Dr Barbara Fielding

Background: Epidemiological studies suggest that metformin may reduce the incidence of cancer in patients with diabetes and multiple late phase clinical trials assessing the potential of repurposing this drug are underway. Transcriptomic profiling of tumour samples is an excellent tool to understand drug bioactivity, identify candidate biomarkers and assess for mechanisms of resistance to therapy. Methods: 36 patients with untreated primary breast cancer were recruited to a window study and transcriptomic profiling of tumour samples carried out before and after metformin treatment. Results: Multiple genes that regulate fatty acid oxidation were upregulated at the transcriptomic level and there was a differential change in expression between 2 previously identified cohorts of patients with distinct metabolic responses. Increase in expression of a mitochondrial fatty oxidation gene composite signature correlated with change in a proliferation gene signature. In vitro assays showed that, in contrast to previous studies in models of normal cells, metformin reduces fatty acid oxidation with a subsequent accumulation of intracellular triglyceride, independent of AMPK activation. Conclusions: We propose that metformin at clinical doses targets fatty acid oxidation in cancer cells with implications for patient selection and drug combinations.

Tetracosahexaenoic acid (24:6ω-3) is an intermediate in the conversion of 18:3ω-3 to 22:6ω-3 in mammals. There is limited information about whether cells can assimilate and metabolize exogenous 24:6ω-3. This study compared the effect of incubation with 24:6ω-3 on the fatty acid composition of two related cell types, primary CD3+ T lymphocytes and Jurkat T cell leukemia, which differ in the integrity of the polyunsaturated fatty acid (PUFA) biosynthesis pathway. 24:6ω-3 was only detected in either cell type when cells were incubated with 24:6ω-3. Incubation with 24:6ω-3 induced similar increments in the amount of 22:6ω-3 in both cell types and modified the homeoviscous adaptations fatty acid composition induced by activation of T lymphocytes. The effect of incubation with 18:3ω-3 compared to 24:6ω-3 on the increment in 22:6ω-3 was tested in Jurkat cells because primary T cells cannot convert 18:3ω-3 to 22:6ω-3. The increment in the 22:6ω-3 content of Jurkat cells incubated with 24:6ω-3 was 19.5-fold greater than that of cells incubated with 18:3ω-3. Acyl-coA oxidase siRNA knockdown decreased the amount of 22:6ω-3 and increased the amount of 24:6ω-3 in Jurkat cells. These findings show exogenous 24:6ω-3 can be incorporated into primary human T lymphocytes and Jurkat cells and induces changes in fatty acid composition consistent with its conversion to 22:6ω-3 via a mechanism involving peroxisomal β-oxidation that is regulated independently from the integrity of the upstream PUFA synthesis pathway. One further implication is that consuming 24:6ω-3 may be an effective alternative means of achieving health benefits attributed to 20:5ω-3 and 22:6ω-3.

BACKGROUND Duodenal mucosal resurfacing (DMR) is a novel day-case endoscopic intervention which results in weight loss-independent reductions in HbA1c in patient with type 2 diabetes mellitus (T2DM). We hypothesized that DMR works by increasing insulin sensitivity and we aimed to investigate the mechanism of action of DMR through longitudinal metabolic phenotyping in humans. METHODSThirty-two insulin-resistant women with polycystic ovary syndrome (PCOS) and obesity were randomised in a double-blinded manner to DMR or sham endoscopy. They underwent measurements of insulin sensitivity using euglycaemic hyperinsulinaemic clamps, insulin secretion using oral glucose tolerance tests and reproductive function using weekly reproductive hormone profiles and ovarian ultrasonography for 6 months post-intervention. RESULTSA small increase in total body insulin sensitivity measured by the clamp was observed in both groups at week 12. An increase in insulin sensitivity, as measured by HOMA-IR, was observed in both groups at week 24. There was an increase in the number of menses (median 2 DMR, 0.5 sham). There were no significant differences between the two groups in these outcomes or insulin secretion. CONCLUSIONSThese findings suggest that DMR does not work by increasing insulin sensitivity in euglycaemic, insulin resistant women with PCOS. The procedure may exert its effects only in the context of hyperglycaemia or pathologically hyperplastic, insulin-desensitised duodenal mucosa.

Longer chain polyunsaturated fatty acids (LCPUFA) ≥ 20 carbons long are required for leukocyte function. These can be obtained from the diet, but there is some evidence that leukocytes can convert essential fatty acids (EFA) into LCPUFA. We used stable isotope tracers to investigate LCPUFA biosynthesis, and the effect of different EFA substrate ratios, in human T lymphocytes. CD3+ T cells were incubated for up to 48 hours with or without concanavalin A in media containing a 18:2n-6 : 18:3n-3 (EFA) ratio of either 5:1 or 8:1, and [13C]18:3n-3 plus [d5]18:2n-6. Mitogen stimulation increased the amounts of 16:1n 7, 18:1n 9, 18:2n 6, 20:3n 6, 20:4n 6, 18:3n 3 and 20:5n 3 in T cells. Expression of the activation marker CD69 preceded increased FADS2 and FADS1 mRNA expression, and increased amounts of [d5]20:2n 6 and [13C]20:3n 3 at 48 hours. 22 carbon n 6 or n 3 LCPUFA synthesis was not detected, consistent with the absence of ELOVL2 expression. An EFA ratio of 8:1 reduced 18:3n 3 conversion and enhanced 20:2n 6 synthesis compared to a 5:1 ratio. [d5]9- and [d5]-13-hydroxyoctadecadienoic (HODE) and [13C]9- and [13C]13-hydroxyoctadecatrienoic acids (HOTrE) were the major labelled oxylipins in culture supernatants; labelled oxylipins ≥ 20 carbons were not detected. An EFA ratio of 8:1 suppressed 9- and 13-HOTrE synthesis, but there was no significant effect on 9- and 13-HODE synthesis. These findings suggest that partitioning of newly assimilated EFA between LCPUFA synthesis and hydroxyoctadecaenoic acid may be a metabolic branch point in T cell EFA metabolism that has implications for understanding the effects of dietary fats on T lymphocyte function.

