Liver-expressed antimicrobial peptide 2 is associated with improved pancreatic insulin secretion in adults with overweight and obesity

Aims: To examine association of liver-expressed antimicrobial peptide 2 (LEAP2), an endogenous ghrelin antagonist with anorexiant effects, to key cardiometabolic risk factors in people with overweight and obesity. Methods: In this cross-sectional study, we sought to identify associations between LEAP2 levels and cardiometabolic risk factors, including body composition (dual X-ray absorptiometry), insulin and glucose metabolism (oral and intravenous glucose tolerance tests and hyperinsulinaemic-euglycaemic clamps), plasma lipids and inflammation markers (ELISA and multiplex assays). Results: In 65 participants with overweight or obesity (63.1% male, mean age 31.3 ± 8.5 years), LEAP2 levels were associated with total body fat, but not with body mass index or waist-hip ratio in both univariable and age- and sex-adjusted models (P < 0.05). Higher LEAP2 level was also positively associated with higher insulin secretion in univariable (P = 0.047) and multivariable models adjusted for age, sex and body fat (P = 0.03), but not with fasting glucose levels (P ≥ 0.05). Higher LEAP2 levels were associated insulin resistance (P = 0.07) after adjustment for age and sex, but the association disappeared after an additional adjustment for body fat (P = 0.2). There was an inverse association between LEAP2 levels and nuclear factor kappa-B (NFκB) activity in the peripheral blood mononuclear cells in age-, sex- and body fat-adjusted models (P = 0.04). There were no associations with cardiovascular risk factors (lipids, blood pressure) or other inflammation markers. Conclusions: These results provide important insights into the association between LEAP2 and cardiometabolic health in a high-risk population of individuals with overweight and obesity. This is a first report of an association between LEAP2 and insulin secretion, insulin sensitivity and NFκB activity. LEAP2 may represent an important potential therapeutic target to promote insulin secretion in people with type 2 diabetes and obesity

Ghrelin-AMPK Signaling Mediates the Neuroprotective Effects of Calorie Restriction in Parkinson's Disease

Jacqeline Bayliss, Romana Stark, Moyra Lemus, Vanessa Santos, Aiysha Thompson, Daniel Rees, Sandra Galic, John Elsworth, Bruce Kemp, Jeffrey Davies, and Zane Andrew

Unacylated-Ghrelin Impairs Hippocampal Neurogenesis and Memory in Mice and Is Altered in Parkinson’s Dementia in Humans

Blood-borne factors regulate adult hippocampal neurogenesis and cognition in mammals. We report that elevating circulating unacylated-ghrelin (UAG), using both pharmacological and genetic methods, reduced hippocampal neurogenesis and plasticity in mice. Spatial memory impairments observed in ghrelin-O-acyl transferase-null (GOAT/) mice that lack acyl-ghrelin (AG) but have high levels of UAG were rescued by acyl-ghrelin. Acyl-ghrelin-mediated neurogenesis in vitro was dependent on non-cell-autonomous BDNF signaling that was inhibited by UAG. These findings suggest that post-translational acylation of ghrelin is important to neurogenesis and memory in mice. To determine relevance in humans, we analyzed circulating AG:UAG in Parkinson disease (PD) patients diagnosed with dementia (PDD), cognitively intact PD patients, and controls. Notably, plasma AG:UAG was only reduced in PDD. Hippocampal ghrelin-receptor expression remained unchanged; however, GOAT+ cell number was reduced in PDD. We identify UAG as a regulator of hippocampal-dependent plasticity and spatial memory and AG:UAG as a putative circulating diagnostic biomarker of dementia

Oral Administration of Skin Gelatin Isolated from Chum Salmon (Oncorhynchus keta) Enhances Wound Healing in Diabetic Rats

