The development of gut hormones as drugs for the treatment of obesity

Obesity is an ever-increasing problem with limited effective treatments available that are safe and tolerable. Gut hormones are naturally occurring endogenous satiety factors that are released in response to food consumption. As well as playing an important role in the regulation of appetite, food intake and body weight, they also have roles in glucose disposal, gastric motility, fuel-type utilisation and energy expenditure. These characteristics make gut hormones attractive drugable targets for the treatment of obesity. However, the vulnerability of gut hormones to degradative enzymes results in rapid clearance rates and limited bioactivity which limits their viability as anti-obesity therapies. This work aims to address two important properties to consider when developing gut hormones as drugs: cause of degradation and immunogenicity. Using pancreatic polypeptide (PP), I developed in vitro and in vivo testing systems to identify the physiologically important mechanisms involved in PP degradation. DPPIV and NEP were identified as important enzymes and enzyme-resistant analogues of PP were designed which demonstrated greater efficacy and slower clearance rates. Subcutaneous (SC) administration of foreign exogenous peptides is known to potentially cause immunogenic reactions against the foreign peptide. The dangers of this reaction can range from mild to lethal. Currently peptide therapeutics require SC administration as oral bioavailability is low. Therefore, an aim of my studies was to explore the impact of modifications to PYY with regard to immunogenicity. To do this I assessed the impact of changes to different regions and specific amino acids of PYY. I successfully identified a region of PYY which, when modified, caused immunogenicity and what substitutions resulted in the most changes to immunogenic properties. In summary, my work demonstrated that by understanding the causes of peptide clearance and immunogenicity, safer and more efficacious analogues of gut hormones can be designed to take forward as potential treatments for obesity.Open Acces

Glucose in the hypothalamic paraventricular nucleus regulates GLP-1 release.

Glucokinase (GK) is highly expressed in the hypothalamic paraventricular nucleus (PVN); however, its role is currently unknown. We found that GK in the PVN acts as part of a glucose-sensing mechanism within the PVN that regulates glucose homeostasis by controlling glucagon-like peptide 1 (GLP-1) release. GLP-1 is released from enteroendocrine L cells in response to oral glucose. Here we identify a brain mechanism critical to the release of GLP-1 in response to oral glucose. We show that increasing expression of GK or injection of glucose into the PVN increases GLP-1 release in response to oral glucose. On the contrary, decreasing expression of GK or injection of nonmetabolizable glucose into the PVN prevents GLP-1 release. Our results demonstrate that gluco-sensitive GK neurons in the PVN are critical to the response to oral glucose and subsequent release of GLP-1

The phototoxicity of aged human retinal melanosomes

The purpose of this study was to determine whether an age-related increase in photoreactivity of human retinal melanosomes (MS) can cause phototoxicity to retinal pigment epithelium (RPE) cells. MS were isolated post mortem from young (20–30 years, young human melanosomes [YHMs]) and old (60–90 years, old human melanosomes [OHMs]) human eyes and from young bovine eyes (bovine melanosomes [BMs]). Confluent cultured ARPE-19 cells were fed equivalent numbers of OHMs or BMs and accumulated similar amounts of melanin as determined by electron paramagnetic resonance assay. Cells with and without MS were either maintained in the dark or exposed to blue light for up to 96 h and assessed for alterations in cell morphology, cell viability and lysosomal integrity. Incubation of cells in dark in the presence of internalized MS or irradiation of cells with blue light in the absence or presence of BMs did not significantly affect cell viability. However, exposures to blue light in the presence of OHMs resulted in abnormal cell morphology, up to ∼75% decrease in mitochondrial activity, loss of lysosomal pH and cell death. OHMs contained significantly less melanin than YHMs, supporting the hypothesis that melanin undergoes degradation during RPE aging. Our results demonstrate that aged MS can be phototoxic to human RPE cells and support a contributing role of MS in RPE aging and in the pathogenesis of age-related macular degeneration

Glucose in the hypothalamic paraventricular nucleus regulates GLP-1 release.

Glucokinase (GK) is highly expressed in the hypothalamic paraventricular nucleus (PVN); however, its role is currently unknown. We found that GK in the PVN acts as part of a glucose-sensing mechanism within the PVN that regulates glucose homeostasis by controlling glucagon-like peptide 1 (GLP-1) release. GLP-1 is released from enteroendocrine L cells in response to oral glucose. Here we identify a brain mechanism critical to the release of GLP-1 in response to oral glucose. We show that increasing expression of GK or injection of glucose into the PVN increases GLP-1 release in response to oral glucose. On the contrary, decreasing expression of GK or injection of nonmetabolizable glucose into the PVN prevents GLP-1 release. Our results demonstrate that gluco-sensitive GK neurons in the PVN are critical to the response to oral glucose and subsequent release of GLP-1

Resistance to lean mass gain in constitutional thinness in free‐living conditions is not overpassed by overfeeding

