Interleukin-33-Activated Islet-Resident Innate Lymphoid Cells Promote Insulin Secretion through Myeloid Cell Retinoic Acid Production.

Pancreatic-islet inflammation contributes to the failure of ß cell insulin secretion during obesity and type 2 diabetes. However, little is known about the nature and function of resident immune cells in this context or in homeostasis. Here we show that interleukin (IL)-33 was produced by islet mesenchymal cells and enhanced by a diabetes milieu (glucose, IL-1ß, and palmitate). IL-33 promoted ß cell function through islet-resident group 2 innate lymphoid cells (ILC2s) that elicited retinoic acid (RA)-producing capacities in macrophages and dendritic cells via the secretion of IL-13 and colony-stimulating factor 2. In turn, local RA signaled to the ß cells to increase insulin secretion. This IL-33-ILC2 axis was activated after acute ß cell stress but was defective during chronic obesity. Accordingly, IL-33 injections rescued islet function in obese mice. Our findings provide evidence that an immunometabolic crosstalk between islet-derived IL-33, ILC2s, and myeloid cells fosters insulin secretion.

A 30-color spectral flow cytometry panel for comprehensive analysis of immune cell composition and macrophage subsets in mouse metabolic organs.

Obesity-induced chronic low-grade inflammation, also known as metaflammation, results from alterations of the immune response in metabolic organs and contributes to the development of fatty liver diseases and type 2 diabetes. The diversity of tissue-resident leukocytes involved in these metabolic dysfunctions warrants an in-depth immunophenotyping in order to elucidate disease etiology. Here, we present a 30-color, full spectrum flow cytometry panel, designed to (i) identify the major innate and adaptive immune cell subsets in murine liver and white adipose tissues and (ii) discriminate various tissue-specific myeloid subsets known to contribute to the development of metabolic dysfunctions. This panel notably allows for distinguishing embryonically-derived liver-resident Kupffer cells from newly recruited monocyte-derived macrophages and KCs. Furthermore, several adipose tissue macrophage (ATM) subsets, including perivascular macrophages, lipid-associated macrophages, and pro-inflammatory CD11c+ ATMs, can also be identified. Finally, the panel includes cell-surface markers that have been associated with metabolic activation of different macrophage and dendritic cell subsets. Altogether, our spectral flow cytometry panel allows for an extensive immunophenotyping of murine metabolic tissues, with a particular focus on metabolically-relevant myeloid cell subsets, and can easily be adjusted to include various new markers if needed.

Soluble mannose receptor induces proinflammatory macrophage activation and metaflammation.

Proinflammatory activation of macrophages in metabolic tissues is critically important in the induction of obesity-induced metaflammation. Here, we demonstrate that the soluble mannose receptor (sMR) plays a direct functional role in both macrophage activation and metaflammation. We show that sMR binds CD45 on macrophages and inhibits its phosphatase activity, leading to an Src/Akt/NF-κB-mediated cellular reprogramming toward an inflammatory phenotype both in vitro and in vivo. Remarkably, increased serum sMR levels were observed in obese mice and humans and directly correlated with body weight. Importantly, enhanced sMR levels increase serum proinflammatory cytokines, activate tissue macrophages, and promote insulin resistance. Altogether, our results reveal sMR as regulator of proinflammatory macrophage activation, which could constitute a therapeutic target for metaflammation and other hyperinflammatory diseases.

The helminth glycoprotein omega-1 improves metabolic homeostasis in obese mice through type 2 immunity-independent inhibition of food intake.

