Soat2 ties cholesterol metabolism to ß-oxidation and glucose tolerance in male mice.

BACKGROUND: Sterol O-acyltransferase 2 (Soat2) encodes acyl-coenzyme A:cholesterol acyltransferase 2 (ACAT2), which synthesizes cholesteryl esters in hepatocytes and enterocytes fated either to storage or to secretion into nascent triglyceride-rich lipoproteins. OBJECTIVES: We aimed to unravel the molecular mechanisms leading to reduced hepatic steatosis when Soat2 is depleted in mice. METHODS: Soat2-/- and wild-type mice were fed a high-fat, a high-carbohydrate, or a chow diet, and parameters of lipid and glucose metabolism were assessed. RESULTS: Glucose, insulin, homeostatic model assessment for insulin resistance (HOMA-IR), oral glucose tolerance (OGTT), and insulin tolerance tests significantly improved in Soat2-/- mice, irrespective of the dietary regimes (2-way ANOVA). The significant positive correlations between area under the curve (AUC) OGTT (r = 0.66, p < 0.05), serum fasting insulin (r = 0.86, p < 0.05), HOMA-IR (r = 0.86, p < 0.05), Adipo-IR (0.87, p < 0.05), hepatic triglycerides (TGs) (r = 0.89, p < 0.05), very-low-density lipoprotein (VLDL)-TG (r = 0.87, p < 0.05) and the hepatic cholesteryl esters in wild-type mice disappeared in Soat2-/- mice. Genetic depletion of Soat2 also increased whole-body oxidation by 30% (p < 0.05) compared to wild-type mice. CONCLUSION: Our data demonstrate that ACAT2-generated cholesteryl esters negatively affect the metabolic control by retaining TG in the liver and that genetic inhibition of Soat2 improves liver steatosis via partitioning of lipids into secretory (VLDL-TG) and oxidative (fatty acids) pathways.

Ghrelin treatment induces rapid and delayed increments of food intake: a heuristic model to explain ghrelin's orexigenic effects.

Ghrelin is a stomach-derived peptide hormone with salient roles in the regulation of energy balance and metabolism. Notably, ghrelin is recognized as the most powerful known circulating orexigenic hormone. Here, we systematically investigated the effects of ghrelin on energy homeostasis and found that ghrelin primarily induces a biphasic effect on food intake that has indirect consequences on energy expenditure and nutrient partitioning. We also found that ghrelin-induced biphasic effect on food intake requires the integrity of Agouti-related peptide/neuropeptide Y-producing neurons of the hypothalamic arcuate nucleus, which seem to display a long-lasting activation after a single systemic injection of ghrelin. Finally, we found that different autonomic, hormonal and metabolic satiation signals transiently counteract ghrelin-induced food intake. Based on our observations, we propose a heuristic model to describe how the orexigenic effect of ghrelin and the anorectic food intake-induced rebound sculpt a timely constrain feeding response to ghrelin.

Insights From Liver-Humanized Mice on Cholesterol Lipoprotein Metabolism and LXR-Agonist Pharmacodynamics in Humans.

BACKGROUND AND AIMS: Genetically modified mice have been used extensively to study human disease. However, the data gained are not always translatable to humans because of major species differences. Liver-humanized mice (LHM) are considered a promising model to study human hepatic and systemic metabolism. Therefore, we aimed to further explore their lipoprotein metabolism and to characterize key hepatic species-related, physiological differences. APPROACH AND RESULTS: Fah-/- , Rag2-/- , and Il2rg-/- knockout mice on the nonobese diabetic (FRGN) background were repopulated with primary human hepatocytes from different donors. Cholesterol lipoprotein profiles of LHM showed a human-like pattern, characterized by a high ratio of low-density lipoprotein to high-density lipoprotein, and dependency on the human donor. This pattern was determined by a higher level of apolipoprotein B100 in circulation, as a result of lower hepatic mRNA editing and low-density lipoprotein receptor expression, and higher levels of circulating proprotein convertase subtilisin/kexin type 9. As a consequence, LHM lipoproteins bind to human aortic proteoglycans in a pattern similar to human lipoproteins. Unexpectedly, cholesteryl ester transfer protein was not required to determine the human-like cholesterol lipoprotein profile. Moreover, LHM treated with GW3965 mimicked the negative lipid outcomes of the first human trial of liver X receptor stimulation (i.e., a dramatic increase of cholesterol and triglycerides in circulation). Innovatively, LHM allowed the characterization of these effects at a molecular level. CONCLUSIONS: LHM represent an interesting translatable model of human hepatic and lipoprotein metabolism. Because several metabolic parameters displayed donor dependency, LHM may also be used in studies for personalized medicine.