Care for diabetic wounds remains a significant clinical problem. The present study was aimed at investigating the effect of skin gelatin from Chum Salmon on defective wound repair in the skin of diabetic rats. Full-thickness excisional skin wounds were made in 48 rats, of which 32 were diabetes. The diabetic rats were orally treated daily for 14 days with skin gelatin from Chum Salmon (2 g/kg) or its vehicle. Sixteen non-diabetic control rats received the same amount of water as vehicle-treated non-diabetic rats. Rats were killed to assess the rate of wound closure, microvessel density (MVD), vascular endothelial growth factor (VEGF), hydroxyproline (HP) contents in wound tissues and nitrate in plasma and wound tissue at 7 and 14 days after wounding. Skin gelatin-treated diabetic rats showed a better wound closure, increased MVD, VEGF, hyproxyproline and NO contents and a reduced extent of inflammatory response. All parameters were significant (P < 0.05) in comparison to vehicle-treated diabetic group. In light of our finding that skin gelatin of Chum Salmon promotes skin wound repair in diabetic rats, we propose that oral administration of Chum Salmon skin gelatin might be a beneficial method for treating wound disorders associated with diabetes

Prevention of Hepatic Steatosis and Hepatic Insulin Resistance by Knockdown of cAMP Response Element-Binding Protein

SummaryIn patients with poorly controlled type 2 diabetes mellitus (T2DM), hepatic insulin resistance and increased gluconeogenesis contribute to fasting and postprandial hyperglycemia. Since cAMP response element-binding protein (CREB) is a key regulator of gluconeogenic gene expression, we hypothesized that decreasing hepatic CREB expression would reduce fasting hyperglycemia in rodent models of T2DM. In order to test this hypothesis, we used a CREB-specific antisense oligonucleotide (ASO) to knock down CREB expression in liver. CREB ASO treatment dramatically reduced fasting plasma glucose concentrations in ZDF rats, ob/ob mice, and an STZ-treated, high-fat-fed rat model of T2DM. Surprisingly, CREB ASO treatment also decreased plasma cholesterol and triglyceride concentrations, as well as hepatic triglyceride content, due to decreases in hepatic lipogenesis. These results suggest that CREB is an attractive therapeutic target for correcting both hepatic insulin resistance and dyslipidemia associated with nonalcoholic fatty liver disease (NAFLD) and T2DM

Metabolic sensing in AgRP neurons integrates homeostatic state with dopamine signalling in the striatum

Agouti-related peptide (AgRP) neurons increase motivation for food, however, whether metabolic sensing of homeostatic state in AgRP neurons potentiates motivation by interacting with dopamine reward systems is unexplored. As a model of impaired metabolic-sensing, we used the AgRP-specific deletion of carnitine acetyltransferase (Crat) in mice. We hypothesised that metabolic sensing in AgRP neurons is required to increase motivation for food reward by modulating accumbal or striatal dopamine release. Studies confirmed that Crat deletion in AgRP neurons (KO) impaired ex vivo glucose-sensing, as well as in vivo responses to peripheral glucose injection or repeated palatable food presentation and consumption. Impaired metabolic-sensing in AgPP neurons reduced acute dopamine release (seconds) to palatable food consumption and during operant responding, as assessed by GRAB-DA photometry in the nucleus accumbens, but not the dorsal striatum. Impaired metabolic-sensing in AgRP neurons suppressed radiolabelled 18F-fDOPA accumulation after ~30 min in the dorsal striatum but not the nucleus accumbens. Impaired metabolic sensing in AgRP neurons suppressed motivated operant responding for sucrose rewards during fasting. Thus, metabolic-sensing in AgRP neurons is required for the appropriate temporal integration and transmission of homeostatic hunger-sensing to dopamine signalling in the striatum