Type 2 immunity plays an essential role in the maintenance of metabolic homeostasis and its disruption during obesity promotes meta-inflammation and insulin resistance. Infection with the helminth parasite Schistosoma mansoni and treatment with its soluble egg antigens (SEA) induce a type 2 immune response in metabolic organs and improve insulin sensitivity and glucose tolerance in obese mice, yet, a causal relationship remains unproven. Here, we investigated the effects and underlying mechanisms of the T2 ribonuclease omega-1 (ω1), one of the major S mansoni immunomodulatory glycoproteins, on metabolic homeostasis. We show that treatment of obese mice with plant-produced recombinant ω1, harboring similar glycan motifs as present on the native molecule, decreased body fat mass, and improved systemic insulin sensitivity and glucose tolerance in a time- and dose-dependent manner. This effect was associated with an increase in white adipose tissue (WAT) type 2 T helper cells, eosinophils, and alternatively activated macrophages, without affecting type 2 innate lymphoid cells. In contrast to SEA, the metabolic effects of ω1 were still observed in obese STAT6-deficient mice with impaired type 2 immunity, indicating that its metabolic effects are independent of the type 2 immune response. Instead, we found that ω1 inhibited food intake, without affecting locomotor activity, WAT thermogenic capacity or whole-body energy expenditure, an effect also occurring in leptin receptor-deficient obese and hyperphagic db/db mice. Altogether, we demonstrate that while the helminth glycoprotein ω1 can induce type 2 immunity, it improves whole-body metabolic homeostasis in obese mice by inhibiting food intake via a STAT6-independent mechanism.

TOTUM-63, a plant-based polyphenol-rich extract, improves glycaemic control in subjects with prediabetes or early stage newly-diagnosed type 2 diabetes in a randomized, double-blind, placebo-controlled trial.

AIM: The plant-based polyphenol-rich extract TOTUM-63 improves glucose homeostasis in various preclinical models of obesity and type 2 diabetes (T2D). A pilot exploratory study showed that TOTUM-63 has good safety and tolerability profiles, and beneficial effects on postprandial glucose control in healthy individuals with overweight. The aim of this study was to assess the effects of TOTUM-63 on glycaemic control in individuals with prediabetes or early stage newly-diagnosed T2D (which does not require pharmacological treatment). MATERIALS AND METHODS: This study was a multicentre, randomized, double-blind, placebo-controlled trial. Individuals with prediabetes or early stage newly-diagnosed T2D and with overweight/abdominal obesity received TOTUM-63 (5 g/day) or placebo for 6 months. The primary outcome was the change in fasting blood glucose. RESULTS: Fifty-one participants (age: 57.1 ± 10 years; body mass index: 31.3 ± 5.7 kg.m2 ; 35 women and 16 men) completed the study (n = 38 TOTUM-63, n = 13 placebo). After 6 months, blood glucose concentration after fasting and after the 2-h oral glucose tolerance test was reduced in the TOTUM-63-treated group compared with the placebo group (placebo-corrected difference between baseline and month 6: -0.71 mmol/L, p < .05, and -1.93 mmol/L, p < .05, respectively). TOTUM-63 was safe and well tolerated and significantly reduced body weight gain (-1.9 kg; p < .05), waist circumference (-4.5 cm; p < .001), circulating triglycerides (-0.54 mmol/L; p < .01) and low-density lipoprotein-cholesterol (-0.38 mmol/L; p < .05) compared with placebo. CONCLUSIONS: TOTUM-63 lowered fasting blood glucose in participants with impaired fasting glycaemia and glucose intolerance. Moreover, TOTUM-63 showed a good safety and tolerability profile and improved several metabolic syndrome features. Therefore, TOTUM-63 is a promising candidate for T2D prevention.

Glucose availability but not changes in pancreatic hormones sensitizes hepatic AMPK activity during nutritional transition in rodents.

The cellular energy sensor AMP-activated protein kinase (AMPK) is a metabolic regulator that mediates adaptation to nutritional variations to maintain a proper energy balance in cells. We show here that suckling-weaning and fasting-refeeding transitions in rodents are associated with changes in AMPK activation and the cellular energy state in the liver. These nutritional transitions were characterized by a metabolic switch from lipid to glucose utilization, orchestrated by modifications in glucose levels and the glucagon/insulin ratio in the bloodstream. We therefore investigated the respective roles of glucose and pancreatic hormones on AMPK activation in mouse primary hepatocytes. We found that glucose starvation transiently activates AMPK, whereas changes in glucagon and insulin levels had no impact on AMPK. Challenge of hepatocytes with metformin-induced metabolic stress strengthened both AMPK activation and cellular energy depletion under limited-glucose conditions, whereas neither glucagon nor insulin altered AMPK activation. Although both insulin and glucagon induced AMPKα phosphorylation at its Ser485/491 residue, they did not affect its activity. Finally, the decrease in cellular ATP levels in response to an energy stress was additionally exacerbated under fasting conditions and by AMPK deficiency in hepatocytes, revealing metabolic inflexibility and emphasizing the importance of AMPK for maintaining hepatic energy charge. Our results suggest that nutritional changes (i.e. glucose availability), rather than the related hormonal changes (i.e. the glucagon/insulin ratio), sensitize AMPK activation to the energetic stress induced by the dietary transition during fasting. This effect is critical for preserving the cellular energy state in the liver.

