Modulating effects of RAMPs on signaling profiles of the glucagon receptor family

Receptor activity-modulating proteins (RAMPs) are accessory molecules that form complexes with specific G protein-coupled receptors (GPCRs) and modulate their functions. It is established that RAMP interacts with the glucagon receptor family of GPCRs but the underlying mechanism is poorly understood. In this study, we used a bioluminescence resonance energy transfer (BRET) approach to comprehensively investigate such interactions. In conjunction with cAMP accumulation, Gα q activation and β-arrestin1/2 recruitment assays, we not only verified the GPCR-RAMP pairs previously reported, but also identified new patterns of GPCR-RAMP interaction. While RAMP1 was able to modify the three signaling events elicited by both glucagon receptor (GCGR) and glucagon-like peptide-1 receptor (GLP-1R), and RAMP2 mainly affected β-arrestin1/2 recruitment by GCGR, GLP-1R and glucagon-like peptide-2 receptor, RAMP3 showed a widespread negative impact on all the family members except for growth hormone-releasing hormone receptor covering the three pathways. Our results suggest that RAMP modulates both G protein dependent and independent signal transduction among the glucagon receptor family members in a receptor-specific manner. Mapping such interactions provides new insights into the role of RAMP in ligand recognition and receptor activation.

Fighting type 2 diabetes: Formulation strategies for peptide-based therapeutics

Diabetes mellitus is a major health problem with increasing prevalence at a global level. The discovery of insulin in the early 1900s represented a major breakthrough in diabetes management, with further milestones being subsequently achieved with the identification of glucagon-like peptide-1 (GLP-1) and the introduction of GLP-1 receptor agonists (GLP-1 RAs) in clinical practice. Moreover, the subcutaneous delivery of biotherapeutics is a well-established route of administration generally preferred over the intravenous route due to better patient compliance and prolonged drug absorption. However, current subcutaneous formulations of GLP-1 RAs present pharmacokinetic problems that lead to adverse reactions and treatment discontinuation. In this review, we discuss the current challenges of subcutaneous administration of peptide-based therapeutics and provide an overview of the formulations available for the different routes of administration with improved bioavailability and reduced frequency of administration.

Frontiers in the Treatment of Diabetes / 医药导报

In recent years,the prevalence of diabetes is increasing year by year with the improvement of people's living standard.The traditional oral anti-hyperglycemic drugs as well as insulin injection therapy can not block the progress of the disease course.The deterioration of glycaemic control results in various acute and chronic complications which seriously affect patients' health and bring huge economic burden.The researchers have been seeking new therapeutic regimens to improve or even reverse diabetes process.This paper focuses on the following novel treatment options such as GLP-1 receptor agonist,DPP4 inhibitors, fixed-dose combination,SGLT2 inhibitors,amylin analogues,dopamine receptor agonist,bile acid sequestrant,bariatric surgery and pancreatic stem cell transplantation.

The effects of astragalus combined with metformin on insulin resistance and plasma amylin level in newly diagnosed type 2 diabetic patients / 国际中医中药杂志

Objective To understand the effect of astragalus on insulin resistance and plasma amylin levels in newly diagnosed type 2 diabetic patients.Methods 88 patients of newly diagnosed type 2 diabetes were randomly divided into three groups:Group A (lifestyle intervention group) contained 30 patients,Group B (metformin treatment group) also contained 30 Patients,and Group C (astragalus and metformin treatment group) contained 28 Patients.Patients in group A were intervened with the control of diet,blood pressure and lipids level; patients in group B were additionally treated with metformin on the basis of group A; patients in group C were additionally treated with metformin and astragalus on the basis of group A.The course for both treatments were 8 weeks.Various clinical and biochemical parameters were detected before and after the treatment in all three groups and enzyme-linked immunosorbent assay was adopted for the detection of plasma amylin.Results Fasting blood glucose levels were significantly decreased after treataent in the three groups (t=-2.696、-4.029、-3.995,P＜0.05) ; insulin resistance index reduced in group B and group C (t=-2.599、-3.813,P＜0.05),the difference between group C and group B was statistically significant (t=-2.334,P＜0.05) ;treatments of group B and group C could improve the beta cell function index (t=2.303、2.384,P＜0.05),and they also could increase the plasma amylin level (t=2.341、3.045,P＜0.05).Conclusion Astragalus and metformin can improve insulin resistance index and increase plasma amylin levels in newly diagnosed type 2 diabetic patients.