Gastric emptying (GE) is the process of food being processed by the stomach and delivered to the small intestine where nutrients such as lipids are absorbed into the blood circulation. The combination of an easy and inexpensive method to measure GE such as the CO2 breath test using the stable isotope [ 13 C]octanoic acid with semi-mechanistic modelling could foster a wider application in nutritional studies to further understand the metabolic response to food. Here, we discuss the use of the [ 13 C]octanoic acid breath test to label the solid phase of a meal, and the factors that influence GE to support mechanistic studies. Furthermore, we give an overview of existing mathematical models for the interpretation of the breath test data and how much nutritional studies could benefit from a physiological based pharmacokinetic model approach.

Scope: Fructose exacerbates post-prandial hypertriacylglycerolaemia. This may be partly due to increased enterocyte de novo lipogenesis (DNL). It is unknown whether this is concentration-dependent or whether fructose has a greater effect on lipid synthesis than glucose. The dose-dependent effects of fructose and glucose on DNL and de novo triacylglycerol (TAG)-glycerol synthesis were investigated in an enterocyte model; Caco-2 cells. Methods and results: Caco-2 cells were treated for 96h with 5mM, 25mM or 50mM fructose or glucose, or 12.5mM fructose/12.5mM glucose mix. DNL was measured following addition of [13C2]-acetate to apical media. In separate experiments, [13C6]-fructose and [13C6]-glucose were used to measure DNL and de novo TAG-glycerol synthesis. DNL from [13C2]-acetate was detected following all treatments, with greater amounts in intracellular than secreted (media) samples (all P

Purpose: UK guidelines recommend dietary saturated fatty acids (SFAs) should not exceed 10% total energy (%TE) for cardiovascular disease prevention, with benefits observed when SFAs are replaced with unsaturated fatty acids (UFAs). This study aimed to assess the efficacy of a dietary exchange model using commercially available foods to replace SFAs with UFAs. Methods: Healthy men (n=109, age 48, SD 11y) recruited to the Reading, Imperial, Surrey, Saturated fat Cholesterol Intervention-1 (RISSCI-1) study (ClinicalTrials.Gov n°NCT03270527) followed two sequential 4-week isoenergetic moderate-fat (34%TE) diets: high-SFA (18%TE SFAs, 16%TE UFAs) and low-SFA (10%TE SFAs, 24%TE UFAs). Dietary intakes were assessed using 4-day weighed diet diaries. Nutrient intakes were analysed using paired t-tests, fasting plasma phospholipid fatty acid (PL-FA) profiles and dietary patterns were analysed using orthogonal partial least square discriminant analyses. Results: Participants exchanged 10.2%TE (SD 4.1) SFAs for 9.7%TE (SD 3.9) UFAs between the high and low-SFA diets, reaching target intakes with minimal effect on other nutrients or energy intakes. Analyses of dietary patterns confirmed successful incorporation of recommended foods from commercially available sources (e.g. dairy products, snacks, oils, and fats), without affecting participants’ overall dietary intakes. Analyses of plasma PL-FAs indicated good compliance to the dietary intervention and foods of varying SFA content. Conclusions: RISSCI-1 dietary exchange model successfully replaced dietary SFAs with UFAs in free-living healthy men using commercially available foods, and without altering their dietary patterns. Further intervention studies are required to confirm utility and feasibility of such food-based dietary fat replacement models at a population level.

Specialized metabolic-sensors in the hypothalamus regulate blood glucose levels by influencing hepatic glucose output and hypoglycemic counter regulatory responses. Hypothalamic reactive oxygen species (ROS) may act as a metabolic signal mediating responses to changes in glucose, other substrates and hormones. The role of ROS in the brain’s control of glucose homeostasis remains unclear. We hypothesized that hydrogen peroxide (H2O2), a relatively stable form of ROS, acts as a sensor of neuronal glucose consumption and availability and that lowering brain H2O2 with the enzyme catalase would lead to systemic responses increasing blood glucose. During hyperinsulinemic euglycemic clamps in rats, ICV catalase infusion resulted in increased hepatic glucose output, which was associated with reduced neuronal activity in the arcuate nucleus of the hypothalamus (ARC). Electrophysiological recordings revealed a subset of ARC neurons expressing pro-opiomelanocortin (POMC) that were inhibited by catalase and excited by H2O2. During hypoglycemic clamps, ICV catalase increased glucagon and epinephrine responses to hypoglycemia, consistent with perceived lower glucose levels. Our data suggest that H2O2 represents an important metabolic cue which, through tuning the electrical activity of key neuronal populations such as POMC neurons, may have a role in the brain’s influence of glucose homeostasis and energy balance.

Purpose of review: There is considerable political and public awareness of new recommendations to reduce sugars and sugar-sweetened beverages in our diets. It is therefore timely to review the most recent changes in guidelines, with a focus on evidence for metabolic health, recent research in the area and gaps in our knowledge. Recent findings: Sufficient evidence links a high intake of sugar to dental caries and obesity, and high intakes of sugar-sweetened beverages in particular to increased risk of type 2 diabetes. This has led to the updating of dietary recommendations related to added sugars in the diet. The effects of specific sugars at usual intakes as part of an isoenergetic diet are less clear. The glycaemic response to food is complex and mediated by many factors, but sugar intake is not necessarily the major component. Summary: There are many challenges faced by healthcare professionals and government bodies in order to improve the health of individuals and nations through evidence-based diets. Sufficiently powered long-term mechanistic studies are still required to provide evidence for the effects of reducing dietary sugars on metabolic health. However, there are many challenges for research scientists in the implementation of these studies.

