Sex Differences Affect the NRF2 Signaling Pathway in the Early Phase of Liver Steatosis: A High-Fat-Diet-Fed Rat Model Supplemented with Liquid Fructose.

Sex differences may play a role in the etiopathogenesis and severity of metabolic dysfunction-associated steatotic liver disease (MASLD), a disorder characterized by excessive fat accumulation associated with increased inflammation and oxidative stress. We previously observed the development of steatosis specifically in female rats fed a high-fat diet enriched with liquid fructose (HFHFr) for 12 weeks. The aim of this study was to better characterize the observed sex differences by focusing on the antioxidant and cytoprotective pathways related to the KEAP1/NRF2 axis. The KEAP1/NRF2 signaling pathway, autophagy process (LC3B and LAMP2), and endoplasmic reticulum stress response (XBP1) were analyzed in liver homogenates in male and female rats that were fed a 12-week HFHFr diet. In females, the HFHFr diet resulted in the initial activation of the KEAP1/NRF2 pathway, which was not followed by the modulation of downstream molecular targets; this was possibly due to the increase in KEAP1 levels preventing the nuclear translocation of NRF2 despite its cytosolic increase. Interestingly, while in both sexes the HFHFr diet resulted in an increase in the levels of LC3BII/LC3BI, a marker of autophagosome formation, only males showed a significant upregulation of LAMP2 and XBP1s; this did not occur in females, suggesting impaired autophagic flux in this sex. Overall, our results suggest that males are characterized by a greater ability to cope with an HFHFr metabolic stimulus mainly through an autophagic-mediated proteostatic process while in females, this is impaired. This might depend at least in part upon the fine modulation of the cytoprotective and antioxidant KEAP1/NRF2 pathway resulting in sex differences in the occurrence and severity of MASLD. These results should be considered to design effective therapeutics for MASLD.

Pemafibrate abrogates SLD in a rat experimental dietary model, inducing a shift in fecal bile acids and microbiota composition.

BACKGROUND AND AIMS: Drugs resolving steatotic liver disease (SLD) could prevent the evolution of metabolic dysfunction associated SLD (MASLD) to more aggressive forms but must show not only efficacy, but also a high safety profile. Repurposing of drugs in clinical use, such as pemafibrate and mirabegron, could facilitate the finding of an effective and safe drug-treatment for SLD. APPROACH AND RESULTS: The SLD High Fat High Fructose (HFHFr) rat model develops steatosis without the influence of other metabolic disturbances, such as obesity, inflammation, or type 2 diabetes. Further, liver fatty acids are provided, as in human pathology, both from dietary origin and de novo lipid synthesis. We used the HFHFr model to evaluate the efficacy of pemafibrate and mirabegron, alone or in combination, in the resolution of SLD, analyzing zoometric, biochemical, histological, transcriptomic, fecal metabolomic and microbiome data. We provide evidence showing that pemafibrate, but not mirabegron, completely reverted liver steatosis, due to a direct effect on liver PPARα-driven fatty acid catabolism, without changes in total energy consumption, subcutaneous, perigonadal and brown fat, blood lipids and body weight. Moreover, pemafibrate treatment showed a neutral effect on whole-body glucose metabolism, but deeply modified fecal bile acid composition and microbiota. CONCLUSIONS: Pemafibrate administration reverts liver steatosis in the HFHFr dietary rat SLD model without altering parameters related to metabolic or organ toxicity. Our results strongly support further clinical research to reposition pemafibrate for the treatment of SLD/MASLD.

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.

Bempedoic acid for nonalcoholic fatty liver disease: evidence and mechanisms of action.

PURPOSE OF REVIEW: Nonalcoholic fatty liver disease (NAFLD) is a highly prevalent progressive condition that lacks a specific pharmacological treatment. ATP-citrate lyase (ACLY) is one of the emergent targets for the treatment of NAFLD. This review aims to summarize the role of ACLY in NAFLD, provide evidence of the beneficial effects of the ACLY inhibitor bempedoic acid (BemA) in NAFLD and discuss the mechanisms involved. RECENT FINDINGS: BemA is effective in reducing hepatic steatosis in several animal models that recapitulate different stages of the disease. Thus, in a dietary model of simple hepatic steatosis in female rats, BemA abrogates the accumulation of liver fat. Apart from ACLY inhibition, BemA has several functions in the liver that contribute to the antisteatotic effect: inhibition of ketohexokinase, induction of patatin-like phospholipase domain-containing protein 3 and increases in both fatty acid ß-oxidation activity and hepatic H 2 S production. In models of the advanced phases of NAFLD, BemA reduces not only steatosis, but also ballooning, lobular inflammation and hepatic fibrosis, by mechanisms involving both hepatocytes and hepatic stellate cells. SUMMARY: BemA, an ACLY inhibitor currently approved for the treatment of hypercholesterolemia, may be a useful drug to treat NAFLD through its antisteatotic, anti-inflammatory and antifibrotic effects.

