Center for Basic Metabolic Research

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Sparre-Ulrich A.H.,Copenhagen University | Sparre-Ulrich A.H.,Center for Basic Metabolic Research | Gabe M.N.,Copenhagen University | Gasbjerg L.S.,Copenhagen University | And 10 more authors.
Biochemical Pharmacology | Year: 2017

Alternative processing of the precursor protein pro-GIP results in endogenously produced GIP(1–30)NH2, that by DPP-4 cleavage in vivo results in the metabolite GIP(3–30)NH2. We showed previously that GIP(3–30)NH2 is a high affinity antagonist of the human GIPR in vitro. Here we determine whether it is suitable for studies of GIP physiology in rats since effects of GIP agonists and antagonists are strictly species-dependent. Transiently transfected COS-7 cells were assessed for cAMP accumulation upon ligand stimulation or assayed in competition binding using human 125I-GIP(1–42) as radioligand. In isolated perfused rat pancreata, insulin, glucagon, and somatostatin-releasing properties were evaluated. Competition binding demonstrated that on the rat GIP receptor (GIPR), rat GIP(3–30)NH2 bound with high affinity (Ki of 17 nM), in contrast to human GIP(3–30)NH2 (Ki of 250 nM). In cAMP studies, rat GIP(3–30)NH2 inhibited GIP(1–42)-induced rat GIPR activation and schild-plot analysis showed competitive antagonism with a pA2 of 13 nM and a slope of 0.9 ± 0.09. Alone, rat GIP(3–30)NH2 displayed weak, low-potent partial agonistic properties (EC50 > 1 μM) with an efficacy of 9.4% at 0.32 μM compared to GIP(1–42). In perfused rat pancreata, rat GIP(3–30)NH2 efficiently antagonized rat GIP(1–42)-induced insulin, somatostatin, and glucagon secretion. In summary, rat GIP(3–30)NH2 is a high affinity competitive GIPR antagonist and effectively antagonizes GIP-mediated G protein-signaling as well as pancreatic hormone release, while human GIP(3–30)NH2, despite a difference of only one amino acid between the two (arginine in position 18 in rat GIP(3–30)NH2; histidine in human), is unsuitable in the rat system. This underlines the importance of species differences in the GIP system, and the limitations of testing human peptides in rodent systems. © 2017 The Author(s)


Sparre-Ulrich A.H.,Copenhagen University | Hansen L.S.,Copenhagen University | Svendsen B.,Center for Basic Metabolic Research | Christensen M.,Copenhagen University | And 4 more authors.
British Journal of Pharmacology | Year: 2016

Background and Purpose Specific, high potency receptor antagonists are valuable tools when evaluating animal and human physiology. Within the glucose-dependent, insulinotropic polypeptide (GIP) system, considerable attention has been given to the presumed GIP receptor antagonist, (Pro3)GIP, and its effect in murine studies. We conducted a pharmacological analysis of this ligand including interspecies differences between the rodent and human GIP system. Experimental Approach Transiently transfected COS-7 cells were assessed for cAMP accumulation upon ligand stimulation and assayed in competition binding using 125I-human GIP. Using isolated perfused pancreata both from wild type and GIP receptor-deficient rodents, insulin-releasing, glucagon-releasing and somatostatin-releasing properties in response to species-specific GIP and (Pro3)GIP analogues were evaluated. Key Results Human (Pro3)GIP is a full agonist at human GIP receptors with similar efficacy (Emax) for cAMP production as human GIP, while both rat and mouse(Pro3)GIP were partial agonists on their corresponding receptors. Rodent GIPs are more potent and efficacious at their receptors than human GIP. In perfused pancreata in the presence of 7 mM glucose, both rodent (Pro3)GIP analogues induced modest insulin, glucagon and somatostatin secretion, corresponding to the partial agonist activities observed in cAMP production. Conclusions and Implications When evaluating new compounds, it is important to consider interspecies differences both at the receptor and ligand level. Thus, in rodent models, human GIP is a comparatively weak partial agonist. Human (Pro3)GIP was not an antagonist at human GIP receptors, so there is still a need for a potent antagonist in order to elucidate the physiology of human GIP. © 2015 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.


