Recent evidence suggests that the classic gut peptide, Peptide YY (PYY), could play a fundamental role in endocrine pancreatic function. In the present study expression of PYY and its NPY receptors on mouse islets and immortalised rodent and human beta-cells was examined together with the effects of both major circulating forms of PYY, namely PYY(1-36) and PYY(3-36), on beta-cell function, murine islet adaptions to insulin deficiency/resistance, as well as direct effects on cultured beta-cell proliferation and apoptosis. In vivo administration of PYY(3-36), but not PYY(1-36), markedly (p < 0.05) decreased food intake in overnight fasted mice. Neither form of PYY affected glucose disposal or insulin secretion following an i.p. glucose challenge. However, in vitro, PYY(1-36) and PYY(3-36) inhibited (p < 0.05 to p < 0.001) glucose, alanine and GLP-1 stimulated insulin secretion from immortalised rodent and human beta-cells, as well as isolated mouse islets, by impeding alterations in membrane potential, [Ca(2+)]i and elevations of cAMP. Mice treated with multiple low dose streptozotocin presented with severe (p < 0.01) loss of beta-cell mass accompanied by notable increases (p < 0.001) in alpha and PP cell numbers. In contrast, hydrocortisone-induced insulin resistance increased islet number (p < 0.01) and beta-cell mass (p < 0.001). PYY expression was consistently observed in alpha-, PP- and delta-, but not beta-cells. Streptozotocin decreased islet PYY co-localisation with PP (p < 0.05) and somatostatin (p < 0.001), whilst hydrocortisone increased PYY co-localisation with glucagon (p < 0.05) in mice. More detailed in vitro investigations revealed that both forms of PYY augmented (p < 0.05 to p < 0.01) immortalised human and rodent beta-cell proliferation and protected against streptozotocin-induced cytotoxicity, to a similar or superior extent as the well characterised beta-cell proliferative and anti-apoptotic agent GLP-1. Taken together, these data highlight the significance and potential offered by modulation of pancreatic islet NPY receptor signalling pathways for preservation of beta-cell mass in diabetes.

Diabetes results from the inability of pancreatic islets to maintain blood glucose concentrations within a normal physiological range. Clinical features are usually not observed until islets begin to fail and irreversible damage has occurred. Diabetes is generally diagnosed based on elevated glucose, which does not distinguish between type 1 and 2 diabetes. Thus, new diagnostic approaches are needed to detect different modes of diabetes before manifestation of disease. During prediabetes (pre-DM), islets undergo stress and release micro (mi) RNAs. Here, we review studies that have measured and tracked miRNAs in the blood for those with recent-onset or longstanding type 1 diabetes, obesity, pre-diabetes, type 2 diabetes, and gestational diabetes. We summarize the findings on miRNA signatures with the potential to stage progression of different modes of diabetes. Advances in identifying selective biomarker signatures may aid in early detection and classification of diabetic conditions and treatments to prevent and reverse diabetes.

Role of GLP-1 and GIP in beta cell compensatory responses to beta cell attack and insulin resistance were examined in C57BL/6 mice lacking functional receptors for GLP-1 and GIP. Mice were treated with multiple low dose streptozotocin or hydrocortisone. Islet parameters were assessed by immunohistochemistry and hormone measurements were determined by specific enzyme linked immunoassays. Wild-type streptozotocin controls exhibited severe diabetes, irregularly shaped islets with lymphocytic infiltration, decreased Ki67/TUNEL ratio with decreased beta cell and increased alpha cell areas. GLP-1 and GIP were co-expressed with glucagon and numbers of alpha cells mainly expressing GLP-1 were increased. In contrast, hydrocortisone treatment and induction of insulin resistance increased islet numbers and area, with enhanced beta cell replication, elevated mass of beta and alpha cells, together with co-expression of GLP-1 and GIP with glucagon in islets. The metabolic responses to streptozotocin in GLP-1RKO and GIPRKO mice were broadly similar to C57BL/6 controls, although decreases in islet numbers and size were more severe. In contrast, both groups of mice lacking functional incretin receptors displayed substantially impaired islet adaptations to insulin resistance induced by hydrocortisone, including marked curtailment of expansion of islet area, beta cell mass and islet number. Our observations cannot be explained by simple changes in circulating incretin concentrations, suggesting that intra-islet GLP-1 and GIP make a significant contribution to islet adaptation, particularly expansion of beta cell mass and compensatory islet compensation to hydrocortisone and insulin resistance.

Islet adaptations to pregnancy were explored in C57BL6/J mice lacking functional receptors for glucagon-like peptide 1 (GLP-1) and gastric inhibitory polypeptide (GIP). Pregnant wild type mice and GIPRKO mice exhibited marked increases in islet and beta cell area, numbers of medium/large sized islets, with positive effects on Ki67/Tunel ratio favouring beta cell growth and enhanced pancreatic insulin content. Alpha cell area and glucagon content were unchanged but prohormone convertases PC2 and PC1/3 together with significant amounts of GLP-1 and GIP were detected in alpha cells. Knockout of GLP-1R abolished these islet adaptations and paradoxically decreased pancreatic insulin, GLP-1 and GIP. This was associated with abolition of normal pregnancy-induced increases in plasma GIP, L-cell numbers, and intestinal GIP and GLP-1 stores. These data indicate that GLP-1 but not GIP is a key mediator of beta cell mass expansion and related adaptations in pregnancy, triggered in part by generation of intra-islet GLP-1.