Effects of Totum-63 on glucose homeostasis and postprandial glycemia: a translational study.

Global prevalence of type 2 diabetes (T2D) is rising and may affect 700 million people by 2045. Totum-63 is a polyphenol-rich natural composition developed to reduce the risk of T2D. We first investigated the effects of Totum-63 supplementation in high-fat diet (HFD)-fed mice for up to 16 wk and thereafter assessed its safety and efficacy (2.5 g or 5 g per day) in 14 overweight men [mean age 51.5 yr, body mass index (BMI) 27.6 kg·m-2] for 4 wk. In HFD-fed mice, Totum-63 reduced body weight and fat mass gain, whereas lean mass was unchanged. Moreover, fecal energy excretion was higher in Totum-63-supplemented mice, suggesting a reduction of calorie absorption in the digestive tract. In the gut, metagenomic analyses of fecal microbiota revealed a partial restoration of HFD-induced microbial imbalance, as shown by principal coordinate analysis of microbiota composition. HFD-induced increase in HOMA-IR score was delayed in supplemented mice, and insulin response to an oral glucose tolerance test was significantly reduced, suggesting that Totum-63 may prevent HFD-related impairments in glucose homeostasis. Interestingly, these improvements could be linked to restored insulin signaling in subcutaneous adipose tissue and soleus muscle. In the liver, HFD-induced steatosis was reduced by 40% (as shown by triglyceride content). In the subsequent study in men, Totum-63 (5 g·day-1) improved glucose and insulin responses to a high-carbohydrate breakfast test (84% kcal carbohydrates). It was well tolerated, with no clinically significant adverse events reported. Collectively, these data suggest that Totum-63 could improve glucose homeostasis in both HFD-fed mice and overweight individuals, presumably through a multitargeted action on different metabolic organs.NEW & NOTEWORTHY Totum-63 is a novel polyphenol-rich natural composition developed to reduce the risk of T2D. Totum-63 showed beneficial effects on glucose homeostasis in HFD-fed mice, presumably through a multitargeted action on different metabolic organs. Totum-63 was well tolerated in humans and improved postprandial glucose and insulin responses to a high-carbohydrate breakfast test.

Effects of a novel polyphenol-rich plant extract on body composition, inflammation, insulin sensitivity, and glucose homeostasis in obese mice.

BACKGROUND/OBJECTIVES: The worldwide prevalence of obesity, metabolic syndrome and type 2 diabetes (T2D) is reaching epidemic proportions that urge the development of new management strategies. Totum-63 is a novel, plant-based polyphenol-rich active principle that has been shown to reduce body weight, fasting glycemia, glucose intolerance, and fatty liver index in obese subjects with prediabetes. Here, we investigated the effects and underlying mechanism(s) of Totum-63 on metabolic homeostasis in insulin-resistant obese mice. METHODS: Male C57Bl6/J mice were fed a high-fat diet for 12 weeks followed by supplementation with Totum-63 for 4 weeks. The effects on whole-body energy and metabolic homeostasis, as well as on tissue-specific inflammation and insulin sensitivity were assessed using a variety of immunometabolic phenotyping tools. RESULTS: Totum-63 decreased body weight and fat mass in obese mice, without affecting lean mass, food intake and locomotor activity, and increased fecal energy excretion and whole-body fatty acid oxidation. Totum-63 reduced fasting plasma glucose, insulin and leptin levels, and improved whole-body insulin sensitivity and peripheral glucose uptake. The expression of insulin receptor ß and the insulin-induced phosphorylation of Akt/PKB were increased in liver, skeletal muscle, white adipose tissue (WAT) and brown adipose tissue (BAT). Hepatic steatosis was also decreased by Totum-63 and associated with a lower expression of genes involved in fatty acid uptake, de novo lipogenesis, inflammation, and fibrosis. Furthermore, a significant reduction in pro-inflammatory macrophages was also observed in epidydimal WAT. Finally, a potent decrease in BAT mass associated with enhanced tissue expression of thermogenic genes was found, suggesting BAT activation by Totum-63. CONCLUSIONS: Our results show that Totum-63 reduces inflammation and improves insulin sensitivity and glucose homeostasis in obese mice through pleiotropic effects on various metabolic organs. Altogether, plant-derived Totum-63 might constitute a promising novel nutritional supplement for alleviating metabolic dysfunctions in obese people with or without T2D.

