Mycoprotein reduces energy intake and postprandial insulin release without altering glucagon-like peptide-1 and peptide tyrosine-tyrosine concentrations in healthy overweight and obese adults: a randomised-controlled trial.

Dietary mycoprotein decreases energy intake in lean individuals. The effects in overweight individuals are unclear, and the mechanisms remain to be elucidated. This study aimed to investigate the effect of mycoprotein on energy intake, appetite regulation, and the metabolic phenotype in overweight and obese volunteers. In two randomised-controlled trials, fifty-five volunteers (age: 31 (95 % CI 27, 35) years), BMI: 28·0 (95 % CI 27·3, 28·7) kg/m2) consumed a test meal containing low (44 g), medium (88 g) or high (132 g) mycoprotein or isoenergetic chicken meals. Visual analogue scales and blood samples were collected to measure appetite, glucose, insulin, peptide tyrosine-tyrosine (PYY) and glucagon-like peptide-1 (GLP-1). Ad libitum energy intake was assessed after 3 h in part A (n 36). Gastric emptying by the paracetamol method, resting energy expenditure and substrate oxidation were recorded in part B (n 14). Metabonomics was used to compare plasma and urine samples in response to the test meals. Mycoprotein reduced energy intake by 10 % (280 kJ (67 kcal)) compared with chicken at the high content (P=0·009). All mycoprotein meals reduced insulin concentrations compared with chicken (incremental AUClow (IAUClow): -8 %, IAUCmedium: -12 %, IAUChigh: -21 %, P=0·004). There was no significant difference in glucose, PYY, GLP-1, gastric emptying rate and energy expenditure. Following chicken intake, paracetamol-glucuronide was positively associated with fullness. After mycoprotein, creatinine and the deamination product of isoleucine, α-keto-ß-methyl-N-valerate, were inversely related to fullness, whereas the ketone body, ß-hydroxybutyrate, was positively associated. In conclusion, mycoprotein reduces energy intake and insulin release in overweight volunteers. The mechanism does not involve changes in PYY and GLP-1. The metabonomics analysis may bring new understanding to the appetite regulatory properties of food.

Improved diagnostic accuracy for neuroendocrine neoplasms using two chromogranin A assays.

CONTEXT: Chromogranin A (Cg A) is the best available diagnostic marker for neuroendocrine neoplasms (NENs). However, clinical interpretation of Cg A results may be limited by considerable heterogeneity between commonly available Cg A assays. Variation in diagnostic accuracy of these assays largely reflects differences in antibody specificities. We compared the diagnostic utility of four Cg A assays [Imperial Supra-regional Assay Service radioimmunoassay (SAS) and three commercial assays, Cisbio, DAKO and Eurodiagnostica]. METHOD: Plasma Cg A was measured using these four assays in 125 patients with NENs, 41 patients with cancers other than NENs and 108 healthy controls. RESULT: There was no significant difference in diagnostic accuracy between any of the four assays alone and no single assay positively identified all patients with NEN. However, concordance between assays was variable. Cisbio and SAS assays were least concordant. We, therefore, hypothesized that using a combination of the least concordant Cg A assays will improve NEN diagnosis by detecting a larger number of Cg A epitopes and hence patients with NEN. Consistent with our hypothesis, multiple logistic regression analysis showed that the combination of Cisbio and SAS assays was more useful than any other combinations or any assay alone in predicting a NEN diagnosis. CONCLUSION: Although individually, all four Cg A assays are similarly useful for the measurement of Cg A in the diagnosis of a NEN, in patients with a suspected NEN, negative results by one assay should prompt analysis by a second assay. The combination of Cisbio and SAS assays may have greatest diagnostic utility.

Gastric emptying, glucose metabolism and gut hormones: evaluation of a common preoperative carbohydrate beverage.