The expansion of lower-body adipose tissue (AT) is paradoxically associated with reduced cardiovascular disease and diabetes risk. We examined whether the beneficial metabolic properties of lower-body AT are related to the production and release of the insulin-sensitizing lipokine palmitoleate (16:1n-7). Using venoarterial difference sampling, we investigated the relative release of 16:1n-7 from lower-body (gluteofemoral) and upper-body (abdominal subcutaneous) AT depots. Paired gluteofemoral and abdominal subcutaneous AT samples were analyzed for triglyceride fatty acid composition and mRNA expression. Finally, the triglyceride fatty acid composition of isolated human preadipocytes was determined. Relative release of 16:1n-7 was markedly higher from gluteofemoral AT compared with abdominal subcutaneous AT. Stearoyl-CoA desaturase 1 (SCD1), the key enzyme involved in endogenous 16:1n-7 production, was more highly expressed in gluteofemoral AT and was associated with greater enrichment of 16:1n-7. Furthermore, isolated human preadipocytes from gluteofemoral AT displayed a higher content of SCD1-derived fatty acids. We demonstrate that human gluteofemoral AT plays a major role in determining systemic concentrations of the lipokine palmitoleate. Moreover, this appears to be an inherent feature of gluteofemoral AT. We propose that the beneficial metabolic properties of lower-body AT may be partly explained by the intrinsically greater production and release of palmitoleate.

The menopause is accompanied by increased risk of obesity, altered body fat distribution and decreased skeletal muscle mass. The resulting decrease in RMR should be accompanied by a compensatory change in energy balance to avoid weight gain. We aimed to investigate habitual energy intake and expenditure in pre- and postmenopausal women matched for abdominal obesity. We recruited fifty-one healthy Caucasian women, BMI > 18·5 and 

Objective: To determine the effect of SGLT2 inhibitor dapagliflozin on glucose flux, lipolysis and ketone body concentrations during insulin withdrawal in people with type 1 diabetes. Research Design and Methods: A double-blind placebo controlled crossover study with a 4-week wash out period was performed in 12 people with type 1 diabetes using insulin pump therapy. Participants received dapagliflozin or placebo in random order for 7 days. Stable isotopes were infused to measure the rate of glucose production (Ra), disappearance (Rd) and lipolysis. At isotopic steady state insulin was withdrawn and the study terminated after 600 minutes or earlier if blood glucose reached 18mmol/L, bicarbonate 27 and

Polyunsaturated fatty acids (PUFAs) are important for immune function. Limited evidence indicates that immune cell activation involves endogenous PUFA synthesis, but this has not been characterised. To address this, we measured metabolism of 18:3n-3 in quiescent and activated peripheral blood mononuclear cells (PBMCs), and in Jurkat T cell leukaemia. PBMCs from men and women (n = 34) were incubated with [1-13C]18:3n-3 with or without Concanavalin A (Con. A). 18:3n-3 conversion was undetectable in unstimulated PBMCs, but up-regulated when stimulated. The main products were 20:3n-3 and 20:4n-3, while 18:4n-3 was undetectable, suggesting initial elongation and 8 desaturation. PUFA synthesis was 17.4-fold greater in Jurkat cells than PBMCs. The major products of 18:3n-3 conversion in Jurkat cells were 20:4n-3, 20:5n-3 and 22:5n-3. 13C Enrichment of 18:4n-3 and 20:3n-3 suggests parallel initial elongation and Δ6 desaturation. The FADS2 inhibitor SC26196 reduced PBMC, but not Jurkat cell, proliferation suggesting PUFA synthesis is involved in regulating mitosis in PBMCs. Con. A stimulation increased FADS2, FADS1, ELOVL5 and ELOVL4 mRNA expression in PBMCs. A single transcript corresponding to the major isoform of FADS2, FADS20001, was detected in PBMCs and Jurkat cells. PBMC activation induced hypermethylation of a 470bp region in the FADS2 5’-regulatory sequence. This region was hypomethylated in Jurkat cells compared to quiescent PBMCs. These findings show that PUFA synthesis involving initial elongation and 8 desaturation is involved in regulating PBMC proliferation and is regulated via transcription possibly by altered DNA methylation. These processes were dysregulated in Jurkat cells. This has implications for understanding the regulation of mitosis in normal and transformed lymphocytes.

Background Android fat distribution (abdominal obesity) is associated with insulin resistance, hepatic steatosis and greater secretion of large very low density lipoprotein (VLDL) particles in men. Since abdominal obesity is becoming increasingly prevalent in women we aimed to investigate the relationship between android fat and hepatic lipid metabolism in pre- and post-menopausal women. Methods and Results We used a combination of stable isotope tracer techniques to investigate intrahepatic fatty acid synthesis and partitioning in 29 lean and 29 abdominally obese women (android fat/total fat 0.065 (0.02-0.08) and 0.095 (0.08-0.11) respectively). Thirty women were pre-menopausal aged 35-45 and they were matched for abdominal obesity with 28 post-menopausal women aged 55-65. As anticipated, abdominal obese women were more insulin resistant with enhanced hepatic secretion of large (404±30 v 268±26 mg/kg lean mass, P