Acetyl-CoA Carboxylase Inhibitor CP640.186 Increases Tubulin Acetylation and Impairs Thrombin-Induced Platelet Aggregation.

Acetyl-CoA carboxylase (ACC) is the first enzyme regulating de novo lipid synthesis via the carboxylation of acetyl-CoA into malonyl-CoA. The inhibition of its activity decreases lipogenesis and, in parallel, increases the acetyl-CoA content, which serves as a substrate for protein acetylation. Several findings support a role for acetylation signaling in coordinating signaling systems that drive platelet cytoskeletal changes and aggregation. Therefore, we investigated the impact of ACC inhibition on tubulin acetylation and platelet functions. Human platelets were incubated 2 h with CP640.186, a pharmacological ACC inhibitor, prior to thrombin stimulation. We have herein demonstrated that CP640.186 treatment does not affect overall platelet lipid content, yet it is associated with increased tubulin acetylation levels, both at the basal state and after thrombin stimulation. This resulted in impaired platelet aggregation. Similar results were obtained using human platelets that were pretreated with tubacin, an inhibitor of tubulin deacetylase HDAC6. In addition, both ACC and HDAC6 inhibitions block key platelet cytoskeleton signaling events, including Rac1 GTPase activation and the phosphorylation of its downstream effector, p21-activated kinase 2 (PAK2). However, neither CP640.186 nor tubacin affects thrombin-induced actin cytoskeleton remodeling, while ACC inhibition results in decreased thrombin-induced reactive oxygen species (ROS) production and extracellular signal-regulated kinase (ERK) phosphorylation. We conclude that when using washed human platelets, ACC inhibition limits tubulin deacetylation upon thrombin stimulation, which in turn impairs platelet aggregation. The mechanism involves a downregulation of the Rac1/PAK2 pathway, being independent of actin cytoskeleton.

AMPK-ACC signaling modulates platelet phospholipids and potentiates thrombus formation.

AMP-activated protein kinase (AMPK) α1 is activated in platelets on thrombin or collagen stimulation, and as a consequence, phosphorylates and inhibits acetyl-CoA carboxylase (ACC). Because ACC is crucial for the synthesis of fatty acids, which are essential for platelet activation, we hypothesized that this enzyme plays a central regulatory role in platelet function. To investigate this, we used a double knock-in (DKI) mouse model in which the AMPK phosphorylation sites Ser79 on ACC1 and Ser212 on ACC2 were mutated to prevent AMPK signaling to ACC. Suppression of ACC phosphorylation promoted injury-induced arterial thrombosis in vivo and enhanced thrombus growth ex vivo on collagen-coated surfaces under flow. After collagen stimulation, loss of AMPK-ACC signaling was associated with amplified thromboxane generation and dense granule secretion. ACC DKI platelets had increased arachidonic acid-containing phosphatidylethanolamine plasmalogen lipids. In conclusion, AMPK-ACC signaling is coupled to the control of thrombosis by specifically modulating thromboxane and granule release in response to collagen. It appears to achieve this by increasing platelet phospholipid content required for the generation of arachidonic acid, a key mediator of platelet activation.