The role of PEPCK-M in glucose homeostasis

Hintergrund: Wenn der Blutglukosespiegel ansteigt, wird Insulin aus der Bauchspeicheldrüse ausgeschüttet und macht so Glukose für die Zellen verfügbar. Der Blutglukosespiegel hängt von drei Faktoren ab: Resorption aus Darm nach der Nahrungsaufnahme, Glukoneogenese (die Synthese von "neuen" Glukosemolekülen in Leber und Niere) und Glykogenolyse (Abbau von Stärke, Glykogen, zu Glukose in Leber und Muskel). Das GTP-abhängige Enzym, Phosphoenolpyruvat Carboxykinase (PEPCK), ermöglicht die Bildung von Phosphoenolpyruvat (PEP) und ist so das Schlüsselenzym für die Glukoneogenese and Glyceroneogenese (die Produktion von "neuen" Glycerolmolekülen, die für die Veresterung von freien Fettsäuren zu Neutralfett, Triglyceriden, benötigt werden). Weniger bekannt ist, dass zwei PEPCK Formen existieren, eine zytosolische (PEPCK-C) und eine mitochondriale (PEPCK-M) Form. Während die Eigenschaften und die Regulation von PEPCK-C bereits intensiv untersucht wurden, war die metabolische Funktion von PEPCK-M bisher unklar. Hypothese: Im Gegensatz zu PEPCK-C, deren Transkription durch Insulin stark gehemmt wird, postulieren wir, dass PEPCK-M im Gegensatz dazu immer exprimiert wird und durch Substratverfügbarkeit und mitochondrialem GTP (mtGTP) reguliert wird. MtGTP wird durch Succinyl-CoA Synthetase gebildet und dient als Sensor für die Aktivität des Krebszyklus. Dieser Energie-sensitive mitochondriale Stoffwechselweg ist an der Glukosehomöostase beteiligt. Methode: Zur Untersuchung der Rolle von PEPCK-M im Glukosestoffwechel wurde PEPCK-M mittels siRNA in INS-1 832/13 Zellen und primäre Hepatozyten sowie mittels Antisense Oligonukleotiden (ASO) in Sprague Dawley Ratten herunterreguliert. Die Insulinsekretion wurde in INS-1 Zellen und die Gluconeogenese in Hepatozyten gemessen. Der PEPCK-M Stoffwechselweg wurde mit Hilfe einer neuentwickelten Strategie analysiert, wobei Stoffwechselprodukte mit 13C markiert und in einem Massenspektrometer gemessen wurden. Mit ASO behandelte Ratten wurden unter Bedingungen eines euglycämischen-hyperinsulinämischen Clamp-Tests sowie nach 36 Stunden Fasten untersucht. Ergebnisse: Wir haben mRNA, Protein und Aktivität von PEPCK-M in INS-1 Zellen und in Langerhans'schen Inselzellen der Ratte und Maus bestätigt. PEPCK-C war in diesen Zellen nicht nachweisbar. Mit unserer neu entwickelten Methode, Stoffwechselprodukte mit 13C zu markieren, zeigten wir, dass PEPCK-M in INS-1 Zellen und Inselzellen substantiell zur PEP Produktion beiträgt. In INS-1 Zellen betrug dieser Anteil 30 % und in Inselzellen der Ratte 41 %. In Gegenwart von Glukose verdreifachte sich der Beitrag von PEPCK-M zur gesamten PEP Produktion. Abschwächung des PEPCK-M Proteins hemmte die Glukose-stimulierte Insulinsekretion, was die zentrale Rolle von PEPCK-M in der mitochondrialen Stoffwechsel-mediierten Insulinsekretion unterstreicht. Desweiteren zeigten wir, dass PEPCK-M in der Leber der Ratte vorkommt und dort wahrscheinlich einen vergleichbaren Stoffwechselweg, die Glukoneogenese, reguliert. Eine Reduktion der PEPCK-M mRNA um 80% resultierte in Hepatozyten in einer von Glukose, Glukagon, oder Insulin unabhängigen Abnahme der Glukoneogenese um 60 %. Zur Bestimmung der Funktion von PEPCK-M in vivo erhielten Ratten eine intraperitoneale Injektion von ASO, um so die PEPCK-M Transkription zu drosseln. So wurde PEPCK-M mRNA um ungefähr 80 % in der Leber, proximalem Tubulus der Niere und weißem Fettgewebe vermindert. In ASO-behandelte Ratten waren Blut-Glukose (159.012.80 vs. 148.573.85 mg/dl, P<0.05) und -Insulin (576 vs. 356 myU/ml, P<0.02) nach Nahrungsaufnahme, nicht aber nach 36 Stunden Fasten reduziert. Der euglykämisch-hyperinsulinämischer Clamp ergab in Ratten mit verminderter PEPCK-M mRNA höhere Insulinsensitivität (Glukoseinfusionsraten: 201 vs. 261 mg/kg/min, P=0.003). Dies konnte nicht durch Unterschiede in der endogenen Glukoseproduktion, Aufnahme in der Muskulatur oder Fettgewebe erklärt werden. Interessanterweise, waren das post-absorptives Leberglykogen um 82 %, Blut-Triglyceride um 42 % und das Gewicht des entfernten Fettpolsters um 25 % niedriger. All dieses lässt auf einen konstitutiven Defekt in der PEP Produktion schließen. Schlussfolgerung: PEPCK-M spielt eine bedeutende Rolle in der Glukosehomöostase. Substrate des Krebszyklus liefern Energie für mtGTP Synthese und weiters PEP in der mitochondrialen Matrix via PEPCK-M. Dieser mitochondriale Stoffwechselweg koppelt die Flussrate durch den Krebszyklus mit jener der Anaplerose und triggert Insulinsekretion der [beta]-Zellen der Bauchspeicheldrüse. Ein