OBJECTIVE: To study the gastric-emptying rate and gut hormonal response of two carbohydrate-rich beverages. A specifically designed carbohydrate-rich beverage is currently used to support the surgical patient metabolically. Fruit-based beverages may also promote recovery, due to natural antioxidant and carbohydrate content. However, gastric emptying of fluids is influenced by its nutrient composition; hence, safety of preoperative carbohydrate loading should be confirmed. Because gut hormones link carbohydrate metabolism and gastric emptying, hormonal responses were studied. METHODS: In eight volunteers, gastric emptying rates of both 400 mL of a ready-to-use beverage (A: Nutricia preOp; 50.4 g carbohydrates-mainly polysaccharides; 260 mOsm/kg) and 400 mL over-the-counter fruit-based lemonade (B: Roosvicee Original; 48 g carbohydrates--mainly fruit-associated saccharides; 805 mOsm/kg) were determined scintigraphically (using hepatate Tc-99(m)) according to a crossover design. Plasma glucose, insulin, C-peptide, glucagon-like peptide (GLP-1), peptide YY, total glucagon, and ghrelin were studied. RESULTS: Gastric emptying showed no differences in residual volumes. Earlier onset in emptying for beverage A versus B was observed (trend), with significantly higher glucose, insulin, C-peptide, and glucagon responses at 15-90 min. GLP-1 was inversely related to residual volume. CONCLUSION: Fruit-based lemonade is a safe alternative for preoperative purposes. It induces a more limited glucose, insulin, and C-peptide response. Later onset in gastric emptying (B versus A: trend), lower glucagon release, and differences in beverage content and osmolarity may have contributed to those differences. Efficient emptying was reflected by early GLP-1 levels.

Gut hypertrophy after gastric bypass is associated with increased glucagon-like peptide 2 and intestinal crypt cell proliferation.

OBJECTIVE: We aimed to determine changes in crypt cell proliferation and glucagon-like peptide-2 (GLP-2) in rodents and man after Roux-en-Y gastric bypass (RYGB). SUMMARY OF BACKGROUND DATA: Roux-en-Y gastric bypass results in sustained weight loss and reduced appetite with only mild gastrointestinal side effects. Glucagon-like peptide-2 released from intestinal l-cells after nutrient intake stimulates intestinal crypt cell proliferation and mitigates the effects of gut injury. METHODS: Wistar rats underwent either RYGB (n = 6) or sham procedure (n = 6) and plasma GLP-2, GLP-1, and gut hormone peptide YY (PYY) were measured after 23 days. Biopsies from the terminal ileum were stained using the antibody to Ki67, which detects cyclins and hence demonstrates cells in the S-phase of the cell cycle. The total number of cells, number of mitosis, and number of labeled cells per crypt were counted. Obese patients (n = 6) undergoing RYGB were evaluated following a 420 kcal meal preoperatively, and 1, 3, 6, 12, and 24 months later for responses in l-cell products such as GLP-2, GLP-1, total PYY, and PYY3-36. RESULTS: Rat GLP-2 levels after RYGB were elevated 91% above sham animals (P = 0.02). At necropsy, mitotic rate (P < 0.001) and cells positive for the antibody Ki67 (P < 0.001) were increased, indicating crypt cell proliferation. Human GLP-2 after RYGB reached a peak at 6 months of 168% (P < 0.01) above preoperative values. Area under the curve for GLP-1 (P < 0.0001), total PYY (P < 0.01), and PYY3-36 (P < 0.05) responses increased progressively over 24 months. CONCLUSIONS: RYGB leads to increased GLP-2 and mucosal crypt cell proliferation. Other gut hormones from l-cells remain elevated for at least 2 years in humans. These findings may account for the restoration of the absorptive surface area of the gut, which limits malabsorption and contributes to the long-term weight loss after RYGB.

Kisspeptin is released from human prostate cancer cell lines but plasma kisspeptin is not elevated in patients with prostate cancer.

Kisspeptin, the product of the KiSS-1 gene, inhibits metastasis and stimulates the hypothalamo-pituitary-gonadal axis. Kisspeptin is therefore a putative target in the treatment of hormone-sensitive malignancies. Prostatic carcinoma remains a significant cause of mortality despite improvements in therapy. The role of kisspeptin in prostatic carcinoma remains undefined. We therefore aimed to investigate release of kisspeptin by prostatic cancer cell lines; investigate expression of KiSS-1 in human prostate tissue; investigate whether patients with prostate carcinoma have elevated plasma kisspeptin. 1) Culture medium from prostatic carcinoma cell lines LNCaP, DU145 and PC3 was assayed for kisspeptin immunoreactivity (-IR). Kisspeptin-IR release was detectable from all three cell lines. The effect of hydroxyflutamide, gefitinib and resveratrol on kisspeptin-IR release from these cell lines was also investigated. No effect of the drugs tested on release of kisspeptin-IR was observed. 2) Expression of KiSS-1 in human prostate tissue (n=4) was investigated using in situ hybridisation. Expression of KiSS-1 was detected in human prostate tissue. 3) Plasma kisspeptin-IR was compared in 92 patients with prostatic carcinoma and 73 male controls. Kisspeptin-IR was not detected in the plasma of either patients with prostate cancer or control patients. We have therefore shown for the first time the release of kisspeptin-IR by prostatic carcinoma cell lines. We have also shown that KiSS-1 is expressed in human prostate tissue, and that circulating levels of kisspeptin-IR are not elevated in patients with prostatic carcinoma. Further work is required to determine the role of kisspeptin in the prostate.