A single day of high-fat diet feeding induces lipid accumulation and insulin resistance in brown adipose tissue in mice.

Brown adipose tissue (BAT) catabolizes glucose and fatty acids to produce heat and thereby contributes to energy expenditure. Long-term high-fat diet (HFD) feeding results in so-called 'whitening' of BAT characterized by increased lipid deposition, mitochondrial dysfunction, and reduced fat oxidation. The aim of the current study was to unravel the rate and related mechanisms by which HFD induces BAT whitening and insulin resistance. Wild-type mice were fed a HFD for 0, 1, 3, or 7 days. Within 1 day of HFD, BAT weight and lipid content were increased. HFD also immediately reduced insulin-stimulated glucose uptake by BAT, indicating rapid induction of insulin resistance. This was accompanied by a tendency toward a reduced uptake of triglyceride-derived fatty acids by BAT. Mitochondrial mass and Ucp1 expression were unaltered, whereas after 3 days of HFD, markers of mitochondrial dynamics suggested induction of a more fused mitochondrial network. Additionally, HFD also increased macrophage markers in BAT after 3 days of HFD. Counterintuitively, the switch to HFD was accompanied by an acute rise in core body temperature. We showed that a single day of HFD feeding is sufficient to induce the first signs of whitening and insulin resistance in BAT, which reduces the uptake of glucose and triglyceride-derived fatty acids. BAT whitening and insulin resistance are likely sustained by reduced mitochondrial oxidation due to changes in mitochondrial dynamics and macrophage infiltration, respectively. Likely, the switch to HFD swiftly induces thermogenesis in other metabolic organs, which allows attenuation of BAT thermogenesis.

SpecPad: device-independent NMR data visualization and processing based on the novel DART programming language and Html5 Web technology.

SpecPad is a new device-independent software program for the visualization and processing of one-dimensional and two-dimensional nuclear magnetic resonance (NMR) time domain (FID) and frequency domain (spectrum) data. It is the result of a project to investigate whether the novel programming language DART, in combination with Html5 Web technology, forms a suitable base to write an NMR data evaluation software which runs on modern computing devices such as Android, iOS, and Windows tablets as well as on Windows, Linux, and Mac OS X desktop PCs and notebooks. Another topic of interest is whether this technique also effectively supports the required sophisticated graphical and computational algorithms. SpecPad is device-independent because DART's compiled executable code is JavaScript and can, therefore, be run by the browsers of PCs and tablets. Because of Html5 browser cache technology, SpecPad may be operated off-line. Network access is only required during data import or export, e.g. via a Cloud service, or for software updates. A professional and easy to use graphical user interface consistent across all hardware platforms supports touch screen features on mobile devices for zooming and panning and for NMR-related interactive operations such as phasing, integration, peak picking, or atom assignment. Copyright © 2017 John Wiley & Sons, Ltd.

Chronic helminth infection and helminth-derived egg antigens promote adipose tissue M2 macrophages and improve insulin sensitivity in obese mice.

Chronic low-grade inflammation associated with obesity contributes to insulin resistance and type 2 diabetes. Helminth parasites are the strongest natural inducers of type 2 immune responses, and short-lived infection with rodent nematodes was reported to improve glucose tolerance in obese mice. Here, we investigated the effects of chronic infection (12 weeks) with Schistosoma mansoni, a helminth that infects millions of humans worldwide, on whole-body metabolic homeostasis and white adipose tissue (WAT) immune cell composition in high-fat diet-induced obese C57BL/6 male mice. Our data indicate that chronic helminth infection reduced body weight gain (-62%), fat mass gain (-89%), and adipocyte size; lowered whole-body insulin resistance (-23%) and glucose intolerance (-16%); and improved peripheral glucose uptake (+25%) and WAT insulin sensitivity. Analysis of immune cell composition by flow cytometry and quantitative PCR (qPCR) revealed that S. mansoni promoted strong increases in WAT eosinophils and alternatively activated (M2) macrophages. Importantly, injections with S. mansoni-soluble egg antigens (SEA) recapitulated the beneficial effect of parasite infection on whole-body metabolic homeostasis and induced type 2 immune responses in WAT and liver. Taken together, we provide novel data suggesting that chronic helminth infection and helminth-derived molecules protect against metabolic disorders by promoting a T helper 2 (Th2) response, eosinophilia, and WAT M2 polarization.