ähnlicher Mechanismus findet sich in der Leber. Substrate des Krebszyklus werden via PEPCK-M zur Produktion von PEP verwendet, das wiederum der Glukose-Neubildung (Glukoneogenese) und Speicherung als Glykogen dient, beziehungsweise um Glycerol-Neubildung (Glyceroneogenese) und Speicherung als Triglycerid.Background: The pancreatic [beta]-cell hormone insulin makes glucose available for its cellular uptake and is secreted in response to an increase in plasma glucose. Circulating plasma glucose is derived from three sources: intestinal absorption after food intake, gluconeogenesis (de novo glucose production in liver and kidney) and glycogenolysis (breakdown of glycogen to glucose in liver and muscle). The GTP-dependent enzyme phosphoenolpyruvate carboxykinase (PEPCK) is the key enzyme that provides phosphoenolpyruvate (PEP) for gluconeogenesis and glyceroneogenesis (de novo glycerol production to store free fatty acids in form of triglycerides). Less known is that there are two isoforms, a cytosolic (PEPCK-C) and a mitochondrial (PEPCK-M) form. Most research focused on the function and regulation of PEPCK-C, and the metabolic role of PEPCK-M has not been established before. Hypothesis: In contrast to PEPCK-C whose transcription is strongly inhibited by insulin, we hypothesized that PEPCK-M is constitutively expressed and instead regulated in response to fuel supply by mitochondrial GTP (mtGTP) that is made by succinyl-CoA synthetase (SCS-GTP) and is a sensor for TCA flux. This energy sensing mitochondrial pathway is involved in glucose homeostasis. Methods: To assess the role of PEPCK-M in glucose homeostasis we silenced PEPCK-M in insulin secreting INS-1 832/13 cells as well as primary rat hepatocytes using siRNA and in Sprague Dawley rats using antisense oligonucleotides (ASO). We measured insulin secretion (INS-1 cells), gluconeogenic rates (hepatocytes) and flux via PEPCK-M. Metabolic flux studies were performed using novel 13C-labeling strategies with analysis by mass spectroscopy. ASO treated rats were studied by euglycemic hyperinsulinemic clamps and a 36-hour fast. Results: We confirmed the presence of PEPCK-M message, protein and activity in INS-1 cells, rat and mouse islets, whereas PEPCK-C was absent. Novel 13C-labeling strategies in INS-1 cells and islets measured substantial contribution of PEPCK-M to the synthesis of PEP. As high as 30 % of PEP in INS-1 cells and 41 % of PEP in rat islets came from PEPCK-M. The contribution of PEPCK-M to overall PEP synthesis was more than tripled with glucose stimulation. Silencing the PEPCK-M gene inhibited glucose-stimulated insulin secretion underscoring its central role in mitochondrial metabolism-mediated insulin secretion. Moreover, we confirmed the existence of PEPCK-M in rodent liver with a similar signaling III pathway regulating hepatic gluconeogenesis. An 80 % reduction of PEPCK-M mRNA in hepatocytes reduced gluconeogenesis by 60 % regardless of the presence of glucose, glucagon, or insulin. To assess the role of PEPCK-M in vivo rats were intraperitoneally injected with ASO that silenced PEPCK-M gene transcription by 80 % in liver, renal proximal tubules and white adipose tissue. PEPCK-M silenced rats had reduced plasma glucose (159.012.80 vs. 148.573.85 mg/dl, P<0.05) and insulin (576 vs. 356 myU/ml, P<0.02) in the fed state but were equivalent following a 36-hour fast. Euglycemic- hyperinsulinemic clamps identified increased insulin sensitivity when PEPCK-M was silenced (GINF: 201 vs. 261 mg/kg/min, P=0.003) that could not be accounted for by differences in endogenous glucose production nor uptake in the muscle or adipose. Interestingly, post-absorptive hepatic glycogen was 82 % lower and plasma triglycerides 42 % lower along with a 25% reduction in fat pad mass suggesting a constitutive defect in PEP production. Conclusion: Our work presented here, support an important role for PEPCK-M in glucose homeostasis. ^Substrate entry into the TCA cycle provides energy for mtGTP synthesis and further PEP synthesis in the mitochondrial matrix by PEPCK-M. In pancreatic [beta]-cells this mitochondrial metabolic flux pathway couples TCA cycle flux with anaplerotic flux to trigger insulin secretion. Similar in liver adequate substrate entry into the TCA cycle drives PEP production by PEPCK-M, which is then used to generate glucose (gluconeogenesis) to be stored as glycogen or glycerol (glyceroneogenesis) to be used for triglyceride synthesis.by Romana StarkAbweichender Titel laut Übersetzung der Verfasserin/des VerfassersWien, Med. Univ., Diss., 2011OeBB(VLID)171399