The hyperphagic effect of ghrelin is inhibited in mice by a diet high in fat.

BACKGROUND & AIMS: Ghrelin is the only peripheral hormone known to increase food intake. It is released from the stomach and is thought to function as a signal of energy deficit and a meal initiator. We generated transgenic mice in which levels of bioactive ghrelin are increased in the stomach and circulation. These mice, as expected, are hyperphagic and glucose intolerant. We investigated whether exposure to a high-fat diet (HFD) would exacerbate this phenotype. METHODS: We investigated the effect of HFD on energy and glucose homeostasis in ghrelin transgenic mice. We determined dietary preference; expression of hypothalamic neuropeptides that control food intake; and, using fast-performance liquid chromatography, the circulating forms of ghrelin. We measured food intake during continuous administration of ghrelin in wild-type mice fed either regular chow or an HFD. RESULTS: Ghrelin transgenic mice were resistant to diet-induced obesity because of their reduced food intake. This was not caused by alterations to food preference, hypothalamic signaling of neuropeptides that control food intake, or the form of circulating acylated ghrelin. Long-term administration of ghrelin to wild-type mice failed to increase ingestion of an HFD but, as expected, increased intake of regular chow. CONCLUSIONS: This is the first report that diets high in fat inhibit the hyperphagic effect of ghrelin; these findings indicate that features of the diet are important determinants of ghrelin's function. This information is important for the development of anti-obesity drugs that target ghrelin signaling.

Ghrelin and peptide YY (PYY) profiles in gastrointestinal tissues and the circulation of the rat during pregnancy and lactation.

Plasma and tissue profiles of gastrointestinal hormones ghrelin and peptide YY (PYY) were investigated in different female rat reproductive states. Neither plasma nor tissue ghrelin concentrations were suppressed during pregnancy despite elevated leptin. The highest concentrations of stomach ghrelin were measured in late pregnancy. PYY concentrations in plasma, descending colon and rectum tissues were increased (P<0.001) throughout pregnancy and lactation. PYY peaked at day 5 of lactation in plasma, as well as descending colon and rectum tissues (proestrus vs day 5 of lactation: 25+/-3.0 pmol/l vs 55+/-8.0 pmol/l; 85+/-4.5 pmol/g wwt vs 418+/-45.0 pmol/g wwt; 23+/-3.0 pmol/g wwt vs 78+/-9.1 pmol/g wwt). This PYY peak was temporally associated with the luteinizing hormone peak on day 1 of lactation. Following weaning, dam adiposity and plasma leptin increased whereas ghrelin stomach peptide decreased. Relative PYY concentrations in the tissues of the gut varied in the different states suggesting regional alterations taking place in the colon. The ascending colon produced the highest concentrations in non-pregnant rats, the descending colon the highest concentrations during lactation with the pregnant rats and the dams postweaning in a transition state between. It is unclear what role the increased PYY in various tissues observed has during pregnancy and lactation as it would be expected to be reduced in these states of greatly increased appetite. PYY may have an influence on maternal dietary adaptation, intestinal hypertrophy and weight gain during pregnancy and lactation although it is still unclear precisely how it acts.

Mice with hyperghrelinemia are hyperphagic and glucose intolerant and have reduced leptin sensitivity.

OBJECTIVE: Ghrelin is the only known peripheral hormone to increase ingestive behavior. However, its role in the physiological regulation of energy homeostasis is unclear because deletion of ghrelin or its receptor does not alter food intake or body weight in mice fed a normal chow diet. We hypothesized that overexpression of ghrelin in its physiological tissues would increase food intake and body weight. RESEARCH DESIGN AND METHODS: We used bacterial artificial chromosome transgenesis to generate a mouse model with increased ghrelin expression and production in the stomach and brain. We investigated the effect of ghrelin overexpression on food intake and body weight. We also measured energy expenditure and determined glucose tolerance, glucose stimulated insulin release, and peripheral insulin sensitivity. RESULTS: Ghrelin transgenic (Tg) mice exhibited increased circulating bioactive ghrelin, which was associated with hyperphagia, increased energy expenditure, glucose intolerance, decreased glucose stimulated insulin secretion, and reduced leptin sensitivity. CONCLUSIONS: This is the first report of a Tg approach suggesting that ghrelin regulates appetite under normal feeding conditions and provides evidence that ghrelin plays a fundamental role in regulating beta-cell function.