Middle-aged overweight South Asian men exhibit a different metabolic adaptation to short-term energy restriction compared with Europeans.

AIMS/HYPOTHESIS: South Asians have a higher risk of developing type 2 diabetes than Europeans. The underlying cause of this excess risk is still poorly understood but might be related to differences in the regulation of energy/nutrient-sensing pathways in metabolic tissues and subsequent changes in whole-body substrate metabolism. In this study, we investigated the whole-body and skeletal muscle metabolic adaptations to short-term energy restriction in South Asian and European volunteers. METHODS: Twenty-four middle-aged overweight South Asian and European men underwent a two-step hyperinsulinaemic-euglycaemic clamp, with skeletal muscle biopsies and indirect calorimetry before and after an 8 day diet very low in energy (very low calorie diet [VLCD]). Abdominal fat distribution and hepatic triacylglycerol content were assessed using MRI and MR spectroscopy. RESULTS: South Asian men had higher hepatic triacylglycerol content than European men, and exhibited elevated clamp insulin levels that probably reflect a lower insulin clearance rate. Despite higher insulin levels, endogenous glucose production rate was similar and glucose disposal rate (Rd) and nonoxidative glucose disposal rate (NOGD) were significantly lower in South Asian than European men, indicating impaired whole-body insulin sensitivity. Energy restriction decreased abdominal fat mass and hepatic triacylglycerol content in both groups. However, the shift induced by energy restriction from glucose towards lipid oxidation observed in European men was impaired in South Asian men, indicating whole-body metabolic inflexibility. Remarkably, although energy restriction improved hepatic insulin sensitivity in both groups, Rd improved only in South Asian men owing to higher NOGD. At the molecular level, an increase in insulin-induced activation of the skeletal muscle mTOR pathway was found in South Asian men, showing that skeletal muscle energy/nutrient-sensing pathways were differentially affected by energy restriction. CONCLUSIONS/INTERPRETATION: We conclude that South Asian men exhibit a different metabolic adaptation to short-term energy restriction than European men. TRIAL REGISTRATION: Dutch trial registry ( www.trialregister.nl ), trial number NTR 2473.

Short-term high-fat diet increases macrophage markers in skeletal muscle accompanied by impaired insulin signalling in healthy male subjects.

Macrophage markers in skeletal muscle of obese subjects are elevated and inversely relate to insulin sensitivity. The present study aimed to investigate whether short-term high-fat high-calorie (HFHC) diet already increases macrophage markers and affects glucose metabolism in skeletal muscle of healthy lean subjects. Muscle biopsies were obtained from 24 healthy lean young men before and after a 5-day HFHC-diet. mRNA expression levels of relevant genes in muscle and glucose, insulin, C-peptide and cholesteryl ester transfer protein (CETP) levels in plasma were measured. In addition, we assessed hepatic triacylglycerol ('triglyceride') (HTG) content by magnetic resonance spectroscopy and subcutaneous white adipose tissue (sWAT) biopsies were analysed histologically from a subset of subjects (n=8). A 5-day HFHC-diet markedly increased skeletal muscle mRNA expression of the general macrophage markers CD68 (3.7-fold, P<0.01) and CD14 (3.2-fold, P<0.01), as well as the M1 macrophage markers MARCO (11.2-fold, P<0.05), CD11c (1.8-fold, P<0.05) and MRC1 (1.7-fold, P<0.05). This was accompanied by down-regulation of SLC2A4 and GYS1 mRNA expression, and elevated plasma glucose (+4%, P<0.001) and insulin (+55%, P<0.001) levels together with homoeostasis model assessment of insulin resistance (HOMA-IR) (+48%, P<0.001), suggesting development of insulin resistance (IR). Furthermore, the HFHC-diet markedly increased HTG (+118%, P<0.001) and plasma CETP levels (+21%, P<0.001), a marker of liver macrophage content, whereas sWAT macrophage content remained unchanged. In conclusion, short-term HFHC-diet increases expression of macrophage markers in skeletal muscle of healthy men accompanied by reduced markers of insulin signalling and development of IR. Therefore, recruitment of macrophages into muscle may be an early event in development of IR in response to short-term HFHC-feeding.