Most postprandial studies have investigated the response of a single meal, yet the ingestion of sequential meals is more typical in a Western society. The aim of this review is to explain how natural and stable isotope tracers of fatty acids have been used to investigate the metabolism of dietary fat after single and multiple meals, with a focus on in vivo measurements of adipose tissue metabolism. When stable isotope tracers are combined with arteriovenous difference measurements, very specific measurements of metabolic flux across tissues can be made. We have found that adipose tissue is a net importer of dietary fat for 5 h following a single test meal and for most of the day during a typical three-meal eating pattern. When dietary fat is cleared from plasma, some fatty acids ‘spillover’ into the plasma and contribute up to 50 % of postprandial plasma non-esterified fatty acid concentrations. Therefore, plasma non-esterified fatty acid concentrations after a meal reflect the balance between intracellular and extracellular lipolysis in adipose tissue. This balance is altered after the acute ingestion of fructose. The enzyme lipoprotein lipase is a key modulator of fatty acid flux in adipose tissue and its rate of action is severely diminished in obese men. In conclusion, in vivo studies of human metabolism can quantify the way that fatty acid trafficking modulates plasma lipid concentrations. The magnitude of fatty acid flux from adipose tissue has implications for ectopic fat deposition in tissues such as the liver and muscle.

Fatty acid binding protein 4 (FABP4) is a fatty acid chaperone, which is induced during adipocyte differentiation. Previously we have shown that endothelial FABP4 is induced by the NOTCH1 signalling pathway, which is involved in mechanisms of resistance to anti-angiogenic tumour therapy1. Here, we investigated the role of FABP4 in endothelial fatty acid metabolism and tumour angiogenesis. We analysed the effect of transient FABP4 knockdown in human umbilical vein endothelial cells on fatty acid metabolism, viability, and angiogenesis. Through therapeutic delivery of siRNA targeting murine FABP4, we investigated the effect of stromal FABP4 knockdown on tumour growth and blood vessel formation. In vitro, siRNA-mediated FABP4 knockdown in endothelial cells led to a marked increase of endothelial fatty acid oxidation, an increase of reactive oxygen species (ROS), and decreased angiogenesis. In vivo, we found that increased NOTCH1 signalling in tumour xenografts led to increased expression of endothelial FABP4 that decreased when NOTCH1 and VEGFA inhibitors were used in combination. Angiogenesis, growth, and metastasis in ovarian tumour xenografts were markedly inhibited by therapeutic siRNA delivery targeting mouse FABP4. Therapeutic targeting of endothelial FABP4 by siRNA in vivo has anti-angiogenic and anti-tumour effects with minimal toxicity and should be investigated further.

Context GLP-1 agonists control postprandial glucose and lipid excursion in type 2 diabetes; however the mechanism(s) are unclear. Objective To determine the mechanism(s) of postprandial lipid and glucose control with lixisenatide (GLP-1 analogue) in type 2 diabetes. Design Randomised, double-blind, cross-over study. Setting Centre for Diabetes, Endocrinology, and Research, Royal Surrey County Hospital, Guildford, UK Patients Eight obese men with type 2 diabetes (57.3±1.9yrs; BMI 30.3±1.0kg/m2, HbA1C 66.5±2.6mmol/mol, [8.2±0.3%]). Interventions Two metabolic studies, four-weeks after lixisenatide or placebo; with cross-over and repetition of studies. Main outcome measures Study one: very-low density lipoprotein (VLDL) and chylomicron (CM) triacylglycerol (TAG) kinetics were measured with iv bolus of [2H5]glycerol in a 12h study, with hourly feeding. Oral [13C]triolein, in a single meal, labelled enterally-derived TAG. Study two: glucose kinetics were measured with [U-13C]glucose in a mixed-meal (plus acetaminophen to measure gastric emptying) and variable iv [6,6-2H2]glucose infusion. Results Study one: CM-TAG (but not VLDL-TAG) pool-size, was lower with lixisenatide (P=0.046). Lixisenatide reduced CM [13C]oleate AUC60-480min concentration (P=0.048) and increased CM-TAG clearance; with no effect on CM-TAG production rate. Study two: postprandial glucose and insulin AUC0-240min were reduced with lixisenatide (P=0.0051, P˂0.05). Total glucose production rate (Ra) (P=0.015), Rameal (P=0.0098) and acetaminophen AUC0-360min (P=0.006) were lower with lixisenatide than placebo. Conclusions Lixisenatide reduced [13C]oleate concentration, derived from a single meal in CM-TAG, as well as glucose Rameal, through delayed gastric emptying. However day-long CM production, measured with repeated meal-feeding, was not reduced by lixisenatide and decreased CM-TAG concentration was due to increased CM-TAG clearance.

Background Epidemiological studies suggest that metformin may reduce the incidence of cancer in patients with diabetes and multiple late phase clinical trials assessing the potential of repurposing this drug are underway. Transcriptomic profiling of tumour samples is an excellent tool to understand drug bioactivity, identify candidate biomarkers and assess for mechanisms of resistance to therapy. Methods Thirty-six patients with untreated primary breast cancer were recruited to a window study and transcriptomic profiling of tumour samples carried out before and after metformin treatment. Results Multiple genes that regulate fatty acid oxidation were upregulated at the transcriptomic level and there was a differential change in expression between two previously identified cohorts of patients with distinct metabolic responses. Increase in expression of a mitochondrial fatty oxidation gene composite signature correlated with change in a proliferation gene signature. In vitro assays showed that, in contrast to previous studies in models of normal cells, metformin reduces fatty acid oxidation with a subsequent accumulation of intracellular triglyceride, independent of AMPK activation. Conclusions We propose that metformin at clinical doses targets fatty acid oxidation in cancer cells with implications for patient selection and drug combinations. Clinical Trial Registration NCT01266486.