Elevated basal and post-feed glucagon-like peptide 1 (GLP-1) concentrations in the neonatal period.

BACKGROUND: Glucagon-like peptide 1 (GLP-1) is an incretin hormone that stimulates glucose-induced insulin secretion, increases beta-cell proliferation, neogenesis and beta-cell mass. In adults, plasma concentrations of amidated GLP-1 are typically within the 5-10 pmol/l range in the fasting state and increases to approximately 50 pmol/l after ingestion of a mixed meal. RESEARCH DESIGN AND METHODS: We measured plasma glucose, insulin and amidated forms of GLP-1 prefeed and then at 20 and 60 min post-feed following ingestion of a 60-70 ml of standard milk feed in preterm (n=10, 34-37 weeks) and term newborn infants (n=12, 37-42 weeks). Reverse-phase fast protein liquid chromatography was used to characterise the molecular nature of the circulating GLP-1. RESULTS: Mean birth weight was 3.18 kg and mean age at sampling for GLP-1 was 7.7 days. The mean basal GLP-1 concentration was 79.1 pmol/l, which increased to 156.6 pmol/l (+/-70.9, P<0.001) and 121.5 pmol/l (+/-59.2) at 20 and 60 min respectively. Reverse-phase chromatography analysis suggested that the majority of GLP-1 immunoreactivity (>75%) represented GLP-1 (7-36) amide and (9-36) amide. CONCLUSIONS: Basal and post-feed amidated GLP-1 concentrations in neonates are grossly raised with the major fractions of circulating GLP-1 being (7-36) amide and (9-36) amide. Elevated GLP-1 concentrations in the newborn period may have a role in regulating maturation of enteroendocrine system and also of increasing pancreatic beta-cell mass and regeneration. The high levels of GLP-1 may be due to immaturity of the dipeptidyl peptidase IV and or lower glomerular filtration rate in the neonatal period. Further studies are required to understand the role of GLP-1 in the neonatal period.

PYY3-36 injection in mice produces an acute anorexigenic effect followed by a delayed orexigenic effect not observed with other anorexigenic gut hormones.

Peptide YY (PYY) is secreted postprandially from the endocrine L cells of the gastrointestinal tract. PYY(3-36), the major circulating form of the peptide, is thought to reduce food intake in humans and rodents via high-affinity binding to the autoinhibitory neuropeptide Y (NPY) receptor within the arcuate nucleus. We studied the effect of early light-phase injection of PYY(3-36) on food intake in mice fasted for 0, 6, 12, 18, 24, and 30 h and show that PYY(3-36) produces an acute anorexigenic effect regardless of the duration of fasting. We also show evidence of a delayed orexigenic effect in ad libitum-fed mice injected with PYY(3-36) in the early light phase. This delayed orexigenic effect also occurs in mice administered a potent analog of PYY(3-36), d-Allo Ile(3) PYY(3-36), but not following injection of other anorectic agents (glucagon-like-peptide 1, oxyntomodulin, and lithium chloride). Early light-phase injection of PYY(3-36) to ad libitum-fed mice resulted in a trend toward increased levels of hypothalamic NPY and agouti-related peptide mRNA and a decrease in proopiomelanocortin mRNA at the beginning of the dark phase. Furthermore, plasma levels of ghrelin were increased significantly, and there was a trend toward decreased plasma PYY(3-36) levels at the beginning of the dark phase. These data indicate that PYY(3-36) injection results in an acute anorexigenic effect followed by a delayed orexigenic effect.

Postprandial ghrelin, cholecystokinin, peptide YY, and appetite before and after weight loss in overweight women with and without polycystic ovary syndrome.