Ghrelin signalling in AgRP neurons links metabolic state to the sensory regulation of AgRP neural activity

Objective: The sensory detection of food and food cues suppresses Agouti related peptide (AgRP) neuronal activity prior to consumption with greatest suppression occurring in response to highly caloric food or interoceptive energy need. However, the interoceptive mechanisms priming an appropriate AgRP neural response to external sensory information of food availability remain unexplored. Since hunger increases plasma ghrelin, we hypothesized that ghrelin receptor (GHSR) signalling on AgRP neurons is a key interoceptive mechanism integrating energy need with external sensory cues predicting caloric availability. Methods: We used in vivo photometry to measure the effects of ghrelin administration or fasting on AgRP neural activity with GCaMP6s and dopamine release in the nucleus accumbens with GRAB-DA in mice lacking ghrelin receptors in AgRP neurons. Results: The deletion of GHSR on AgRP neurons prevented ghrelin-induced food intake, motivation and AgRP activity. The presentation of food (peanut butter pellet) or a wooden dowel suppressed AgRP activity in fasted WT but not mice lacking GHSRs in AgRP neurons. Similarly, peanut butter and a wooden dowel increased dopamine release in the nucleus accumbens after ip ghrelin injection in WT but not mice lacking GHSRs in AgRP neurons. No difference in dopamine release was observed in fasted mice. Finally, ip ghrelin administration did not directly increase dopamine neural activity in the ventral tegmental area. Conclusions: Our results suggest that AgRP GHSRs integrate an interoceptive state of energy need with external sensory information to produce an optimal change in AgRP neural activity. Thus, ghrelin signalling on AgRP neurons is more than just a feedback signal to increase AgRP activity during hunger