Newer drugs in the management of diabetes mellitus.

Modern life style with present days technological advances have made human life sedentary. This is causing increasing prevalence of obesity and physical inactivity amongst population. The number of cases of diabetes worldwide in the year 2000 among adults 20 years of age is estimated to be 171 million in recent reports and is said to rise to more than 300 million by 2025. The raised plasma glucose levels give rise to complications in the form of microvascular and macrovascular complications diminished quality of life with reduced life expectancy. The currently available drugs used in the management of type II DM are not completely satisfactory in regard of controlling blood glucose level, many of the times they are associated with undesirable side effects. Hence there is continuous ongoing work in development of newer drugs, which are safe, efficacious and potent as well as free of undesirable effects such as sustained hypoglycaemia. Fortunately there are newer drug, few of them approved while other still knocking the door from the classes of drug such as GLP-1Mimetic, DPP-4 Inhibitors and others. Here we have tried to cover them in brief.

Delayed Response of Amylin Levels after an Oral Glucose Challenge in Children with Prader-Willi Syndrome

PURPOSE: Amylin secretion is increased parallel to insulin in obese subjects. Despite their marked obesity, a state of relative hypoinsulinemia occurs in children with Prader-Willi syndrome (PWS). Based on the hypothesis that amylin levels may be relatively low in PWS children, contributing to their excessive appetite, we studied amylin levels after oral glucose loading in children with PWS and overweight controls. MATERIALS AND METHODS: Plasma levels of amylin, glucagon, insulin, and glucose were measured at 0, 30, 60, 90, and 120 min after a glucose challenge in children with PWS (n = 18) and overweight controls (n = 25); the relationships among the variables were investigated in these two groups. RESULTS: Amylin levels were significantly correlated with insulin during fasting and during the oral glucose tolerance test in both groups. Amylin levels between 0 and 60 min after glucose loading were statistically different between the two groups. They were lower in children with PWS than in the controls between 0 and 30 min after glucose loading. CONCLUSION: The relatively low levels of amylin, compared to those in overweight controls, during the early phase of glucose loading in patients with PWS, may contribute, in part, to the excessive appetite of PWS patients as compared to the overweight controls.

Selection and preliminary identification of human Fab fragement antibody against amylin from phage antibody library / 中国免疫学杂志

Objective:To obtain antibodies against amylin from a 'naive' human Fab fragment antibody phage diasplay library and to analyze the specificity of antigen binding activity.Methods:Panning and screening Fab antibody from the antibody library,the positive clones with well reactivity to amylin were selected after five times selection of 'adsorption-elution-enrichment'.Then the plasmid DNA which was extracted from the clones,was digested with Spe Ⅰ and Nhe Ⅰ to delete gⅢ (about 660 bp).The digested 47 000 bp DNA which was purified after separation of bands from agarose gel was ligated with T4-DNA ligase.The constructed expressing phagemids were transformed to the BL21(DE3)pLysS,soluble Fab was expressed in it by the induction of IPTG and its characteristics and specificity were determined by ELISA and Western blot.Results:Soluble Fab antibodies were expressed in E.coli.According with molecular weight of IgG Fab,protein band of about 47 kD was shown by SDS-PAGE.Western blot using the goat anti human IgG-HRP showed their binding activities.ELISA showed their specificity with amylin antigens and they did not react with bovine serum albumin.Conclusion:The high level expression and identification of the soluble human anti- amylin Fab fragment antibodies has been obtained successfully,which lays a solid foundation for further researching about the biological and pathological activities of amylin.