BACKGROUND: Polycystic ovary syndrome (PCOS) is a common condition associated with obesity and with reproductive and metabolic dysfunction. Abnormalities in appetite regulation in PCOS patients may contribute to difficulties in weight management. OBJECTIVE: We aimed to examine appetite, appetite hormones, and ad libitum food consumption before and after weight loss in overweight women with and without PCOS. DESIGN: Overweight age- and weight-matched women with (n = 14) and without (n = 14) PCOS undertook an 8-wk energy-restricted diet (5185.3 +/- 141.6 kJ/d). At baseline and study end, subjects consumed a test meal (936 kJ; 25% of energy from protein, 9% from fat, and 67% from carbohydrate). Subjective appetite and circulating glucose, insulin, ghrelin, cholecystokinin, and peptide YY were assessed at 0, 15, 30, 45, 60, 90, 120, and 180 min. A mixed buffet lunch was then offered to assess ad libitum food intake. RESULTS: Weight loss (4.2 +/- 3.9 kg) did not differ significantly between the 2 groups. Women with PCOS had significantly (P = 0.023) lower ghrelin concentrations before and after weight loss than did women without PCOS. The degree of postprandial ghrelin suppression was lower at weeks 0 (P = 0.048) and 8 (P = 0.069) in women with PCOS than in women without PCOS. There were no significant differences between the 2 groups in appetite responses, buffet consumption, or fasting or postprandial peptide YY and cholecystokinin before or after weight loss. CONCLUSIONS: PCOS was associated with lower fasting ghrelin and a smaller postprandial ghrelin suppression both before and after weight loss but was not associated with other postprandial gut peptides, subjective satiety, or food intake. It is not clear whether appetite regulation is impaired in PCOS.

Relation of fasting plasma peptide YY to glucose metabolism and cardiovascular risk factors after restrictive bariatric surgery.

INTRODUCTION: Surgically induced weight loss results in reduction of comorbidities in severely obese humans. Reversal of abnormal secretion of appetite-regulating gut hormones such as peptide YY (PYY) could be contributing to the improvement of cardiovascular risk factors. METHODS: Severely obese patients (n = 42, BMI = 45.7 +/- 5.3 kg/m(2)) underwent clinical examination and blood sampling for measurement of PYY, plasma lipids, oral glucose tolerance testing and assessment of insulin secretion (HOMA-%B) and action (HOMA-R, QUICKI) before and during 12 months following gastric banding. Comparisons were made at each time point of the study as well as across the total study period. RESULTS: Weight loss after bariatric surgery resulted in improvement of insulin resistance by 54% (p < 0.03) and plasma triglycerides by 26% (p < 0.01) without changes in fasting PYY (16.2 +/- 8.7 pmol/l at baseline, 15.1 +/- 6.3 pmol/l at 12 months). Fasting PYY correlated negatively with plasma total cholesterol at baseline (p = 0.02) but was not associated with body weight, body mass or abdominal diameter. Individual changes in PYY (DeltaPYY) related to changes in insulin (Deltafasting insulin) at 12 months (r = -0.582, p = 0.02) and HOMA-B at 6 months (r = -0.677, p = 0.006) and 12 months (r = -0.660, p = 0.007). Diabetic status had no impact on these correlations. DISCUSSION: PYY correlates with a major cardiovascular risk factor and surrogate parameters of insulin secretion but not to weight loss or body mass in severe obesity.

Intestinal growth in parenterally-fed rats induced by the combined effects of glucagon-like peptide 2 and epidermal growth factor.

BACKGROUND: Parenteral nutrition and the absence of luminal feeding result in impaired intestinal growth and differentiation of enterocytes. Glucagon-like peptide 2 (GLP-2) and epidermal growth factor (EGF) have each been shown to have trophic effects on the intestine, and thus have the potential to benefit patients fed parenterally, such as those with intestinal failure from short bowel syndrome. We report studies aimed to determine whether there may be synergistic effects of these 2 peptides. METHODS: Rats were established on parenteral nutrition (PN) and infused for 6 days with GLP-2 (20 microg/d), EGF (20 microg/d), or GLP-2 + EGF (20 microg/d of each). These groups were compared with untreated PN-fed and orally-fed controls. Tissue was obtained from small intestine and colon to determine growth, proliferation, and representative gene expression. RESULTS: Small intestinal weight was increased by 75%, 43%, and 116% in the GLP-2, EGF, and GLP-2 + EGF groups, respectively, compared with PN controls (all p < .001). Cell proliferation increased with GLP-2, EGF, and GLP-2 + EGF in proximal small intestine by factors of 2.3, 1.7, and 3.4 respectively (p < .001). A synergistic effect on villous and crypt area was observed in the proximal small intestine when GLP-2 and EGF were combined (p < .05). GLP-2 had no effect in the colon, unlike EGF. Further studies showed GLP-2 + EGF significantly increased expression in distal small intestine of transcripts for the bile acid transport protein IBABP (p < .05) and showed a significant correlation between the expression of IBABP and the transcription factor HNF-4. CONCLUSIONS: Both GLP-2 and EGF upregulate growth of the small intestine, and this is augmented when GLP-2 and EGF are combined. These findings may lead to improved treatment of patients receiving PN.