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.

Chronic liquid fructose supplementation does not cause liver tumorigenesis but elicits clear sex differences in the metabolic response in Sprague-Dawley rats.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) has increased over the last decades and may evolve into hepatocellular carcinoma (HCC). As HCC is challenging to treat, knowledge on the modifiable risk factors for NAFLD/HCC (e.g. hyper caloric diets rich in fructose) is essential. OBJECTIVE AND DESIGN: We used a model of diethyl nitrosamine-induced hepatocarcinogenesis to investigate the liver cancer-promoting effects of a diet supplemented with 10% liquid fructose, administered to male and female rats for 11 months. A subset of the fructose-supplemented rats received resveratrol (RVT) in the last 4 months of treatment. RESULTS AND DISCUSSION: Rat livers showed no de visu or histological evidence of liver tumorigenesis. However, we observed metabolic abnormalities that could be related to cancer development mainly in the female fructose-supplemented rats, such as increases in weight, adiposity and hepatic triglyceride levels, as well as hyperglycaemia, hyperuricemia, hyperleptinemia and a reduced insulin sensitivity index, which were partially reversed by RVT. Therefore, we performed a targeted analysis of 84 cancer-related genes in the female liver samples, which revealed expression changes associated with cancer-related pathways. Analysis of individual genes indicated that some changes increased the risk of hepatocarcinogenesis (Sfrp2, Ccl5, Socs3, and Gstp1), while others exerted a protective/preventive effect (Bcl2 and Cdh1). CONCLUSION: Our data clearly demonstrate that chronic fructose supplementation, as the sole dietary intervention, does not cause HCC development in rats.

Nicotinamide Protects Against Diet-Induced Body Weight Gain, Increases Energy Expenditure, and Induces White Adipose Tissue Beiging.

SCOPE: Interventions that boost NAD+ availability are of potential therapeutic interest for obesity treatment. The potential of nicotinamide (NAM), the amide form of vitamin B3 and a physiological precursor of nicotinamide adenine dinucleotide (NAD)+ , in preventing weight gain has not previously been studied in vivo. Other NAD+ precursors have been shown to decrease weight gain; however, their impact on adipose tissue is not addressed. METHODS AND RESULTS: Two doses of NAM (high dose: 1% and low dose: 0.25%) are given by drinking water to C57BL/6J male mice, starting at the same time as the high-fat diet feeding. NAM supplementation protects against diet-induced obesity by augmenting global body energy expenditure in C57BL/6J male mice. The manipulation markedly alters adipose morphology and metabolism, particularly in inguinal (i) white adipose tissue (iWAT). An increased number of brown and beige adipocyte clusters, protein abundance of uncoupling protein 1 (UCP1), mitochondrial activity, adipose NAD+ , and phosphorylated AMP-activated protein kinase (P-AMPK) levels are observed in the iWAT of treated mice. Notably, a significant improvement in hepatic steatosis, inflammation, and glucose tolerance is also observed in NAM high-dose treated mice. CONCLUSION: NAM influences whole-body energy expenditure by driving changes in the adipose phenotype. Thus, NAM is an attractive potential treatment for preventing obesity and associated complications.

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.

Mesenteric arterial dysfunction in the UC Davis Type 2 Diabetes Mellitus rat model is dependent on pre-diabetic versus diabetic status and is sexually dimorphic.

Previous reports suggest that diabetes may differentially affect the vascular beds of females and males. However, there is insufficient evidence to establish the timeline of the vascular dysfunction in diabetes, specifically in relation to sex. Here, we determined whether mesenteric arterial function is altered in UC Davis Type-2 Diabetes Mellitus (UCD-T2DM) rats and if this occurs as early as the pre-diabetic stage of the disease. Specifically, we investigated whether vascular dysfunction differs between pre-diabetic or diabetic status and if this varies by sex. We measured the responses to endothelium-dependent and -independent vasorelaxant as well as vasoconstrictor agents and explored the potential mechanisms involved in sex-specific development of arterial dysfunction in UCD-T2DM rats. In addition, indices of insulin sensitivity were assessed. We report the reduced insulin sensitivity in pre-diabetic males and diabetic females. Vascular relaxation to acetylcholine was impaired to a greater extent in mesenteric artery from males in the pre-diabetic stage than in their female counterparts. In contrast, the arteries from females with diabetes exhibited a greater impairment to acetylcholine compared with diabetic males. Additionally, the sensitivity of mesenteric artery to contractile agents in females, but not in males, after the onset of diabetes was increased. Our data suggest that the reduced insulin sensitivity through AKT may predispose vessels to injury in the pre-diabetic stage in males. On the other hand, reduced insulin sensitivity as well as enhanced responsiveness to contractile agents may predispose arteries to injury in the diabetic stage in females.