Central CCL2 signaling onto MCH neurons mediates metabolic and behavioral adaptation to inflammation.

Sickness behavior defines the endocrine, autonomic, behavioral, and metabolic responses associated with infection. While inflammatory responses were suggested to be instrumental in the loss of appetite and body weight, the molecular underpinning remains unknown. Here, we show that systemic or central lipopolysaccharide (LPS) injection results in specific hypothalamic changes characterized by a precocious increase in the chemokine ligand 2 (CCL2) followed by an increase in pro-inflammatory cytokines and a decrease in the orexigenic neuropeptide melanin-concentrating hormone (MCH). We therefore hypothesized that CCL2 could be the central relay for the loss in body weight induced by the inflammatory signal LPS. We find that central delivery of CCL2 promotes neuroinflammation and the decrease in MCH and body weight. MCH neurons express CCL2 receptor and respond to CCL2 by decreasing both electrical activity and MCH release. Pharmacological or genetic inhibition of CCL2 signaling opposes the response to LPS at both molecular and physiologic levels. We conclude that CCL2 signaling onto MCH neurons represents a core mechanism that relays peripheral inflammation to sickness behavior.

Lipoprotein lipase in hypothalamus is a key regulator of body weight gain and glucose homeostasis in mice.

AIMS/HYPOTHESIS: Regulation of energy balance involves the participation of many factors, including nutrients, among which are circulating lipids, acting as peripheral signals informing the central nervous system of the energy status of the organism. It has been shown that neuronal lipoprotein lipase (LPL) participates in the control of energy balance by hydrolysing lipid particles enriched in triacylglycerols. Here, we tested the hypothesis that LPL in the mediobasal hypothalamus (MBH), a well-known nucleus implicated in the regulation of metabolic homeostasis, could also contribute to the regulation of body weight and glucose homeostasis. METHODS: We injected an adeno-associated virus (AAV) expressing Cre-green fluorescent protein into the MBH of Lpl-floxed mice (and wild-type mice) to specifically decrease LPL activity in the MBH. In parallel, we injected an AAV overexpressing Lpl into the MBH of wild-type mice. We then studied energy homeostasis and hypothalamic ceramide content. RESULTS: The partial deletion of Lpl in the MBH in mice led to an increase in body weight compared with controls (37.72 ± 0.7 g vs 28.46 ± 0.12, p < 0.001) associated with a decrease in locomotor activity. These mice developed hyperinsulinaemia and glucose intolerance. This phenotype also displayed reduced expression of Cers1 in the hypothalamus as well as decreased concentration of several C18 species of ceramides and a 3-fold decrease in total ceramide intensity. Conversely, overexpression of Lpl specifically in the MBH induced a decrease in body weight. CONCLUSIONS/INTERPRETATION: Our study shows that LPL in the MBH is an important regulator of body weight and glucose homeostasis.

Muscle expression of a malonyl-CoA-insensitive carnitine palmitoyltransferase-1 protects mice against high-fat/high-sucrose diet-induced insulin resistance.