International audienceBackgroundConstitutional thinness (CT), a non‐malnourished underweight state with no eating disorders, is characterized by weight gain resistance to high fat diet. Data issued from muscle biopsies suggested blunted anabolic mechanisms in free‐living state. Weight and metabolic responses to protein caloric supplementation has not been yet explored in CT.MethodsA 2 week overfeeding (additional 600 kcal, 30 g protein, 72 g carbohydrate, and 21 g fat) was performed to compare two groups of CTs (12 women and 11 men) to normal‐weight controls (12 women and 10 men). Bodyweight, food intake, energy expenditure, body composition, nitrogen balance, appetite hormones profiles, and urine metabolome were monitored before and after overfeeding.ResultsBefore overfeeding, positive energy gap was found in both CT genders (309 ± 370 kcal in CT‐F and 332 ± 709 kcal in CT‐M) associated with higher relative protein intake per kilo (1.74 ± 0.32 g/kg/day in CT‐F vs. 1.16 ± 0.23 in C‐F, P < 0.0001; 1.56 ± 0.36 in CT‐M vs. 1.22 ± 0.32 in C‐M, P = 0.03), lower nitrogen (7.26 ± 2.36 g/day in CT‐F vs. 11.41 ± 3.64 in C‐F, P = 0.003; 9.70 ± 3.85 in CT‐M vs. 14.14 ± 4.19 in C‐M, P = 0.02), but higher essential amino acids urinary excretion (CT/C fold change of 1.13 for leucine and 1.14 for arginine) in free‐living conditions. After overfeeding, CTs presented an accentuated positive energy gap, still higher than in controls (675 ± 540 in CTs vs. 379 ± 427 in C, P = 0.04). Increase in lean mass was induced in both controls genders but not in CTs (a trend was noticed in CT women), despite a similar nitrogen balance after overfeeding (5.06 ± 4.33 g/day in CTs vs. 4.28 ± 3.15 in controls, P = 0.49). Higher anorectic gut hormones' tone, glucagon‐like peptide 1 and peptide tyrosine tyrosine, during test meal and higher snacking frequency were noticed before and after overfeeding in CTs.ConclusionsThe blunted muscle energy mechanism, previously described in CTs in free‐living state, is associated with basal saturated protein turn over suggested by the concordance of positive nitrogen balance and an increased urine excretion of several essential amino acids. This saturation cannot be overpassed by increasing this spontaneous high‐protein intake suggesting a resistance to lean mass gain in CT phenotype

Elsevier Editorial System(tm) for Journal of Surgical Research Manuscript Draft Manuscript Number: Title: Learning curve of vessel cannulation in rats using CUSUM Analysis Extensive experience in rodent surgery and training junior surgeons

Extensive experience in rodent surgery and training junior surgeons Cover Letter Dear Ms/Mr, We would be grateful if you would consider our manuscript entitled &quot;Learning curve of cannulation of jugular and femoral vein in rats using Cumulative Summation Analysis&quot; for publication in The Journal of Surgical Research. Our study investigates the learning curve required for competency and proficiency at blood vessel cannulation in rodents. Vessel cannulation in rats, such as of the femoral and jugular vein, is a technically challenging procedure even for an experienced surgeon. In this study we analyse the results of 200 cannulations of the femoral and jugular vein in rats performed within one year by a single surgeon. The use of Cumulative Summation analysis allows calculating the required experience to most effectively cannulate the jugular and femoral vein, which is approximately 50 and 100 cases, respectively. These results highlight the difficulty involved in performing such micro-surgical operations in rodents. The knowledge of the average time taken to cannulate these vessels allows experiments to be optimally planned, avoiding unexpected time over-runs. Furthermore, our results bring into the spotlight the need for training and direct supervision for the inexperienced researchers, as there is an ongoing need to reduce animal use in research through minimising adverse events. To the authors&apos; knowledge this is the first study to report the learning curve for mastering the technique of catheterisation of the femoral and jugular vein in rats. We feel it will be of interest to readers of The Journal of Surgical Research, and particularly those using rodent microsurgery. Subject category: Research Authors&apos; contributions I.C., J.C., R.S. carried out the design and coordinated the study, participated in all of the experiments and prepared the manuscript. J.M., P.G., V.C., F.P., K.M., S.B. provided assistance in the design of the study, coordinated and carried out most of the experiments and participated in manuscript preparation. T.T., F.P., K.M. and S.B. approved the final version of the manuscript. All authors have read and approved the content of the manuscript. -3 - Abstract Background

The post-prandial secretion of Peptide YY1-36 and 3-36 in obesity is differentially increased after gastric bypass versus sleeve gastrectomy

OBJECTIVES: Peptide tyrosine tyrosine (PYY) exists as two species, PYY1-36 and PYY3-36, with distinct effects on insulin secretion and appetite regulation. The detailed effects of bariatric surgery on PYY1-36 and PYY3-36 secretion are not known as previous studies have used non-specific immunoassays to measure total PYY. Our objective was to characterise the effect of sleeve gastrectomy (SG) and Roux-en-Y gastric bypass (RYGB) on fasting and post-prandial PYY1-36 and PYY3-36 secretion using a newly developed liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay. DESIGN AND SUBJECTS: Observational study in 10 healthy non-obese volunteers and 30 participants with obesity who underwent RYGB (n=24) or SG (n=6) at the Imperial Weight Centre [NCT01945840]. Participants were studied using a standardised mixed meal test (MMT) before and 1 year after surgery. The outcome measure was PYY1-36 and PYY3-36 concentration. RESULTS: Pre-surgery, the fasting and post-prandial levels of PYY1-36 and PYY3-36 were low, with minimal responses to the MMT, and these did not differ from healthy non-obese volunteers. The postprandial secretion of both PYY1-36 and PYY3-36 at 1 year was amplified after RYGB, but not SG, with the response being significantly higher in RYGB compared to SG. CONCLUSIONS: There appears to be no difference in PYY secretion between non-obese and obese volunteers at baseline. At 1 year after surgery, RYGB, but not SG, is associated with increased post-prandial secretion of PYY1-36 and PYY3-36, which may account for long-term differences in efficacy and adverse effects between the two types of surgery