Low-density lipoprotein receptor-related protein 1 deficiency in cardiomyocytes reduces susceptibility to insulin resistance and obesity.

BACKGROUND: Low-density lipoprotein receptor-related protein 1 (LRP1) plays a key role in fatty acid metabolism and glucose homeostasis. In the context of dyslipemia, LRP1 is upregulated in the heart. Our aim was to evaluate the impact of cardiomyocyte LRP1 deficiency on high fat diet (HFD)-induced cardiac and metabolic alterations, and to explore the potential mechanisms involved. METHODS: We used TnT-iCre transgenic mice with thoroughly tested suitability to delete genes exclusively in cardiomyocytes to generate an experimental mouse model with conditional Lrp1 deficiency in cardiomyocytes (TNT-iCre+-LRP1flox/flox). FINDINGS: Mice with Lrp1-deficient cardiomyocytes (cm-Lrp1-/-) have a normal cardiac function combined with a favorable metabolic phenotype against HFD-induced glucose intolerance and obesity. Glucose intolerance protection was linked to higher hepatic fatty acid oxidation (FAO), lower liver steatosis and increased whole-body energy expenditure. Proteomic studies of the heart revealed decreased levels of cardiac pro-atrial natriuretic peptide (pro-ANP), which was parallel to higher ANP circulating levels. cm-Lrp1-/- mice showed ANP signaling activation that was linked to increased fatty acid (FA) uptake and increased AMPK/ ACC phosphorylation in the liver. Natriuretic peptide receptor A (NPR-A) antagonist completely abolished ANP signaling and metabolic protection in cm-Lrp1-/- mice. CONCLUSIONS: These results indicate that an ANP-dependent axis controlled by cardiac LRP1 levels modulates AMPK activity in the liver, energy homeostasis and whole-body metabolism.

Effects of Maternal Fructose Intake on Perinatal ER-Stress: A Defective XBP1s Nuclear Translocation Affects the ER-stress Resolution.

Endoplasmic reticulum (ER) homeostasis is crucial to appropriate cell functioning, and when disturbed, a safeguard system called unfolded protein response (UPR) is activated. Fructose consumption modifies ER homeostasis and has been related to metabolic syndrome. However, fructose sweetened beverages intake is allowed during gestation. Therefore, we investigate whether maternal fructose intake affects the ER status and induces UPR. Thus, administrating liquid fructose (10% w/v) to pregnant rats partially activated the ER-stress in maternal and fetal liver and placenta. In fact, a fructose-induced increase in the levels of pIRE1 (phosphorylated inositol requiring enzyme-1) and its downstream effector, X-box binding protein-1 spliced form (XBP1s), was observed. XBP1s is a key transcription factor, however, XBP1s nuclear translocation and the expression of its target genes were reduced in the liver of the carbohydrate-fed mothers, and specifically diminished in the fetal liver and placenta in the fructose-fed mothers. These XBP1s target genes belong to the ER-associated protein degradation (ERAD) system, used to buffer ER-stress and to restore ER-homeostasis. It is known that XBP1s needs to form a complex with diverse proteins to migrate into the nucleus. Since methylglyoxal (MGO) content, a precursor of advanced glycation endproducts (AGE), was augmented in the three tissues in the fructose-fed mothers and has been related to interfere with the functioning of many proteins, the role of MGO in XBP1s migration should not be discarded. In conclusion, maternal fructose intake produces ER-stress, but without XBP1s nuclear migration. Therefore, a complete activation of UPR that would resolve ER-stress is lacking. A state of fructose-induced oxidative stress is probably involved.

mTOR is a Key Protein Involved in the Metabolic Effects of Simple Sugars.

