fed Rat Model.

SCOPE: The aim of this study is to delineate the contribution of dietary saturated fatty acids (FA) versus liquid fructose to fatty liver and hypertriglyceridemia. METHODS AND RESULTS: Three groups of female rats are maintained for 3 months in standard chow (CT); High-fat diet (46.9% of fat-derived calories, rich in palmitic and stearic FA, HFD); and HFD with 10% w/v fructose in drinking water (HFHFr). Zoometric parameters, plasma biochemistry, and liver Oil-Red O (ORO) staining, lipidomics, and expression of proteins involved in FA metabolism are analyzed. Both diets increase ingested calories without modifying body weight. Only the HFHFr diet increases liver triglycerides (x11.0), with hypertriglyceridemia (x1.7) and reduces FA ß-oxidation (x0.7), and increases liver FA markers of DNL (de novo lipogenesis). Whereas HFD livers show a high content of ceramides, HFHFr samples show unchanged ceramides, and an increase in diacylglycerols. Only the HFHFr diet leads to a marked increase in the expression of enzymes involved in DNL and triglyceride metabolism, such as carbohydrate response element binding protein ß (ChREBPß, x3.2), a transcription factor that regulates DNL, and patatin-like phospholipase domain-containing 3 (PNPLA3, x2.6), a lipase that mobilizes stored triglycerides for VLDL secretion. CONCLUSION: The addition of liquid-fructose to dietary FA is determinant in liver steatosis and hypertriglyceridemia production, through increased DNL and PNPLA3 expression, and reduced FA catabolism.

Bempedoic Acid Restores Liver H2S Production in a Female Sprague-Dawley Rat Dietary Model of Non-Alcoholic Fatty Liver.

We previously demonstrated that treatment with BemA (bempedoic acid), an inhibitor of ATP citrate lyase, significantly reduces fatty liver in a model of liver steatosis (HFHFr-female Sprague-Dawley rat fed a high-fat high-fructose diet). Since the hepatic production of the gasotransmitter H2S is impaired in liver disorders, we were interested in determining if the production of H2S was altered in our HFHFr model and whether the administration of BemA reversed these changes. We used stored liver samples from a previous study to determine the total and enzymatic H2S production, as well as the expression of CBS (cystathionine ß-synthase), CSE (cystathionine γ-lyase), and 3MST (3-mercaptopiruvate sulfurtransferase), and the expression/activity of FXR (farnesoid X receptor), a transcription factor involved in regulating CSE expression. Our data show that the HFHFr diet reduces the total and enzymatic production of liver H2S, mainly by decreasing the expression of CBS and CSE. Furthermore, BemA treatment restored H2S production, increasing the expression of CBS and CSE, providing evidence for the involvement of FXR transcriptional activity and the mTORC1 (mammalian target of rapamycin1)/S6K1 (ribosomal protein S6 kinase beta-1)/PGC1α (peroxisome proliferator receptor gamma coactivator1α) pathway.

Bempedoic acid as a PPARα activator: new perspectives for hepatic steatosis treatment in a female rat experimental model. / El ácido bempedoico como activador PPARα: Nuevas perspectivas para el tratamiento de la esteatosis hepática en un modelo experimental de rata hembra.