Peripheral cannabinoid 1 receptor blockade activates brown adipose tissue and diminishes dyslipidemia and obesity.

The endocannabinoid system is an important player in energy metabolism by regulating appetite, lipolysis, and energy expenditure. Chronic blockade of the cannabinoid 1 receptor (CB1R) leads to long-term maintenance of weight loss and reduction of dyslipidemia in experimental and human obesity. The molecular mechanism by which CB1R blockade reverses dyslipidemia in obesity has not yet been clarified. In this study, we showed that CB1R blockade with the systemic CB1R blocker rimonabant enhanced whole-body energy expenditure and activated brown adipose tissue (BAT), indicated by increased expression of genes involved in BAT thermogenesis and decreased lipid droplet size in BAT. This was accompanied by selectively increased triglyceride (TG) uptake by BAT and lower plasma TG levels. Interestingly, the effects on BAT activation were still present at thermoneutrality and could be recapitulated by using the strictly peripheral CB1R antagonist AM6545, indicating direct peripheral activation of BAT. Indeed, CB1R blockade directly activated T37i brown adipocytes, resulting in enhanced uncoupled respiration, most likely via enhancing cAMP/PKA signaling via the adrenergic receptor pathway. Our data indicate that selective targeting of the peripheral CB1R in BAT has therapeutic potential in attenuating dyslipidemia and obesity.

Helminth infections and type 2 diabetes: a cluster-randomized placebo controlled SUGARSPIN trial in Nangapanda, Flores, Indonesia.

BACKGROUND: Insulin resistance is a strong predictor of the development of type 2 diabetes mellitus. Chronic helminth infections might protect against insulin resistance via a caloric restriction state and indirectly via T-helper-2 polarization of the immune system. Therefore the elimination of helminths might remove this beneficial effect on insulin resistance. METHODS/DESIGN: To determine whether soil-transmitted helminth infections are associated with a better whole-body insulin sensitivity and whether this protection is reversible by anthelmintic treatment, a household-based cluster-randomized, double blind, placebo-controlled trial was conducted in the area of Nangapanda on Flores Island, Indonesia, an area endemic for soil-transmitted helminth infections. The trial incorporates three monthly treatment with albendazole or matching placebo for one year, whereby each treatment round consists of three consecutive days of supervised drug intake. The presence of soil-transmitted helminths will be evaluated in faeces using microscopy and/or PCR. The primary outcome of the study will be changes in insulin resistance as assessed by HOMA-IR, while the secondary outcomes will be changes in body mass index, waist circumference, fasting blood glucose, 2 h-glucose levels after oral glucose tolerance test, HbA1c, serum lipid levels, immunological parameters, and efficacy of anthelmintic treatment. DISCUSSION: The study will provide data on the effect of helminth infections on insulin resistance. It will assess the relationship between helminth infection status and immune responses as well as metabolic parameters, allowing the establishment of a link between inflammation and whole-body metabolic homeostasis. In addition, it will give information on anthelmintic treatment efficacy and effectiveness. TRIAL REGISTRATION: This study has been approved by the ethical committee of Faculty of Medicine Universitas Indonesia (ref: 549/H2.F1/ETIK/2013), and has been filed by the ethics committee of Leiden University Medical Center, clinical trial number: ISRCTN75636394. The study is reported in accordance with the CONSORT guidelines for cluster-randomised trials.

Regulation of skeletal muscle energy/nutrient-sensing pathways during metabolic adaptation to fasting in healthy humans.