mpaired regulation of immune function characterised by chronic inflammation together with a declining protective immune response is a major challenge to healthy ageing. It is therefore important to understand the mechanisms that regulate immune function and the impact of ageing upon such processes. Appropriate induction and resolution of the immune response requires adequate availability of polyunsaturated fatty acids (PUFAs) for incorporation into cell membranes. However, humans are unable to synthesise PUFAs de novo and are dependent upon dietary intake for pre-formed PUFAs or synthesis by the liver from the essential fatty acids, linoleic acid (LA, 18:2n-6) and alpha-linolenic acid (aLNA, 18:3n-3). We have shown that activation of peripheral blood mononuclear cells (PBMCs) 37 increases PUFA biosynthesis from essential fatty acids via a mechanism that involves altered epigenetic regulation of a key gene in the pathway. Moreover, induction of PUFA synthesis is directly involved in the regulation of lymphocyte activation and proliferation. The aim of the BBSRC responsive mode award ‘How does polyunsaturated fatty acid biosynthesis regulate T lymphocyte function?’ is to determine how PUFA biosynthesis regulates T cell function and the effect of ageing on this process. The project will identify points of regulation in the biosynthetic pathway and how these might influence the capacity for up-regulation of PUFA synthesis in older individuals. We will use stable isotope tracers of LA and aLNA to determine whether newly synthesised PUFAs are preferential substrates for synthesis of lipid mediators and whether they are involved in formation of membrane microdomains that mediate cell signalling.

Background: Observational studies often infer hepatic de novo lipogenesis (DNL) by measuring circulating fatty acid (FA) markers; however it remains to be elucidated whether these markers accurately reflect hepatic DNL. Objective: We investigated associations between fasting hepatic DNL and proposed FA markers of DNL in subjects consuming their habitual diet. Design: Fasting hepatic DNL was assessed using ²H2O (heavy water) in 149 non-diabetic men and women and measuring the synthesis of very low-density lipoprotein triglyceride (VLDL-TG) palmitate. FA markers of blood lipid fractions was determined by gas chromatography (GC). Results: Neither the lipogenic index (16:0/18:2n-6) nor the SCD index (16:1n-7/16:0) in VLDL-TG were associated with isotopically assessed DNL (r=0.13, P=0.1 and r=-0.08, P=0.35, respectively). The relative abundance (mol%) of 14:0, 16:0 and 18:0 in VLDL-TG were weakly (r≤0.35) associated with DNL whereas abundance of 16:1n-7, 18:1n-7 and 18:1n-9 were not associated. When the cohort was split by median DNL, only abundance of 14:0 and 18:0 in VLDL-TG could discriminate between subjects having high (11.5%) and low (3.8%) fasting hepatic DNL. Based on a subgroup, FA markers in total plasma TG, plasma cholesteryl esters, plasma phospholipids and red blood cell phospholipids were generally not associated with DNL. Conclusions: The usefulness of circulating FAs as markers of hepatic DNL in healthy individuals consuming their habitual diet is limited due to their inability to clearly discriminate between individuals with low and high fasting hepatic DNL.

Impaired adipose tissue insulin signalling is a critical feature of insulin resistance. Here we identify a pathway linking the lipolytic enzyme hormone-sensitive lipase (HSL) to insulin action via the glucose-responsive transcription factor ChREBP and its target, the fatty acid elongase ELOVL6. Genetic inhibition of HSL in human adipocytes and mouse adipose tissue results in enhanced insulin sensitivity and induction of ELOVL6. ELOVL6 promotes an increase in phospholipid oleic acid, which modifies plasma membrane fluidity and enhances insulin signalling. HSL deficiency–mediated effects are suppressed by gene silencing of ChREBP and ELOVL6. Mechanistically, physical interaction between HSL, independent of lipase activity, and the isoform activated by glucose metabolism ChREBPα impairs ChREBPα translocation into the nucleus and induction of ChREBPβ, the isoform with high transcriptional activity that is strongly associated with whole-body insulin sensitivity. Targeting the HSL–ChREBP interaction may allow therapeutic strategies for the restoration of insulin sensitivity.

Potatoes have been an affordable, staple part of the diet for many hundreds of years. Recently however, there has been a decline in consumption, perhaps influenced by erroneous reports of being an unhealthy food. This review provides an overview of the nutritional value of potatoes and examines the evidence for associations between potato consumption and non-communicable diseases. Potatoes are an important source of micronutrients, such as vitamin C, vitamin B6, potassium, folate, and iron and contribute a significant amount of fibre to the diet. However, nutrient content is affected by cooking method; boiling causes leaching of water-soluble nutrients, whereas frying can increase the resistant starch content of the cooked potato. Epidemiological studies have reported associations between potato intake and obesity, type 2 diabetes and cardiovascular disease. However, results are contradictory and confounded by lack of detail on cooking methods. Indeed, potatoes have been reported to be more satiating than other starchy carbohydrates, such as pasta and rice, which may aid weight maintenance. Future research should consider cooking methods in the study design in order to reduce confounding factors and further explore the health impact of this food.