Impaired skeletal muscle mitochondrial fatty acid oxidation (mFAO) has been implicated in the etiology of insulin resistance. Carnitine palmitoyltransferase-1 (CPT1) is a key regulatory enzyme of mFAO whose activity is inhibited by malonyl-CoA, a lipogenic intermediate. Whereas increasing CPT1 activity in vitro has been shown to exert a protective effect against lipid-induced insulin resistance in skeletal muscle cells, only a few studies have addressed this issue in vivo. We thus examined whether a direct modulation of muscle CPT1/malonyl-CoA partnership is detrimental or beneficial for insulin sensitivity in the context of diet-induced obesity. By using a Cre-LoxP recombination approach, we generated mice with skeletal muscle-specific and inducible expression of a mutated CPT1 form (CPT1mt) that is active but insensitive to malonyl-CoA inhibition. When fed control chow, homozygous CPT1mt transgenic (dbTg) mice exhibited decreased CPT1 sensitivity to malonyl-CoA inhibition in isolated muscle mitochondria, which was sufficient to substantially increase ex vivo muscle mFAO capacity and whole body fatty acid utilization in vivo. Moreover, dbTg mice were less prone to high-fat/high-sucrose (HFHS) diet-induced insulin resistance and muscle lipotoxicity despite similar body weight gain, adiposity, and muscle malonyl-CoA content. Interestingly, these CPT1mt-protective effects in dbTg-HFHS mice were associated with preserved muscle insulin signaling, increased muscle glycogen content, and upregulation of key genes involved in muscle glucose metabolism. These beneficial effects of muscle CPT1mt expression suggest that a direct modulation of the malonyl-CoA/CPT1 partnership in skeletal muscle could represent a potential strategy to prevent obesity-induced insulin resistance.

Hypothalamic AgRP-neurons control peripheral substrate utilization and nutrient partitioning.

Obesity-related diseases such as diabetes and dyslipidemia result from metabolic alterations including the defective conversion, storage and utilization of nutrients, but the central mechanisms that regulate this process of nutrient partitioning remain elusive. As positive regulators of feeding behaviour, agouti-related protein (AgRP) producing neurons are indispensible for the hypothalamic integration of energy balance. Here, we demonstrate a role for AgRP-neurons in the control of nutrient partitioning. We report that ablation of AgRP-neurons leads to a change in autonomic output onto liver, muscle and pancreas affecting the relative balance between lipids and carbohydrates metabolism. As a consequence, mice lacking AgRP-neurons become obese and hyperinsulinemic on regular chow but display reduced body weight gain and paradoxical improvement in glucose tolerance on high-fat diet. These results provide a direct demonstration of a role for AgRP-neurons in the coordination of efferent organ activity and nutrient partitioning, providing a mechanistic link between obesity and obesity-related disorders.

Intestinal deletion of leptin signaling alters activity of nutrient transporters and delayed the onset of obesity in mice.

The importance of B-isoform of leptin receptor (LEPR-B) signaling in the hypothalamus, pancreas, or liver has been well characterized, but in the intestine, a unique site of entry for dietary nutrition into the body, it has been relatively ignored. To address this question, we characterized a mouse model deficient for LEPR-B specifically in intestinal epithelial cells (IECs). (IEC)LEPR-B-knockout (KO) and wild-type (WT) mice were generated by Cre-Lox strategy and fed a normal or high-fat diet (HFD). The analyses of the animals involved histology and immunohistochemistry of intestinal mucosa, indirect calorimetric measurements, whole-body composition, and expression and activities of nutrient transporters. (IEC)LEPR-B-KO mice exhibited a 2-fold increase in length of jejunal villi and have normal growth on a normal diet but were less susceptible (P<0.01) to HFD-induced obesity. No differences occurred in energy intake and expenditure between (IEC)LEPR-B-WT and -KO mice, but (IEC)LEPR-B-KO mice fed an HFD showed increased excreted fats (P<0.05). Activities of the Na(+)/glucose cotransporter SGLT-1 and GLUT2 were unaffected in LEPR-B-KO jejunum, while GLUT5-mediated fructose transport and PepT1-mediated peptide transport were substantially reduced (P<0.01). These data demonstrate that intestinal LEPR-B signaling is important for the onset of diet-induced obesity. They suggest that intestinal LEPR-B could be a potential per os target for prevention against obesity.

Myostatin is a key mediator between energy metabolism and endurance capacity of skeletal muscle.