Persson F.,Steno Diabetes Center | Rossing P.,University of Aarhus | Rossing P.,Center for Basic Metabolic Research | Parving H.-H.,University of Aarhus | Parving H.-H.,Copenhagen University
Journal of Hypertension | Year: 2013

OBJECTIVE: Urinary levels of renin-angiotensin-aldosterone system (RAAS) components may reflect renal RAAS activity and/or the renal efficacy of RAAS inhibition. Our aim was to determine whether urinary angiotensinogen and renin are circulating RAAS-independent markers during RAAS blockade. METHODS: Urinary and plasma levels of angiotensinogen, renin, and albumin were measured in 22 patients with type 2 diabetes, hypertension, and albuminuria, during 2-month treatment periods with placebo, aliskiren, irbesartan, or their combination in random order in a crossover study. RESULTS: Aliskiren and irbesartan both increased plasma renin 3-4-fold, and above 10-fold when combined. Irbesartan decreased plasma angiotensinogen by approximately 25%, and no changes in plasma angiotensinogen were observed during the combination. Urine contained aliskiren at micromolar levels, blocking urinary renin by above 90%. Both blockers reduced urinary angiotensinogen, significant for irbesartan only. Combination blockade reduced urinary angiotensinogen even further. Reductions in urinary angiotensinogen paralleled albuminuria changes, and the urine/plasma concentration ratio of angiotensinogen was identical to that of albumin under all conditions. In contrast, urinary renin did not follow albumin, and remained unaltered after all treatments. Yet, the urine/plasma concentration ratio of renin was more than 100-fold higher than that of angiotensinogen and albumin, and approximately 4-fold reduced by single RAAS blockade, and more than 10-fold by dual RAAS blockade. CONCLUSIONS: Aliskiren filters into urine and influences urinary renin measurements. The urine/plasma renin ratio, but not urinary renin alone, may reflect the renal efficacy of RAAS blockade. Urinary angiotensinogen is a marker of filtration barrier damage rather than intrarenal RAAS activity. © 2013 Wolters Kluwer Health / Lippincott Williams & Wilkins.


Ebdrup B.H.,Center for Neuropsychiatric Schizophrenia Research | Knop F.K.,Glostrup | Madsen A.,Copenhagen University | Mortensen H.B.,Copenhagen University | And 4 more authors.
Journal of Clinical Psychiatry | Year: 2014

Objective: Treatment with antipsychotic drugs is widely associated with metabolic side effects such as weight gain and disturbed glucose metabolism, but the pathophysiologic mechanisms are unclear. Method: Fifty nondiabetic (fasting plasma glucose ≤ 7.0 mmol/L), antipsychotic-treated male patients (ICD-10 diagnosis code F20, F21, F22, F25, F28, or F60; mean±SD age = 33.0±6.7 years; body mass index [BMI; kg/ m2] = 26.0±4.7; waist circumference = 95.9±13.3 cm; glycated hemoglobin A1c [HbA1c] = 5.7%±0.3%) and 93 age- and waist circumference-matched healthy male controls (age = 33±7.3 years; BMI = 26.1±3.9; waist circumference = 94.6±11.9 cm; HbA1c = 5.7%±0.3%) participated in this cross-sectional study. Blood was sampled in the fasting state and 90 minutes after ingestion of a standardized liquid meal (2,268 kJ). The primary outcomes were glucometabolic hormones and cardiovascular risk markers. Data were collected between March 2008 and February 2010. Results: Compared to healthy controls, patients were characterized by elevated fasting levels of proinsulin, C-peptide, and glucose-dependent insulinotropic polypeptide (GIP) (P < .05) and higher postprandial levels of insulin, proinsulin, C-peptide, and GIP (P ≤ .02). Also, patients exhibited elevated plasma levels of C-reactive protein and signs of dyslipidemia. Fasting plasma levels of insulin, glucagon, glucagon-like peptide-1 (GLP-1), ghrelin, leptin, adiponectin, tumor necrosis factor-α, plasminogen activator inhibitor-1, and interleukin-6 and postprandial levels of glucagon, GLP-1, ghrelin, leptin, and adiponectin did not differ between groups. Conclusions: Presenting with an insulin resistant-like pattern, including beta cell hypersecretion and elevated GIP levels, nondiabetic antipsychotic-treated patients display emerging signs of dysmetabolism and a compromised cardiovascular risk profile. The appetite-regulating hormones GLP-1 and ghrelin appear not to be influenced by antipsychotic treatment. Our findings provide new clinical insight into the pathophysiology associated with metabolic side effects of antipsychotic treatment and put emphasis on the importance of implementing metabolic screening into psychiatric practice. © Copyright 2014 Physicians Postgraduate Press, Inc.