Hypothalamic cocaine- and amphetamine-regulated transcript (CART) and agouti-related protein (AgRP) neurons coexpress the NOP1 receptor and nociceptin alters CART and AgRP release.

Nociceptin or orphanin FQ (N/OFQ) and its receptor NOP1 are expressed in hypothalamic nuclei involved in energy homeostasis. N/OFQ administered by intracerebroventricular or arcuate nucleus (ARC) injection increases food intake in satiated rats. The mechanisms by which N/OFQ increases food intake are unknown. We hypothesized that N/OFQ may regulate hypothalamic neurons containing peptides involved in the control of food intake such as cocaine- and amphetamine-regulated transcript (CART), alphaMSH, neuropeptide Y (NPY), and agouti-related protein (AgRP). We investigated the ability of N/OFQ to alter the release of CART, alphaMSH, NPY, and AgRP using ex vivo medial basal hypothalamic explants. Incubation of hypothalamic explants with N/OFQ (1, 10, 100 nM) resulted in significant changes in CART and AgRP release. One hundred nanomoles N/OFQ caused a 33% decrease in release of CART (55-102) immunoreactivity (IR) and increased release of AgRP-IR to 163% but produced no change in either alphaMSH-IR or NPY-IR. Double immunocytochemistry/in situ hybridization demonstrated that CART-IR and NOP1 mRNA are colocalized throughout the hypothalamus, in particular in the paraventricular nucleus, lateral hypothalamus, zona incerta, and ARC, providing an anatomical basis for N/OFQ action on CART release. Dual in situ hybridization demonstrated that AgRP neurons in the ARC also express the NOP1 receptor. Our data suggest that nociceptin via the NOP1 receptor may increase food intake by decreasing the release of the anorectic peptide CART and increasing the release of the orexigenic peptide AgRP.

Impaired circulating glucagon-like peptide-1 response to oral glucose in women with previous gestational diabetes.

BACKGROUND AND OBJECTIVE: Women with previous gestational diabetes (pGDM) are at risk of developing Type 2 diabetes. Glucagon-like peptide-1 (GLP-1) potentiates the insulin response to oral glucose, and its secretion is diminished in Type 2 diabetes. The aim of the study was to see if decreased GLP-1 secretion might be an early abnormality in the progression to Type 2 diabetes and would therefore be diminished in women with pGDM. PATIENTS AND METHODS: Eleven women with pGDM and previously documented normal glucose tolerance and 11 control women underwent a 75 g oral glucose tolerance test (OGTT). Circulating plasma glucose, insulin, nonesterified fatty acids (NEFA) and GLP-1 concentrations were sampled. RESULTS: One of the women with pGDM had impaired glucose tolerance and was excluded from the study. All other women had normal glucose tolerance. The women with pGDM had higher fasting glucose concentrations than controls (5.1; 4.9-5.3 vs. 4.8; 4.4-5.1 mmol/l, median; interquartile range, P = 0.04) and greater circulating glucose area under the curve (AUC) following the oral glucose load (930; 818-1015 vs. 668; 584-737 min x mmol/l, P = 0.0007). Fasting insulin concentrations and total insulin AUC were similar. The initial (0-30 min) insulin response was decreased in the pGDM women (AUC 3981; 2783-4795 vs. 6167; 5009-8145 min x pmol/l, P = 0.05). The initial (0-30 min) GLP-1 response was reduced in the pGDM women (AUC 816; 663-984 vs. 1163; 872-2024 min x pmol/l, P = 0.02). CONCLUSION: A reduced initial GLP-1 response to oral glucose may therefore be an early abnormality in the progression to Type 2 diabetes.

Evidence against redox regulation of energy homoeostasis in humans at high altitude.