Myostatin (Mstn) participates in the regulation of skeletal muscle size and has emerged as a regulator of muscle metabolism. Here, we hypothesized that lack of myostatin profoundly depresses oxidative phosphorylation-dependent muscle function. Toward this end, we explored Mstn(-/-) mice as a model for the constitutive absence of myostatin and AAV-mediated overexpression of myostatin propeptide as a model of myostatin blockade in adult wild-type mice. We show that muscles from Mstn(-/-) mice, although larger and stronger, fatigue extremely rapidly. Myostatin deficiency shifts muscle from aerobic toward anaerobic energy metabolism, as evidenced by decreased mitochondrial respiration, reduced expression of PPAR transcriptional regulators, increased enolase activity, and exercise-induced lactic acidosis. As a consequence, constitutively reduced myostatin signaling diminishes exercise capacity, while the hypermuscular state of Mstn(-/-) mice increases oxygen consumption and the energy cost of running. We wondered whether these results are the mere consequence of the congenital fiber-type switch toward a glycolytic phenotype of constitutive Mstn(-/-) mice. Hence, we overexpressed myostatin propeptide in adult mice, which did not affect fiber-type distribution, while nonetheless causing increased muscle fatigability, diminished exercise capacity, and decreased Pparb/d and Pgc1a expression. In conclusion, our results suggest that myostatin endows skeletal muscle with high oxidative capacity and low fatigability, thus regulating the delicate balance between muscle mass, muscle force, energy metabolism, and endurance capacity.

Whole-body deletion of Endospanin 1 protects from obesity-associated deleterious metabolic alterations.

The importance of the proper localization of most receptors at the cell surface is often underestimated, although this feature is essential for optimal receptor response. Endospanin 1 (Endo1) (also known as OBRGRP or LEPROT) is a protein generated from the same gene as the human leptin receptor and regulates the trafficking of proteins to the surface, including the leptin receptor. The systemic role of Endo1 on whole-body metabolism has not been studied so far. Here, we report that general Endo1-KO mice fed a high-fat diet develop metabolically healthy obesity with lipid repartitioning in organs and preferential accumulation of fat in adipose tissue, limited systematic inflammation, and better controlled glucose homeostasis. Mechanistically, Endo1 interacts with the lipid translocase CD36, thus regulating its surface abundance and lipid uptake in adipocytes. In humans, the level of Endo1 transcripts is increased in the adipose tissue of patients with obesity, but low levels rather correlate with a profile of metabolically healthy obesity. We suggest here that Endo1, most likely by controlling CD36 cell surface abundance and lipid uptake in adipocytes, dissociates obesity from diabetes and that its absence participates in metabolically healthy obesity.

Laforin, a dual specificity phosphatase involved in Lafora disease, regulates insulin response and whole-body energy balance in mice.

Laforin is a dual specificity protein phosphatase involved in Lafora disease (LD), a fatal form of progressive myoclonus epilepsy characterized by neurodegeneration and the presence of intracellular polyglucosan inclusions (Lafora bodies) in different tissues. In this work, we describe that mice lacking laforin (epm2a-/-) have enhanced insulin response leading to altered whole-body energy balance. This enhanced insulin response overactivates the Akt pathway which increases glucose uptake in the heart, resulting in increased glycogen levels and the formation of polyglucosan inclusions. In addition, enhanced insulin response resulted in increased liver lipid biosynthesis, resulting in hepatic steatosis. On the contrary, overexpression in rat hepatoma FTO2B cells of native laforin but not of a form lacking phosphatase activity (C266S) resulted in attenuation of insulin signaling. These results define laforin as a new regulator of insulin sensitivity, which provides novel insights into LD pathogenesis and identifies this phosphatase as a potential novel component of the insulin signaling cascade.

Beige differentiation of adipose depots in mice lacking prolactin receptor protects against high-fat-diet-induced obesity.

Stimulating conversion of white fat to metabolically active adipocytes (beige fat) constitutes a promising strategy against weight gain and its deleterious associated-disorders. We provide direct evidence that prolactin (PRL), best known for its actions on the mammary gland, plays a pivotal role in energy balance through the control of adipocyte differentiation and fate. Here we show that lack of prolactin receptor (PRLR) causes resistance to high-fat-diet-induced obesity due to enhanced energy expenditure and increased metabolic rate. Mutant mice displayed reduced fat mass associated with appearance of massive brown-like adipocyte foci in perirenal and subcutaneous but not in gonadal fat depots under a high-fat diet. Positron emission tomography imaging further demonstrated the occurrence of these thermogenic brown fat depots in adult mice, providing additional support for recruitable brown-like adipocytes (beigeing) in white fat depots. Consistent with the activation of brown adipose tissue, PRLR inactivation increases expression of master genes controlling brown adipocyte fate (PRDM16) and mitochondrial function (PGC1α, UCP1). Altered pRb/Foxc2 expression suggests that this PRL-regulated pathway may contribute to beige cell commitment. Together, these results provide direct genetic evidence that PRLR affects energy balance and metabolic adaptation in rodents via effects on brown adipose tissue differentiation and function.