We hypothesised that probiotic supplementation (PRO) increases the absorption and oxidation of orally ingested maltodextrin during 2h endurance cycling, thereby sparing muscle glycogen for a subsequent time trial (simulating a road race). Measurements were made of lipid and carbohydrate oxidation, plasma metabolites and insulin, gastrointestinal permeability, and subjective symptoms of discomfort. Seven male cyclists were randomized to PRO (bacterial composition given in methods) or placebo (PLC) for four weeks, separated by a 14-day washout period. After each period, cyclists consumed a 10% maltodextrin solution (initial 8 mL·kg-1 bolus and 2 mL·kg-1 every 15 min) while exercising for 2h at 55% Wmax followed by a 100 kJ time trial. PRO resulted in small increases in peak oxidation rates of the ingested maltodextrin (0.84 ± 0.10 vs 0.77 ± 0.09 g·min-1, P = 0.016), and mean total carbohydrate oxidation (2.20 ± 0.25 vs 1.87 ± 0.39 g·min-1, P = 0.038), while fat oxidation was reduced (0.40 ± 0.11 vs 0.55 ± 0.10 g·min-1, P = 0.021) . During PRO small but significant increases were seen in glucose absorption, plasma glucose and insulin concentration and decreases in NEFA and glycerol. Differences between markers of GI damage and permeability and time trial performance were not significant (P > 0.05). In contrast to the hypothesis, PRO led to minimal increases in absorption and oxidation of the ingested maltodextrin and small reductions in fat oxidation, while having no effect on subsequent time trial performance.

This pilot study investigated the effects of chilling and reheating a pasta-based meal on the postprandial glycaemic response. In this single-blind crossover study, 10 healthy volunteers consumed identical pasta meals (pasta, olive oil and tomato sauce), served either freshly prepared, chilled or chilled/reheated, on three separate randomised occasions. Capillary blood samples were taken for two hours postprandially. A significant difference in glucose Incremental Area Under the Curve (IAUC) was observed (p = 0.006), with the greatest difference observed between the freshly cooked and chilled/reheated meals (p = 0.041). Significant differences in incremental peak glucose were also observed (p = 0.018). These results suggest that making simple changes to domestic food processing methods can reduce the glycaemic excursion following a pasta meal, with the potential for health benefit.

Abstract: Dietary fructose has been linked to an increased post-prandial triglyceride (TG) level, which is an established independent risk factor for cardiovascular disease. Although much research has focused on the effects of fructose consumption on liver-derived very-low density lipoprotein (VLDL), emerging evidence also suggests that fructose may raise post-prandial TG levels by affecting the metabolism of enterocytes of the small intestine. Enterocytes have become well recognised for their ability to transiently store lipids following a meal and to thus control post-prandial TG levels according to the rate of chylomicron (CM) lipoprotein synthesis and secretion. The influence of fructose consumption on several aspects of enterocyte lipid metabolism are discussed, including de novo lipogenesis, apolipoprotein B48 and CM-TG production, based on the findings of animal and human isotopic tracer studies. Methodological issues affecting the interpretation of fructose studies conducted to date are highlighted, including the accurate separation of CM and VLDL. Although the available evidence to date is limited, disruption of enterocyte lipid metabolism may make a meaningful contribution to the hypertriglyceridaemia often associated with fructose consumption.

The purpose of this study was to determine whether or not partial acylglycerols, resulting from triacylglycerol hydrolysis, accumulate during the process of fat deposition in vivo. We measured mono-, di- and triacylglycerol levels in arterialized and adipose tissue venous plasma in normal subjects before and after a test meal. The mean concentrations of partial acylglycerols were consistently low in all plasma samples, accounting for less than 2% of the total acylglycerol measurement. The absence of monoacylglycerol accumulation in the adipose tissue venous plasma samples indicates that the disposal of monoacylglycerol by the adipose tissue was not rate limiting for complete hydrolysis of triacylglycerol in vivo. After heparin was injected intravenously 300 min after the high-fat meal, the mean concentration of monoacylglycerol in arterialized plasma increased from 1.6 ± 0.6 μmol/l to 106 ± 19 μmol/l (P = 0.01), whereas the mean concentration of diacylglycerol did not change (13.0 ± 2.3 μmol/l before, 17.4 ± 4.2 μmol/l after).

Dietary sugars are linked to the development of non-alcoholic fatty liver disease (NAFLD) and dyslipidaemia, but it is unknown if NAFLD itself influences the effects of sugars on plasma lipoproteins. To study this further, men with NAFLD (n=11) and low liver fat ‘controls’ (n= 14) were fed two iso-energetic diets, high or low in sugars (26% or 6% total energy) for 12 weeks, in a randomised, cross-over design. Fasting plasma lipid and lipoprotein kinetics were measured after each diet by stable isotope trace-labelling. There were significant differences in the production and catabolic rates of VLDL subclasses between men with NAFLD and controls, in response to the high and low sugar diets. Men with NAFLD had higher plasma concentrations of VLDL1-triacylglycerol (TAG) after the high (P

Objective. To investigate the mechanism for increased ketogenesis following treatment with SGLT2 inhibitor, dapagliflozin in people with type 2 diabetes. Research, Design & Methods. This was a double-blind placebo-controlled crossover study with a 4-week washout period. Participants received dapagliflozin or placebo in random order for 4 weeks. After each treatment, they ingested 30ml of olive oil containing [U-13C] palmitate to measure ketogenesis with blood sampling for 480 min. Stable isotopes of glucose and glycerol were infused to measure glucose flux and lipolysis respectively at 450-480 min. Results. Glucose excretion rate was higher and peripheral glucose uptake lower with dapagliflozin than placebo. Plasma beta-hydroxybutyrate (BOHB) concentrations and [13C2] BOHB concentrations were higher and glucose concentrations lower with dapagliflozin than placebo. Non-esterified fatty acids (NEFA) were higher with dapagliflozin at 300 and 420 min but lipolysis at 450-480 min was not different. Triacylglycerol (TAG) at all time points and endogenous glucose production rate at 450-480 min were not different between treatments. Conclusions. The increase in ketone enrichment from the ingested palmitic acid tracer suggests meal derived fatty acids contribute to the increase in ketones during treatment with dapagliflozin. The increase in BOHB concentration with dapagliflozin, occurred with only minimal changes in plasma NEFA concentration and no change in lipolysis. This suggests a metabolic switch to increase ketogenesis within the liver.