One of the most important threats to global human health is the increasing incidences of metabolic pathologies (including obesity, type 2 diabetes and non-alcoholic fatty liver disease), which is paralleled by increasing consumptions of hypercaloric diets enriched in simple sugars. The challenge is to identify the metabolic pathways affected by the excessive consumption of these dietary components when they are consumed in excess, to unravel the molecular mechanisms leading to metabolic pathologies and identify novel therapeutic targets to manage them. Mechanistic (mammalian) target of rapamycin (mTOR) has emerged as one of the key molecular nodes that integrate extracellular signals, such as energy status and nutrient availability, to trigger cell responses that could lead to the above-mentioned diseases through the regulation of lipid and glucose metabolism. By activating mTOR signalling, excessive consumption of simple sugars (such as fructose and glucose), could modulate hepatic gluconeogenesis, lipogenesis and fatty acid uptake and catabolism and thus lipid deposition in the liver. In the present review we will discuss some of the most recent studies showing the central role of mTOR in the metabolic effects of excessive simple sugar consumption.

Chronic fructose intake does not induce liver steatosis and inflammation in female Sprague-Dawley rats, but causes hypertriglyceridemia related to decreased VLDL receptor expression.

PURPOSE: Sugar-sweetened beverage intake is a risk factor for insulin resistance, dyslipidemia, fatty liver, and steatohepatitis (NASH). Sub-chronic supplementation of liquid fructose, but not glucose, in female rats increases liver and plasma triglycerides without inflammation. We hypothesized that chronic supplementation of fructose would cause NASH and liver insulin resistance. METHODS: We supplemented female Sprague-Dawley rats with water or either fructose or glucose 10% w/v solutions under isocaloric conditions for 7 months. At the end, plasma analytes, insulin, and adiponectin were determined, as well as liver triglyceride content and the expression of key genes controlling inflammation, fatty acid synthesis and oxidation, endoplasmic reticulum stress, and plasma VLDL clearance, by biochemical and histological methods. RESULTS: Although sugar-supplemented rats increased their energy intake by 50-60%, we found no manifestation of liver steatosis, fibrosis or necrosis, unchanged plasma or tissue markers of inflammation or fibrosis, and reduced liver expression of gluconeogenic enzymes, despite both sugars increased fatty acid synthesis, mTORC1, and IRE1 activity, while decreasing fatty acid oxidation and PPARα activity. Only fructose-supplemented rats were hypertriglyceridemic, showing a reduced expression of VLDL receptor and lipoprotein lipase in skeletal muscle and vWAT. Glucose-supplemented rats showed increased adiponectinemia, which would explain the different metabolic outcomes of the two sugars. CONCLUSIONS: Chronic liquid simple sugar supplementation, as the sole risk factor, is not enough for female rats to develop NASH and increased liver gluconeogenesis. Nevertheless, under isocaloric conditions, only fructose induced hypertriglyceridemia, thus confirming that also the type of nutrient matters in the development of metabolic diseases.

Chronic Liquid Fructose, but not Glucose, Supplementation Selectively Induces Visceral Adipose Tissue Leptin Resistance and Hypertrophy in Female Sprague-Dawley Rats.

SCOPE: The effect of chronic supplementation with simple-sugar solutions on leptin signaling in liver, hypothalamus, and visceral white adipose tissue (vWAT) is studied, which is designed to mimic the temporal pattern of consumption by humans. METHODS AND RESULTS: Solutions of fructose or glucose are isocalorically supplemented (7 months) in female Sprague-Dawley rats consuming ad libitum rodent chow. After sacrifice, plasma and tissue samples (liver, hypothalamus, and vWAT) are collected. Zoometric parameters, plasma analytes, and the tissue expression and activity of markers of leptin signaling are determined by biochemical and molecular biological methods. The two sugars cause different types of adiposopathy. Both sugars induce increases in plasma nonesterified fatty acids, and leptin resistance in the liver and the hypothalamus. Only fructose-supplemented rats show hyperleptinemia, and increased body weight due to a hypertrophy of vWAT, with no signs of leptin-mediated lipolysis. Glucose-supplemented rats show no significant changes in these parameters but present elevated plasma adiponectin concentrations, lipolysis, and inflammatory markers in vWAT, indicating a shift to a nonexpandable adipose tissue phenotype. CONCLUSION: Chronic consumption of fructose places a greater burden on metabolic homeostasis than equivalent consumption of glucose, inducing hyperleptinemia, generalized leptin resistance, and increased body weight due to expanded, hypertrophic vWAT.

Impairment of Novel Object Recognition Memory and Brain Insulin Signaling in Fructose- but Not Glucose-Drinking Female Rats.