Novel Therapies for Type 2 Diabetes Mellitus / 대한소아내분비학회지

Type 2 diabetes mellitus (T2DM) is a progressive disorder caused by a combination of insulin resistance and betacell dysfunction. The role of new hormones and systems in maintaining blood glucose homeostasis has recently been recognized. This recognition has led to the development of several novel classes of medications, including the incretin mimetic agents (glucagon like polypeptide-1 analogs and dipeptidyl peptidase IV inhibitors), the amylin analog and the endocannabinoid-1 receptor blocker. This review looks at these new agents in terms of their mode of action, pharmacokinetics and use in clinical practice. The new agents offer treatment options in select adult patients now, however, the efficacy and the safety has to be evaluated thoroughly by long term studies before the application to pediatric patients.

The effects of amylin on the islet β-cells voltage-gated L-calcium channels in rats / 中华内科杂志

Objective To observe the effect of amylin on the islet β-cells voltage-gated L-calcium channels in rats. Method Patch clamp technique was employed in the observation of the features and changes of electric current of islet β-cells voltage-gated L-calcium channels before and after using amylin. Results In the glucose environment of 5. 5 mmoL/L, the electric current of rat islet β-cells voltage-gated L-calcium channels was activated at-40 mV and reached the peak at about +20 mV, with a peak value of about-120 pA and the insulin secretion level was(0. 76±0. 12) μg/L. Under the stimulation of glucose of 16. 7 mmol/L, the peak current voltage moved to the left and increased up to-140 pA and the level of insulin secretion measured (1.78±0. 13) μg/L. Hatch islet β-cells in amylin at the concentrations of 0. 5, 1.0, 5.0 and 10.0 μmol/L, respectively. It was observed that in the 0. 5 μmol/L and 1.0 μmol/L groups,there was no remarkable change in the peak potential activation voltage, current, and insulin secretion volume in comparison with the control group. However, in the environment of 5.5 mmol/L glucose, the increase of activation voltage of the 5.0 and 10.0 μmol/L groups was-30 mV, with the peak current reduced to approximately-80 pA and-60 pA and the insulin secretion decreased to (0. 49±0. 11) μg/L and (0. 36±0. 12) μg/L respectively. Under the concentration of 16. 7 mmol/L glucose, the activation voltage increased from-40 mV up to-30 mV and the peak current reduced to-80 pA and-40 pA. In the meantime, the insulin secretion decreased respectively to (1.20±0. 13) μg/L and (0. 89±0. 14) μg/L, which is of significance. Conclusion The secretion of insulin is synchronized with the opening of the islet β-cells voltage-gated L-calcium channels at the stimulation of glucose. The amylin inhibition of the insulin secretion is also synchronized with the opening of islet β-cells voltage-gated L-calcium channels and it's in a positive concentration-dependent manner.

Effect of Amylin on Secretion of Insulin and Glucagon in Rats / 上海第二医科大学学报

Objective To study the effect of amylin on the secretion of glucagon and insulin in isolated rat islets. Methods The models of isolated rat islets were established by collagenase digestion and dextran gradient centrifugation. The influence of various concentrations of amylin on both glucagon and insulin secretion was studied. Results Various concentrations of glucose increased the insulin secretion and decreased the glucagon secretion.Compared with the control, islets exposed to amylin (10~ -10 to 10~ -5 mol/L) for 1 hour showed decreasing glucagon secretion as the glucose increased (0, 5.6 and 11.2 mmol/L) (P

Effect of amylin on biomechanical properties of the femur in glucocorticoid-induced osteoporosis rats / 医学研究生学报