Objectives: To comprehensively survey the sugar and nutrient contents of yogurt products available in UK supermarkets, in particular those marketed to children. Design: A cross-sectional survey of yogurt products available in the UK’s supermarkets in November 2016. Methods: Data were collected from five major online UK supermarkets and a process flow strategy was used to place yogurts into eight categories: children’s, dairy alternatives, dessert, drinks, fruit, flavoured, natural/Greek style and organic. A comprehensive database of product information for 921 unique products was created and analysed. Results: The total sugar, fat, protein, calcium and energy contents were highly variable across categories, and the ranges were extremely broad. Although lower than the dessert category, the medians (range) of the total sugar content of children’s (10.8 g/100 g (4.8–14.5)), fruit (11.9 g/100 g (4.6–21.3)), flavoured (12.0 g/100 g (0.1–18.8)) and organic (13.1 g/100 g (3.8–16.9)) yogurt products were all well above 10 g/100 g, and represented >45% of total energy. Only two out of 101 children’s yogurt and fromage frais products surveyed qualified as low sugar (≤5 g/100 g). Natural/Greek yogurts had dramatically lower sugar contents (5.0 g/100 g (1.6, 9.5), largely lactose) than all other categories. While low-fat (

Background: Interesterified (IE) fats are widely used in place of trans fats; however, little is known about their metabolism. Objective: To test the impact of a commonly consumed IE versus a non-IE equivalent fat on in vivo postprandial and in vitro lipid metabolism, compared with a reference oil (rapeseed oil; RO). Design: A double-blinded, 3-phase crossover, randomized controlled trial was performed in healthy adults (n=20) aged 45-75 years. Postprandial plasma triacylglycerol (TG) and lipoprotein responses (including stable isotope tracing) to a test meal (50g fat) were evaluated over 8 hours. The test fats were IE 80:20 palm stearin/palm kernel fat, an identical non-IE fat, and RO (control). In vitro, mechanisms of digestion were explored using a dynamic gastric model (DGM). Results: Plasma TG 8h incremental area under the curves were lower following non-IE versus RO (-1.7 mmol/L.h (95% confidence interval -3.3, -0.0)), but there were no differences between IE and RO nor IE and non-IE. Low density lipoprotein (LDL) particles were smaller following IE and non-IE versus RO (P=0.005). Extra, extra large (XXL)-, extra large (XL)- and large (L)- VLDL particle concentrations were higher following IE and non-IE versus RO at 6-8 h (P

A Nutrition Society member-led meeting was held on 9 January 2020 at The University of Surrey, UK. Sixty people registered for the event, and all were invited to participate, either through chairing a session, presenting a ‘3 min lightning talk’ or by presenting a poster. The meeting consisted of an introduction to the topic by Dr Barbara Fielding, with presentations from eight invited speakers. There were also eight lightning talks and a poster session. The meeting aimed to highlight recent research that has used stable isotope tracer techniques to understand human metabolism. Such studies have irrefutably shaped our current understanding of metabolism and yet remain a mystery to many. The meeting aimed to de-mystify their use in nutrition research.

A sensitive method has been developed to measure specific mono-, di- and triacylglycerol concentrations in human plasma, using thin-layer chromatography and enzymatic assay. The levels of partial acylglycerols in human plasma from fasting subjects were lower than previous reports had suggested and amounted to less than 3% of the total acylglycerols. After heparin injection the plasma monoacylglycerol concentration increased markedly (P < 0.01) while the triacylglycerol concentration decreased significantly (P < 0.001). The plasma diacylglycerol concentration did not change significantly although it increased as a percentage of the total (P < 0.05); after heparin partial acylglycerols accounted for more than 10% of the total. After a high-fat meal the plasma concentrations of di- and triacylglycerol increased approximately two-fold (P < 0.005) but no significant change was observed for mono-acylglycerol. The percentage contribution of partial acylglycerols was unchanged (2.6% fasting, 2.4% postprandially).

A high fructose intake exacerbates postprandial plasma triacylglycerol (TAG) concentration, an independent risk factor for cardiovascular disease, although it is unclear whether this is due to increased production or impaired clearance of triacylglycerol (TAG)-rich lipoproteins. We determined the in vivo acute effect of fructose on postprandial intestinal and hepatic lipoprotein TAG kinetics and de novo lipogenesis (DNL). Five overweight men were studied twice, 4 weeks apart. They consumed hourly mixed-nutrient drinks that were high-fructose (30% energy) or low-fructose (

There is net outward flow of fatty acids from adipose tissue in the fasted state but net inward flow and storage in the postprandial state. We investigated how this is regulated. Arteriovenous differences were measured across a subcutaneous adipose depot in six normal subjects before and for 5 h after a meal containing 80 g fat and 80 g carbohydrate. In five further experiments, insulin was infused at 40 mU · m -2 · min -1 from 30 min after the meal, clamping the plasma glucose. Net transcapillary fatty acid flow changed from negative (outward flow from tissue to capillaries) in the postabsorptive state to consistently positive (net inward flow, implying fat storage) after the meal despite continued net efflux of fatty acids into venous blood. In the 'clamped' experiments (with additional insulin), net fatty acid efflux in the venous blood was suppressed and positive transcapillary flux (storage) was more marked. Regulation of fatty acid flow appeared to depend on coordinated changes in hormone-sensitive lipase (HSL) and lipoprotein lipase (LPL) action and fatty acid esterification. Additional insulin caused no further suppression of HSL or activation of LPL but markedly stimulated fatty acid retention (presumed to represent esterification). In the absence of additional insulin, a high proportion of the fatty acids liberated by LPL are released into the venous plasma in both postabsorptive and postprandial states. We hypothesize that this 'loss' of fatty acids is necessary to give precise control to the pathway of fat storage.