INTRODUCTION: In its initial stages, nonalcoholic fatty liver disease presents hypertriglyceridemia and accumulation of lipids in the liver (hepatic steatosis). Bempedoic acid is an ATP:citrate lyase inhibitor that promotes a dual inhibition of the synthesis of cholesterol and fatty acids. However, its effect in the prevention / treatment of hepatic steatosis and hypertriglyceridemia has not been investigated. The aim of our work has been to elucidate whether bempedoic acid, through a mechanism other than ATP:citrate lyase inhibition, reverses these metabolic alterations. EXPERIMENTAL DESIGN: The study was carried out in female Sprague-Dawley rats fed, for three months, with a high fat diet supplemented with fructose (10% w/v) in drinking water. During the last month, bempedoic acid (30mg/kg/day) was administered to a group of animals. Zoometric and plasmatic parameters were analyzed, gene and protein expression analysis were performed in liver samples and PPAR-PPRE binding activity was determined. RESULTS: Our interventional model developed hepatic steatosis and hypertriglyceridemia. Despite an increase in total caloric intake, there was no increase in body weight of the animals. The administration of bempedoic acid significantly reduced hepatic steatosis and promoted a marked hepatocyte hypertrophy. There was a 66% increase in the liver weight of the animals treated with the drug that was not accompanied by modifications in the markers of inflammation, oxidative stress, or endoplasmic reticulum stress. Bempedoic acid activated the peroxisome proliferator activated nuclear receptor (PPARα) and its target genes. CONCLUSIONS: Bempedoic acid could be an effective therapy for the treatment of fatty liver and associated cardiovascular risk. Bempedoic acid has other mechanisms of action besides the inhibition of ATP: citrate lyase, such as the activation of PPARα, which could explain the reduction in hepatic steatosis and the increase in liver weight observed in animals treated with the drug.

Effects of a Low Dose of Caffeine Alone or as Part of a Green Coffee Extract, in a Rat Dietary Model of Lean Non-Alcoholic Fatty Liver Disease without Inflammation.

Non-alcoholic fatty liver disease is a highly prevalent condition without specific pharmacological treatment, characterized in the initial stages by hepatic steatosis. It was suggested that lipid infiltration in the liver might be reduced by caffeine through anti-inflammatory, antioxidative, and fatty acid metabolism-related mechanisms. We investigated the effects of caffeine (CAF) and green coffee extract (GCE) on hepatic lipids in lean female rats with steatosis. For three months, female Sprague-Dawley rats were fed a standard diet or a cocoa butter-based high-fat diet plus 10% liquid fructose. In the last month, the high-fat diet was supplemented or not with CAF or a GCE, providing 5 mg/kg of CAF. Plasma lipid levels and the hepatic expression of molecules involved in lipid metabolism were determined. Lipidomic analysis was performed in liver samples. The diet caused hepatic steatosis without obesity, inflammation, endoplasmic reticulum stress, or hepatic insulin resistance. Neither CAF nor GCE alleviated hepatic steatosis, but GCE-treated rats showed lower hepatic triglyceride levels compared to the CAF group. The GCE effects could be related to reductions of hepatic (i) mTOR phosphorylation, leading to higher nuclear lipin-1 levels and limiting lipogenic gene expression; (ii) diacylglycerol levels; (iii) hexosylceramide/ceramide ratios; and (iv) very-low-density lipoprotein receptor expression. In conclusion, a low dose of CAF did not reduce hepatic steatosis in lean female rats, but the same dose provided as a green coffee extract led to lower liver triglyceride levels.

The Effect of Silymarin Flavonolignans and Their Sulfated Conjugates on Platelet Aggregation and Blood Vessels Ex Vivo.

Silymarin is a traditional drug and food supplement employed for numerous liver disorders. The available studies indicate that its activities may be broader, in particular due to claimed benefits in some cardiovascular diseases, but the contributions of individual silymarin components are unclear. Therefore, we tested silymarin flavonolignans as pure diastereomers as well as their sulfated metabolites for potential vasorelaxant and antiplatelet effects in isolated rat aorta and in human blood, respectively. Eleven compounds from a panel of 17 tested exhibited a vasorelaxant effect, with half maximal effective concentrations (EC50) ranging from 20 to 100 µM, and some substances retained certain activity even in the range of hundreds of nM. Stereomers A were generally more potent as vasorelaxants than stereomers B. Interestingly, the most active compound was a metabolite-silychristin-19-O-sulfate. Although initial experiments showed that silybin, 2,3-dehydrosilybin, and 2,3-dehydrosilychristin were able to substantially block platelet aggregation, their effects were rapidly abolished with decreasing concentration, and were negligible at concentrations ≤100 µM. In conclusion, metabolites of silymarin flavonolignans seem to have biologically relevant vasodilatory properties, but the effect of silymarin components on platelets is low or negligible.