Objective:To investigate the effect of amylin on biomechanical property of the femur in glucocorticoid-induced osteoporosis rats. Methods:Four groups of female Wistar rats(3 months old) were treated for 12 weeks as follows: ⅠNormal Control;ⅡDXM;Ⅲ DXM+AMY;Ⅳ DXM+Vitamin D_3.By intramuscular injection of dexamethasone(DXM)1mg/kg twice a week during the first 8 weeks,the animal model of Glucocorticoid induced osteoporosis was established.After 12 weeks,biomechanical properties of the femur were measured. Results:After the treatment with AMY,Bone structural mechanical(diameter) and geometrical parameters(displacement,peak load) of the femur were significantly increased(vs.DXM group,P

Effects of amylin on bone minernal density and structure parameters of bone tissue in glucocorticoid-induced osteoporosis rats / 医学研究生学报

Objectives:To investigate the effects of amylin on bone minernal density and structure parameters of bone tissue in glucocorticoid induced osteoporosis rats. Methods:Four groups of female Wistar rats (3 months old) were treated for 12 weeks as follows: ⅠNormal Control, ⅡDXM, Ⅲ DXM+AMY;Ⅳ DXM+Vitamin Da 3. By intramuscular injection of dexamethasone(XM)1mg/kg twice a week during the first 8 weeks, the animal model of Glucocorticoid induced osteopoprosis was established. After 12 weeks, BMD of the lumbar vertebrae and the femural bone were measured by DEXA. The bone morphology of the lumbar vertebrae was also determined. Results:①After the treatment with AMY, bone mineral density (BMD) was significantly increased at the lumbar spine and the femural bone. ( vs. DXM group, P

Effect of islet amyloid polypeptide(Amylin) on glucose metabolism and secretion of insulin in the rats / 中国糖尿病杂志

The effect of continuous amylin infusion on glucose tolerance in rats and the effect of amylin on glucose-stimulated insulin secretion from isolated rat pancreatic islets were sudied. Three groups of animals formed the study of glucose tolerance: Group 1 received salined infusion; Group2 , amylin at 2. 6 nmol?kg-1/h;and group 3,amylin at 26 nmol**kg-1/h. The total infusion time was 60 min. The insulin level and serum glucose were not altered at lower dose of amylin. However glucose levels markedly rose at 2.7,12 min. after beginning IVGTT, without significant change of insulin levels in group 3. The response of insulin to 16. 7 mol glucose-stimulated isolated rat pancretic islets was singnificantly inhibiled by the addition of amylin 10 umol.

Influence of amylin on apoptosis of human pancreatic islet ?-cells and its molecular mechanism / 中国病理生理杂志

AIM: To investigate the molecular mechanism of amylin in inducing apoptosis of human pancreatic islet ?-cells. METHODS: Human pancreatic islet cells were isolated and cultured. The cells were treated with amylin or amylin and aminoguanidine (AG group) for 24 h, respectively. Apoptosis of pancreatic islet ?-cells was studied by in situ TUNEL method combined with double staining for insulin and ELISA. The levels of insulin, NO 2 -/NO 3 - and glutathione (GSH), p53 mRNA and bcl-2 mRNA were also detected. RESULTS: (1) The enrichment factor and the apoptosis rate of pancreatic islet ?-cells in amylin group were markedly higher than that in control group and AG group ( P

Association of serum proinsulin with insulin resistance,and serum proamylin with islet beta cell function / 第三军医大学学报

Objective To study the serum levels of proinsulin(PI),insulin(INS),proamylin(PA) and amylin(AMY) in abnormal glucose metabolism individuals and to explore the relationship between PI and insulin resistance(IR) and between PA and islet beta cell function.Methods Totally 79 subjects who received Oral glucose tolerance test(OGTT) and insulin release test in our department from March to May 2008 were divided into 4 groups according to the results of OGTT,that is,normal group,impaired glucose regulation(IGR) group,T2DM 1[fasting blood-glucose(FBG)15 mmol/L) group.All subjects underwent examination of anthropometry and serum bio-chemical indicators,HOMA-IR and HOMA-B were calculated.Results No significant difference was found in the serum levels of PI or AMY among the 4 groups,but the level of PA,PI/INS and PA/AMY among the 4 groups did have significant differences(P