Bak M.J.,Center for Basic Metabolic Research | Bak M.J.,Copenhagen University | Albrechtsen N.W.,Center for Basic Metabolic Research | Pedersen J.,Center for Basic Metabolic Research | And 7 more authors.
European Journal of Endocrinology | Year: 2014

Aim: To determine the specificity and sensitivity of assays carried out using commercially available kits for glucagon and/or oxyntomodulin measurements. Methods: Ten different assay kits used for the measurement of either glucagon or oxyntomodulin concentrations were obtained. Solutions of synthetic glucagon (proglucagon (PG) residues 33-61), oxyntomodulin (PG residues 33-69) and glicentin (PG residues 1-69) were prepared and peptide concentrations were verified by quantitative amino acid analysis and a processing-independent in-house RIA. Peptides were added to the matrix (assay buffer) supplied with the kits (concentration range: 1.25-300 pmol/l) and to human plasma and recoveries were determined. Assays yielding meaningful results were analysed for precision and sensitivity by repeated analysis and ability to discriminate low concentrations. Results and conclusion: Three assays were specific for glucagon (carried out using the Millipore (Billerica, MA, USA), Bio-Rad (Sundbyberg, Sweden), and ALPCO (Salem, NH, USA) and Yanaihara Institute (Shizuoka, Japan) kits), but none was specific for oxyntomodulin. The assay carried out using the Phoenix (Burlingame, CA, USA) glucagon kit measured the concentrations of all three peptides (total glucagon) equally. Sensitivity and precision were generally poor; the assay carried out using the Millipore RIA kit performed best with a sensitivity around 10 pmol/l. Assays carried out using the BlueGene (Shanghai, China), USCN LIFE (Wuhan, China) (oxyntomodulin andglucagon),MyBioSource (San Diego, CA, USA)and Phoenix oxyntomodulin kits yielded inconsistent results. © 2014 European Society of Endocrinology Printed in Great Britain.


PubMed | Center for Basic Metabolic Research and Copenhagen University
Type: Comparative Study | Journal: British journal of pharmacology | Year: 2015

Specific, high potency receptor antagonists are valuable tools when evaluating animal and human physiology. Within the glucose-dependent, insulinotropic polypeptide (GIP) system, considerable attention has been given to the presumed GIP receptor antagonist, (Pro3)GIP, and its effect in murine studies. We conducted a pharmacological analysis of this ligand including interspecies differences between the rodent and human GIP system.Transiently transfected COS-7 cells were assessed for cAMP accumulation upon ligand stimulation and assayed in competition binding using (125) I-human GIP. Using isolated perfused pancreata both from wild type and GIP receptor-deficient rodents, insulin-releasing, glucagon-releasing and somatostatin-releasing properties in response to species-specific GIP and (Pro3)GIP analogues were evaluated.Human (Pro3)GIP is a full agonist at human GIP receptors with similar efficacy (Emax ) for cAMP production as human GIP, while both rat and mouse(Pro3)GIP were partial agonists on their corresponding receptors. Rodent GIPs are more potent and efficacious at their receptors than human GIP. In perfused pancreata in the presence of 7mM glucose, both rodent (Pro3)GIP analogues induced modest insulin, glucagon and somatostatin secretion, corresponding to the partial agonist activities observed in cAMP production.When evaluating new compounds, it is important to consider interspecies differences both at the receptor and ligand level. Thus, in rodent models, human GIP is a comparatively weak partial agonist. Human (Pro3)GIP was not an antagonist at human GIP receptors, so there is still a need for a potent antagonist in order to elucidate the physiology of human GIP.

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