The present study examined if free radicals and associated inflammatory sequelae influenced metabolic biomarkers involved in the neuro-endocrinological regulation of energy homoeostasis at high altitude. Sixteen mountaineers (11 males/five females) were matched for physical fitness and caloric intake and assigned in a double-blind manner to either antioxidant (n=8) or placebo (n=8) supplementation, which was enforced for 7 days at sea level and during an 11-day ascent to 4780 m. Enteral prophylaxis incorporated a daily bolus dose of 1 g of L-ascorbate, 400 international units of D,L-alpha-tocopherol acetate and 600 mg of alpha-lipoic acid. EPR (electron paramagnetic resonance) spectroscopic detection of PBN (alpha-phenyl-tert-butylnitrone) adducts confirmed an increase in the venous concentration of carbon-centred radicals at high altitude in the placebo group, whereas a decrease was observed in the antioxidant group (P<0.05 compared with that at sea level). EPR detection of DMSO/A*- (DMSO-supplemented ascorbate free radical) demonstrated that the increase in carbon-centred radicals at high altitude was associated with a decrease in ascorbate (r2=0.63; P<0.05). Ascent to high altitude (pooled placebo+antioxidant groups) also increased the expression of pro-inflammatory cytokines (P<0.05 compared with that at sea level) and biomarkers of skeletal tissue damage (P<0.05). Despite a general decrease in leptin, insulin and glucose at high altitude (pooled placebo+antioxidant groups; P<0.05 compared with that at sea level), persistent anorexia resulted in a selective loss of body fat (P<0.05). In conclusion, antioxidant prophylaxis decreased the concentration of carbon-centred radicals at high altitude (P<0.05 compared with the placebo group), but did not influence markers of inflammation, appetite-related peptides, ad libitum nutrient intake or body composition. Thus free radicals do not appear to be involved in the inflammatory response and subsequent control of eating behaviour at high altitude.

Investigation of the chronic effects of NPY by subcutaneous implantation of 6-23 cells producing NPY in WAG rats.

OBJECTIVE: In this experiment, we studied the chronic effects of NPY, as there were no data on long-term effects of NPY in vivo. METHODS: Complementary DNA encoding NPY was isolated, sequenced and cloned into the expression vector, pCEP4. The 6-23 clone 6 cell line was transfected with this clone. Two groups of 10 adult male WAG rats (180-250 g body weight) were injected with either untransfected 6-23 clone 6 or 6-23 clone 6 transfected with NPY cDNA [6-23 (NPY)]. After 8 weeks, the animals were killed, their plasma assayed for insulin. Pancreatic glucagon (PG), by RIA, and plasma glucose were measured. RESULTS: The transfected cells were shown to be producing fully processed, bioactive NPY. The expression of NPY was also confirmed by Northern blot analysis. The animals injected with 6-23 (NPY) cells gained significantly more weight than the controls, (on day 54, 31.89 +/- 3.56 vs. 24.1 +/- 4.12 g, n = 10, P < 0.05). Plasma insulin and PG increased significantly in NPY animals compared to controls. The total RNA extracted from tumours was analysed by Northern blotting and showed NPY mRNA expression in NPY animals, but not in controls. CONCLUSION: The long-term effects of NPY was confirmed by injection of the cells producing this peptide.

The relationship of ghrelin to biochemical and anthropometric markers of adult growth hormone deficiency.

INTRODUCTION: Ghrelin is the natural ligand of the growth hormone secretagogue receptor (GHS-R) and potently stimulates GH release in humans. Ghrelin is found in the hypothalamus, but most circulating ghrelin is derived from the stomach. Ghrelin stimulates food intake but circulating levels are low in obesity. We hypothesized that GH deficiency (GHD) might be associated with increased circulating ghrelin concentrations as a result of low GH levels. We therefore measured circulating ghrelin concentrations, leptin and body composition in subjects with GHD and healthy controls. METHODS: Subjects with GHD (n = 18) were compared to healthy control subjects (n = 18), matched for body mass index (BMI). They underwent assessment of body composition [waist circumference, BMI and percentage body fat (using bioimpedance)]. Plasma ghrelin, leptin, insulin, GH and IGF-1 were measured in the fasting state. Plasma ghrelin was measured using a specific radioimmunassay, and the other hormones using commercially available assays. RESULTS: The groups were well-matched for BMI (GHD vs. control; 32.9 +/- 10.8 vs. 31.3 +/- 11.7, P = ns) and waist circumference (GHD vs. control; 102.9 +/- 20.0 vs. 99.8 +/- 25.2, P = ns), but percentage body fat (GHD vs. control; 37.0 +/- 9.1 vs. 29.4 +/- 13.0, P = 0.06) tended to be higher in the GHD group. As expected, IGF-1 was lower in GHD (GHD vs. control; 12.5 +/- 6.8 vs. 19.2 +/- 5.8 nmol/l, P = 0.003). Ghrelin [GHD vs. controls; geometric mean (95% CI); 828.8 (95% CI 639.9-1074.2) vs. 487.9 (95% CI 297.2-800.2) pmol/l] and leptin [GHD vs. controls; 13.2 (95% CI 6.6-26.5) vs. 7.9 (95% CI 3.7-16.9) ng/ml] were similar in the two groups. Plasma ghrelin correlated inversely with waist circumference and waist hip ratio in GHD subjects (r = -0.6, P = 0.02) but not with IGF-1 or GH concentrations. There was no significant correlation in the control subjects. CONCLUSION: Circulating ghrelin concentrations are influenced by body fat distribution, but not by levels of either GH or IGF-1. However, given that obesity is associated with reduced ghrelin concentrations and that GHD is commonly associated with increased body fat, it is possible that these two opposing influences on circulating ghrelin levels result in normal concentrations in subjects with GHD.