Excessive sugar intake has been related to cognitive alterations, but it remains unclear whether these effects are related exclusively to increased energy intake, and the molecular mechanisms involved are not fully understood. We supplemented Sprague-Dawley female rats with 10% w/v fructose in drinking water or with isocaloric glucose solution for 7 months. Cognitive function was assessed through the Morris water maze (MWM) and the novel object recognition (NOR) tests. Plasma parameters and protein/mRNA expression in the frontal cortex and hippocampus were determined. Results showed that only fructose-supplemented rats displayed postprandial and fasting hypertriglyceridemia (1.4 and 1.9-fold, p < 0.05) and a significant reduction in the discrimination index in the NOR test, whereas the results of the MWM test showed no differences between groups. Fructose-drinking rats displayed an abnormal glucose tolerance test and impaired insulin signaling in the frontal cortex, as revealed by significant reductions in insulin receptor substrate-2 protein levels (0.77-fold, p < 0.05) and Akt phosphorylation (0.72-fold, p < 0.05), and increased insulin-degrading enzyme levels (1.86-fold, p < 0.001). Fructose supplementation reduced the expression of antioxidant enzymes and altered the amount of proteins involved in mitochondrial fusion/fission in the frontal cortex. In conclusion, cognitive deficits induced by chronic liquid fructose consumption are not exclusively related to increased caloric intake and are correlated with hypertriglyceridemia, impaired insulin signaling, increased oxidative stress and altered mitochondrial dynamics, especially in the frontal cortex.

Human hepatic lipase overexpression in mice induces hepatic steatosis and obesity through promoting hepatic lipogenesis and white adipose tissue lipolysis and fatty acid uptake.

Human hepatic lipase (hHL) is mainly localized on the hepatocyte cell surface where it hydrolyzes lipids from remnant lipoproteins and high density lipoproteins and promotes their hepatic selective uptake. Furthermore, hepatic lipase (HL) is closely associated with obesity in multiple studies. Therefore, HL may play a key role on lipid homeostasis in liver and white adipose tissue (WAT). In the present study, we aimed to evaluate the effects of hHL expression on hepatic and white adipose triglyceride metabolism in vivo. Experiments were carried out in hHL transgenic and wild-type mice fed a Western-type diet. Triglyceride metabolism studies included ß-oxidation and de novo lipogenesis in liver and WAT, hepatic triglyceride secretion, and adipose lipoprotein lipase (LPL)-mediated free fatty acid (FFA) lipolysis and influx. The expression of hHL promoted hepatic triglyceride accumulation and de novo lipogenesis without affecting triglyceride secretion, and this was associated with an upregulation of Srebf1 as well as the main genes controlling the synthesis of fatty acids. Transgenic mice also exhibited more adiposity and an increased LPL-mediated FFA influx into the WAT without affecting glucose tolerance. Our results demonstrate that hHL promoted hepatic steatosis in mice mainly by upregulating de novo lipogenesis. HL also upregulated WAT LPL and promoted triglyceride-rich lipoprotein hydrolysis and adipose FFA uptake. These data support the important role of hHL in regulating hepatic lipid homeostasis and confirm the broad cardiometabolic role of HL.

The Addition of Liquid Fructose to a Western-Type Diet in LDL-R-/- Mice Induces Liver Inflammation and Fibrogenesis Markers without Disrupting Insulin Receptor Signalling after an Insulin Challenge.

A high consumption of fat and simple sugars, especially fructose, has been related to the development of insulin resistance, but the mechanisms involved in the effects of these nutrients are not fully understood. This study investigates the effects of a Western-type diet and liquid fructose supplementation, alone and combined, on insulin signalling and inflammation in low-density lipoprotein (LDL) receptor-deficient mice (LDL-R-/-). LDL-R-/- mice were fed chow or Western diet ±15% fructose solution for 12 weeks. Plasma glucose and insulin, and the expression of genes related to inflammation in the liver and visceral white adipose tissue (vWAT), were analysed. V-akt murine thymoma viral oncogene homolog-2 (Akt) activation was measured in the liver of the mice after a single injection of saline or insulin. None of the dietary interventions caused inflammation in vWAT, whereas the Western diet induced hepatic inflammation, which was further enhanced by liquid fructose, leading also to a significant increase in fibrogenesis markers. However, there was no change in plasma glucose or insulin, or insulin-induced Akt phosphorylation. In conclusion, hepatic inflammation and fibrogenesis markers induced by a Western diet supplemented with liquid fructose in LDL-R-/- mice are not associated with a significant impairment of hepatic insulin signalling.