Mice with humanized livers reveal the role of hepatocyte clocks in rhythmic behavior.

The synchronization of circadian clock depends on a central pacemaker located in the suprachiasmatic nuclei. However, the potential feedback of peripheral signals on the central clock remains poorly characterized. To explore whether peripheral organ circadian clocks may affect the central pacemaker, we used a chimeric model in which mouse hepatocytes were replaced by human hepatocytes. Liver humanization led to reprogrammed diurnal gene expression and advanced the phase of the liver circadian clock that extended to muscle and the entire rhythmic physiology. Similar to clock-deficient mice, liver-humanized mice shifted their rhythmic physiology more rapidly to the light phase under day feeding. Our results indicate that hepatocyte clocks can affect the central pacemaker and offer potential perspectives to apprehend pathologies associated with altered circadian physiology.

CRISPR/Cas9-mediated inactivation of the phosphatase activity of soluble epoxide hydrolase prevents obesity and cardiac ischemic injury.

INTRODUCTION: Although the physiological role of the C-terminal hydrolase domain of the soluble epoxide hydrolase (sEH-H) is well investigated, the function of its N-terminal phosphatase activity (sEH-P) remains unknown. OBJECTIVES: This study aimed to assess in vivo the physiological role of sEH-P. METHODS: CRISPR/Cas9 was used to generate a novel knock-in (KI) rat line lacking the sEH-P activity. RESULTS: The sEH-P KI rats has a decreased metabolism of lysophosphatidic acids to monoacyglycerols. KI rats grew almost normally but with less weight and fat mass gain while insulin sensitivity was increased compared to wild-type rats. This lean phenotype was more marked in males than in female KI rats and mainly due to decreased food consumption and enhanced energy expenditure. In fact, sEH-P KI rats had an increased lipolysis allowing to supply fatty acids as fuel to potentiate brown adipose thermogenesis under resting condition and upon cold exposure. The potentiation of thermogenesis was abolished when blocking PPARγ, a nuclear receptor activated by intracellular lysophosphatidic acids, but also when inhibiting simultaneously sEH-H, showing a functional interaction between the two domains. Furthermore, sEH-P KI rats fed a high-fat diet did not gain as much weight as the wild-type rats, did not have increased fat mass and did not develop insulin resistance or hepatic steatosis. In addition, sEH-P KI rats exhibited enhanced basal cardiac mitochondrial activity associated with an enhanced left ventricular contractility and were protected against cardiac ischemia-reperfusion injury. CONCLUSION: Our study reveals that sEH-P is a key player in energy and fat metabolism and contributes together with sEH-H to the regulation of cardiometabolic homeostasis. The development of pharmacological inhibitors of sEH-P appears of crucial importance to evaluate the interest of this promising therapeutic strategy in the management of obesity and cardiac ischemic complications.

Homocysteine Metabolism Pathway Is Involved in the Control of Glucose Homeostasis: A Cystathionine Beta Synthase Deficiency Study in Mouse.