Background Experimental data suggest that apolipoprotein (apo) C-II and C-III regulate triglyceride-rich lipoprotein (TRL) metabolism, but there are limited studies in humans. We investigated the metabolic associations of TRLs with apoC-II and apoC-III concentrations and kinetics in women. Material and methods The kinetics of plasma apoC-II, apoC-III and very low-density lipoprotein (VLDL) apoB-100 and triglycerides were measured in the postabsorptive state using stable isotopic techniques and compartmental modeling in 60 women with wide-ranging body mass index (19.5-32.9kg/m2). Results Plasma apoC-II and apoC-III concentrations were positively associated with the concentration of plasma triglycerides, VLDL1- and VLDL2- apoB-100 and triglyceride (all P

The menopause is accompanied by increased risk of obesity, altered body fat distribution, and decreased skeletal muscle mass. The resulting decrease in RMR should be accompanied by a compensatory change in energy balance to avoid weight gain. We aimed to investigate habitual energy intake and expenditure in pre- and post-menopausal women matched for abdominal obesity. We recruited fifty-one healthy Caucasian women, BMI >18.5 and < 35, aged 35 - 45 years (pre-menopausal, n=26) and 55-65 years (post-menopausal, n=25). Energy intake was measured using 3-day diet diaries and dietary fat quality assessed using adipose tissue fatty acid biomarkers. RMR was measured using indirect calorimetry, and total (TEE) and activity energy expenditure using a combined accelerometer and heart rate monitor. Post-menopausal women had lower RMR and TEE and spent significantly less time undertaking moderate exercise than pre-menopausal women. Post-menopausal women had a tendency for a lower calorie intake, and a similar macronutrient intake but a significantly lower adipose tissue n-6/n-3 index (24.6 (1.6) v 37.7 (3.1), P

Mycobacterium tuberculosis (Mtb) infects macrophages and macrophage-derived foam cells, a hallmark of granulomata in tuberculous lesions. We analysed the effects of lipid accumulation in human primary macrophages and quantified strong triglyceride and phospholipid remodelling which depended on the dietary fatty acid used for the assay. The enrichment of >70% in triglyceride and phospholipids can alter cell membrane properties, signalling and phagocytosis in macrophages. In conventional macrophage cultures, cells are heterogeneous, small or large macrophages. In foam cells, a third population of 30% of cells with increased granularity can be detected. We found that foam cell formation is heterogenous and that lipid accumulation and foam cell formation reduces the phagocytosis of Mtb. Under the conditions tested, cell death was highly prevalent in macrophages, whereas foam cells were largely protected from this effect. Foam cells also supported slower Mtb replication, yet this had no discernible impact on the intracellular efficacy of four different antitubercular drugs. Foam cell formation had a significant impact in the inflammatory potential of the cells. TNF-α, IL-1β and prototypical chemokines were increased. The ratio of inflammatory IL-1β, TNF-α and IL-6 versus anti-inflammatory IL-10 was significantly higher in response to Mtb versus LPS, and was increased in foam cells compared to macrophages, suggestive of increased pro-inflammatory properties. Cytokine production correlated with NF-κB activation in our models. We conclude that foam cell formation reduces the host cell avidity for, and phagocytosis of, Mtb while protecting the cells from death. This protective effect is associated with enhanced inflammatory potential of foam cells and restricted intracellular growth of Mtb.

Purpose of review In 2004, the ‘omega-3 index’ was described as the sum of eicosapentaenoic acid (EPA, 20:5 n-3) and docosahexaenoic acid (DHA, 22:6 n-3) in red blood cells (RBCs) as an index of coronary heart disease mortality. This review outlines new evidence to support the omega-3 index as a tool to inform disease prognosis. Recent findings Recent studies have reported differential metabolism of EPA and DHA. High dose supplementation with EPA and DHA led to increased levels of RBC DHA that were associated with decreased liver fat. EPA and DHA in RBCs were associated with reduced mortality in a prospective study of patients with cardiac disease; the strongest association was with EPA. A diet containing 9.5 g alpha linolenic acid lead to an increase in EPA but not DHA status in middle aged women. Summary: Dietary intake or supplementation studies with n-3 fatty acids should include measurement of n-3 status in a standardised way. The omega-3 index, reflecting EPA and DHA status throughout the body, is convenient and may be appropriate in some cases, but since EPA and DHA assimilate differently in membranes, and have different potency, measurement of individual fatty acid composition in RBCs may be more informative

Different lipid fractions in humans have characteristic fatty acid profiles and these are maintained partly through diet and to a lesser extent through endogenous synthesis. The enzyme stearoyl-CoA desaturase (SCD; EC 1.14.99.5) is the rate-limiting enzyme in the synthesis of monounsaturated fatty acids such as palmitoleic acid (16:1 n-7) and oleic acid (18:1 n-9). These are the two most abundant monounsaturated fatty acids in human plasma lipids, membranes and adipose tissue. Although in quantitative terms, the endogenous synthesis of fatty acids in humans is not great in most circumstances, it is becoming increasingly evident that SCD plays important structural and metabolic roles. In addition, 16:1 n-7 has been purported to act as a beneficial 'lipokine' in an animal model. Research in humans has relied on indirect measurements of SCD1 activity and therefore, much of our understanding has come from work on animal models. However, results have been somewhat counterintuitive and confusing, so the purpose of this review is to try to summarise our current understanding of this fascinating enzyme.