A role for arcuate cocaine and amphetamine-regulated transcript in hyperphagia, thermogenesis, and cold adaptation.

We have recently shown that injection of the hypothalamic peptide cocaine and amphetamine regulated transcript (CART) into discrete hypothalamic nuclei stimulates food intake. This stimulation was particularly marked in the arcuate nucleus. Here we show that twice daily intra-arcuate injection of 0.2 nmole CART peptide for 7 days was associated with a 60% higher daytime food intake, an 85% higher thermogenic response to the beta3 agonist BRL 35135, and a 60% increase in brown adipose tissue UCP-1 mRNA. In a separate study, using stereotactically targeted gene transfer, a CART transgene was delivered by using polyethylenimine to the arcuate nucleus of adult rats. Food intake was increased significantly during ad libitum feeding and following periods of food withdrawal and food restriction in CART over-expressing animals. CART over-expressing animals lost 12% more weight than controls following a 24-h fast. Brown adipose tissue uncoupling protein-1 (UCP-1) mRNA levels (collected Day 25) were 80% higher in CART over-expressing animals. Finally, by using quantitative in situ hybridization, we found that chronic cold exposure (20 days at 4oC) increased arcuate nucleus CART mRNA by 124%. Together with the orexigenic and thermogenic effects of CART, this finding suggests a role for arcuate nucleus CART in cold adaptation.

A randomised four-intervention crossover study investigating the effect of carbohydrates on daytime profiles of insulin, glucose, non-esterified fatty acids and triacylglycerols in middle-aged men.

Postprandial concentrations of glucose, insulin and triacylglycerols (TG) correlate to risk for CHD. Carbohydrates affect many metabolites that could have a potential effect on cardiovascular risk factors. The objective of the present study was to examine, using a randomised prospective study, the acute (day 1) and ad libitum medium-term (day 24) effects of four diets: a high-fat diet (HIGH-FAT; 50 % fat, >34 % monounsaturated fatty acids); a low-glycaemic index (GI) diet (LOW-GI; high-carbohydrate, low-GI); a high-sucrose diet (SUCROSE; high carbohydrate increase of 90 g sucrose/d); a high-GI diet (HIGH-GI; high-carbohydrate, high-GI). Daytime profiles (8 h) (breakfast, lunch and tea) of lipid and carbohydrate metabolism were completed during day 1 and day 24. Seventeen middle-aged men with one or more cardiac risk factors completed the study. There was no change from day 1 or between diets in fasting glucose, lipids or homeostatic assessment model (HOMA) on day 24. The HIGH-FAT compared with the three high-carbohydrate diets was associated with lower postprandial insulin and glucose but higher postprandial TG and non-esterified fatty acids (NEFA). There was a significant increase in the 6 h (15.00 hours) TG concentration (day 1, 2.6 (sem 0.3) mmol/l v. day 24, 3.3 (sem 0.3) mmol/l; P<0.01) on the SUCROSE diet. Postprandial HOMA (i.e. incremental area under the curve (IAUC) glucose (mmol/l per min)xIAUC insulin/22.5 (mU/l per min)) median changes from day 1 to day 24 were -61, -43, -20 and +31 % for the HIGH-FAT, LOW-GI, SUCROSE and HIGH-GI diets respectively. The HIGH-GI percentage change was significantly different from the other three diets (P<0.001). Despite being advised to maintain an identical energy intake there was a significant weight change (-0.27 (sem 0.3) kg; P<0.02) on the LOW-GI diet compared with the SUCROSE diet (+0.84 (sem 0.3) kg). In conclusion the HIGH-FAT diet had a beneficial effect on postprandial glucose and insulin over time but it was associated with higher postprandial concentrations of TG and NEFA. Conversely the HIGH-GI diet appeared to increase postprandial insulin resistance over the study period.