Cystathionine beta synthase (CBS) catalyzes the first step of the transsulfuration pathway from homocysteine to cystathionine, and its deficiency leads to hyperhomocysteinemia (HHcy) in humans and rodents. To date, scarce information is available about the HHcy effect on insulin secretion, and the link between CBS activity and the setting of type 2 diabetes is still unknown. We aimed to decipher the consequences of an inborn defect in CBS on glucose homeostasis in mice. We used a mouse model heterozygous for CBS (CBS+/-) that presented a mild HHcy. Other groups were supplemented with methionine in drinking water to increase the mild to intermediate HHcy, and were submitted to a high-fat diet (HFD). We measured the food intake, body weight gain, body composition, glucose homeostasis, plasma homocysteine level, and CBS activity. We evidenced a defect in the stimulated insulin secretion in CBS+/- mice with mild and intermediate HHcy, while mice with intermediate HHcy under HFD presented an improvement in insulin sensitivity that compensated for the decreased insulin secretion and permitted them to maintain a glucose tolerance similar to the CBS+/+ mice. Islets isolated from CBS+/- mice maintained their ability to respond to the elevated glucose levels, and we showed that a lower parasympathetic tone could, at least in part, be responsible for the insulin secretion defect. Our results emphasize the important role of Hcy metabolic enzymes in insulin secretion and overall glucose homeostasis.

Tanycytes control hypothalamic liraglutide uptake and its anti-obesity actions.

Liraglutide, an anti-diabetic drug and agonist of the glucagon-like peptide one receptor (GLP1R), has recently been approved to treat obesity in individuals with or without type 2 diabetes. Despite its extensive metabolic benefits, the mechanism and site of action of liraglutide remain unclear. Here, we demonstrate that liraglutide is shuttled to target cells in the mouse hypothalamus by specialized ependymoglial cells called tanycytes, bypassing the blood-brain barrier. Selectively silencing GLP1R in tanycytes or inhibiting tanycytic transcytosis by botulinum neurotoxin expression not only hampers liraglutide transport into the brain and its activation of target hypothalamic neurons, but also blocks its anti-obesity effects on food intake, body weight and fat mass, and fatty acid oxidation. Collectively, these striking data indicate that the liraglutide-induced activation of hypothalamic neurons and its downstream metabolic effects are mediated by its tanycytic transport into the mediobasal hypothalamus, strengthening the notion of tanycytes as key regulators of metabolic homeostasis.

Hypothalamic astrocytes control systemic glucose metabolism and energy balance.

The hypothalamus is key in the control of energy balance. However, strategies targeting hypothalamic neurons have failed to provide viable options to treat most metabolic diseases. Conversely, the role of astrocytes in systemic metabolic control has remained largely unexplored. Here, we show that obesity promotes anatomically restricted remodeling of hypothalamic astrocyte activity. In the paraventricular nucleus (PVN) of the hypothalamus, chemogenetic manipulation of astrocytes results in bidirectional control of neighboring neuron activity, autonomic outflow, glucose metabolism, and energy balance. This process recruits a mechanism involving the astrocytic control of ambient glutamate levels, which becomes defective in obesity. Positive or negative chemogenetic manipulation of PVN astrocyte Ca2+ signals, respectively, worsens or improves metabolic status of diet-induced obese mice. Collectively, these findings highlight a yet unappreciated role for astrocytes in the direct control of systemic metabolism and suggest potential targets for anti-obesity strategy.

Acute changes in systemic glycemia gate access and action of GLP-1R agonist on brain structures controlling energy homeostasis.

Therapies based on glucagon-like peptide-1 (GLP-1) long-acting analogs and insulin are often used in the treatment of metabolic diseases. Both insulin and GLP-1 receptors are expressed in metabolically relevant brain regions, suggesting a cooperative action. However, the mechanisms underlying the synergistic actions of insulin and GLP-1R agonists remain elusive. In this study, we show that insulin-induced hypoglycemia enhances GLP-1R agonists entry in hypothalamic and area, leading to enhanced whole-body fat oxidation. Mechanistically, this phenomenon relies on the release of tanycyctic vascular endothelial growth factor A, which is selectively impaired after calorie-rich diet exposure. In humans, low blood glucose also correlates with enhanced blood-to-brain passage of insulin, suggesting that blood glucose gates the passage other energy-related signals in the brain. This study implies that the preventing hyperglycemia is important to harnessing the full benefit of GLP-1R agonist entry in the brain and action onto lipid mobilization and body weight loss.

Further Evidence that Habitual Consumption of Sucralose with, but Not without, Carbohydrate Alters Glucose Metabolism.

In this letter, Dalenberg et al. provide a point-by-point response to the critique offered by Kahn and Sievenpiper. They also offer new evidence to support their original finding from a study they conducted in mice.