During fasting, rapid metabolic adaptations are required to maintain energy homeostasis. This occurs by a coordinated regulation of energy/nutrient-sensing pathways leading to transcriptional activation and repression of specific sets of genes. The aim of the study was to investigate how short-term fasting affects whole body energy homeostasis and skeletal muscle energy/nutrient-sensing pathways and transcriptome in humans. For this purpose, 12 young healthy men were studied during a 24-h fast. Whole body glucose/lipid oxidation rates were determined by indirect calorimetry, and blood and skeletal muscle biopsies were collected and analyzed at baseline and after 10 and 24 h of fasting. As expected, fasting induced a time-dependent decrease in plasma insulin and leptin levels, whereas levels of ketone bodies and free fatty acids increased. This was associated with a metabolic shift from glucose toward lipid oxidation. At the molecular level, activation of the protein kinase B (PKB/Akt) and mammalian target of rapamycin pathways was time-dependently reduced in skeletal muscle during fasting, whereas the AMP-activated protein kinase activity remained unaffected. Furthermore, we report some changes in the phosphorylation and/or content of forkhead protein 1, sirtuin 1, and class IIa histone deacetylase 4, suggesting that these pathways might be involved in the transcriptional adaptation to fasting. Finally, transcriptome profiling identified genes that were significantly regulated by fasting in skeletal muscle at both early and late time points. Collectively, our study provides a comprehensive map of the main energy/nutrient-sensing pathways and transcriptomic changes during short-term adaptation to fasting in human skeletal muscle.

The degree of liver injury determines the role of p21 in liver regeneration and hepatocarcinogenesis in mice.

UNLABELLED: Hepatocellular carcinoma (HCC) frequently arises in the context of chronic injury that promotes DNA damage and chromosomal aberrations. The cyclin-dependent kinase inhibitor p21 is an important transcriptional target of several tumor suppressors, which promotes cell cycle arrest in response to many stimuli. The aim of this study was to further delineate the role of p21 in the liver during moderate and severe injury and to specify its role in the initiation and progression of HCC. Deletion of p21 led to continuous hepatocyte proliferation in mice with severe injury allowing animal survival but also facilitated rapid tumor development, suggesting that control of compensatory proliferation by high levels of p21 is critical to the prevention of tumor development. Unexpectedly, however, liver regeneration and hepatocarcinogenesis was impaired in p21-deficient mice with moderate injury. Mechanistically, loss of p21 was compensated by activation of Sestrin2, which impaired mitogenic mammalian target of rapamycin (mTOR) signaling and activated cytoprotective Nrf2 signaling. CONCLUSION: The degree of liver injury and the strength of p21 activation determine its effects on liver regeneration and tumor development in the liver. Moreover, our data uncover a molecular link in the complex mTOR, Nrf2, and p53/p21-signaling network through activation of Sestrin2, which regulates hepatocyte proliferation and tumor development in mice with liver injury.

A polyphenol-rich plant extract prevents hypercholesterolemia and modulates gut microbiota in western diet-fed mice.

Introduction: Totum-070 is a combination of five plant extracts enriched in polyphenols to target hypercholesterolemia, one of the main risk factors for cardiovascular diseases. The aim of this study was to investigate the effects of Totum-070 on cholesterol levels in an animal model of diet-induced hypercholesterolemia. Methods: C57BL/6JOlaHsd male mice were fed a Western diet and received Totum-070, or not, by daily gavage (1g/kg and 3g/kg body weight) for 6 weeks. Results: The Western diet induced obesity, fat accumulation, hepatic steatosis and increased plasma cholesterol compared with the control group. All these metabolic perturbations were alleviated by Totum-070 supplementation in a dose-dependent manner. Lipid excretion in feces was higher in mice supplemented with Totum-070, suggesting inhibition of intestinal lipid absorption. Totum-070 also increased the fecal concentration of short chain fatty acids, demonstrating a direct effect on intestinal microbiota. Discussion: The characterization of fecal microbiota by 16S amplicon sequencing showed that Totum-070 supplementation modulated the dysbiosis associated with metabolic disorders. Specifically, Totum-070 increased the relative abundance of Muribaculum (a beneficial bacterium) and reduced that of Lactococcus (a genus positively correlated with increased plasma cholesterol level). Together, these findings indicate that the cholesterol-lowering effect of Totum-070 bioactive molecules could be mediated through multiple actions on the intestine and gut microbiota.