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Aims/hypothesis Pancreatic islets produce non-coding microRNAs (miRNAs) that regulate islet cell function and survival. Our earlier investigations revealed that human islets undergo significant damage due to various types of stresses following transplantation and release miRNAs. Here, we sought to identify and validate exosomal miRNAs (exo-miRNAs) produced by human islets under conditions of cellular stress, preceding loss of cell function and death. We also aimed to identify islet stress signalling pathways targeted by exo-miRNAs to elucidate potential regulatory roles in islet cell stress. Methods Human islets were subjected to proinflammatory cytokine and hypoxic cell stress and miRNA from exosomes was isolated for RNA sequencing and analysis. Stress-induced exo-miRNAs were evaluated for kinetics of expression and release by intact islets for up to 48 h exposure to cytokines and hypoxia. A subset of stress-induced exo-miRNAs were assessed for recovery and detection as biomarkers of islet cell stress in a diabetic nude mouse xenotransplant model and in patients undergoing total pancreatectomy with islet auto-transplantation (TPIAT). Genes and signalling pathways targeted by stress-induced exo-miRNAs were identified by Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and direct interactions of miRNAs with downstream signalling targets were validated in human islet cells using the miRNA Tests for Read Analysis and Prediction (MirTrap) system. Results Global exo-miRNA sequencing revealed that 879 miRNA species were released from human islets and 190 islet exo-miRNAs were differentially expressed in response to proinflammatory cytokines, hypoxia or both. Release of exo-miRNAs hsa-miR-29b-3p and hsa-miR-216a-5p was detected within 6 h of exposure to cytokines and hypoxia. The remaining subset of stressinduced exo-miRNAs, including hsa-miR-148a-3p and islet cell damage marker hsa-miR-375, showed delayed release at 24-48 h, correlating with apoptosis and cell death. Stress and damage exo-miRNAs were significantly elevated in the circulation in human-to-mouse xenotransplant models and in human transplant recipients. Elevated blood exo-miRNAs negatively correlated with post-transplant islet function based on comparisons of stress and damage exo-miRNA indices with Secretory Unit of Islet Transplant Objects (SUITO) indices. KEGG analysis and further validation of exo-miRNA targets by MirTrap analysis revealed significant enrichment of islet mRNAs involved in phosphoinositide 3-kinase/Akt and mitogen-activated protein kinase signalling pathways. Conclusions/interpretation The study identifies exo-miRNAs differentially expressed and released by islets in response to damage and stress. These exo-miRNAs could serve as potential biomarkers for assessing islet damage and predicting outcomes in islet transplantation. Notably, exo-miRNAs 29b-3p and 216a-5p could be detected in islets prior to damage-released miRNAs and indicators of cellular apoptosis and death. Thus, these stress-induced exo-miRNAs may have potential diag...

Modulation of neuropeptide receptors is important for pancreatic β-cell function. Here, islet distribution and effects of the neurotensin (NT) receptor modulators, xenin and NT, was examined. Xenin, but not NT, significantly improved glucose disposal and insulin secretion, in mice. However, both peptides stimulated insulin secretion from rodent β-cells at 5.6 mm glucose, with xenin having similar insulinotropic actions at 16.7 mm glucose. In contrast, NT inhibited glucose-induced insulin secretion. Similar observations were made in human 1.1B4 β-cells and isolated mouse islets. Interestingly, similar xenin levels were recorded in pancreatic and small intestinal tissue. Arginine and glucose stimulated xenin release from islets. Streptozotocin treatment decreased and hydrocortisone treatment increased β-cell mass in mice. Xenin co-localisation with glucagon was increased by streptozotocin, but unaltered in hydrocortisone mice. This corresponded to elevated plasma xenin levels in streptozotocin mice. In addition, co-localisation of xenin with insulin was increased by hydrocortisone, and decreased by streptozotocin. Further in vitro investigations revealed that xenin and NT protected β-cells against streptozotocin-induced cytotoxicity. Xenin augmented rodent and human β-cell proliferation, whereas NT displayed proliferative actions only in human β-cells. These data highlight the involvement of NT signalling pathways for the possible modulation of β-cell function.

Aims/hypothesis We designed a chemically modified, enzyme-resistant peptide with triple-acting properties based on human glucagon with amino acid substitutions aligned to strategic positions in the sequence of glucose-dependent insulinotropic polypeptide (GIP). Methods Y 1 -dA 2 -I 12 -N 17 -V 18 -I 27 -G 28,29 -glucagon (termed YAG-glucagon) was incubated with dipeptidylpeptidase IV (DPP-IV) to assess stability, BRIN-BD11 cells to evaluate insulin secretion, and receptor-transfected cells to examine cAMP production. Acute glucose-lowering and insulinotropic properties of YAG-glucagon were assessed in National Institutes of Health (NIH) Swiss mice, while longer-term actions on glucose homeostasis, insulin secretion, food intake and body weight were examined in high-fat-fed mice. Results YAG-glucagon was resistant to DPP-IV, increased in vitro insulin secretion (1.5-3-fold; p<0.001) and stimulated cAMP production in GIP receptor-, glucagon-like peptide-1 (GLP-1) receptor-and glucagon receptortransfected cells. Plasma glucose levels were significantly reduced (by 51%; p<0.01) and insulin concentrations increased (1.2-fold; p<0.01) after acute injection of YAGglucagon in NIH Swiss mice. Acute actions were countered by established GIP, GLP-1 and glucagon antagonists. In high-fat-fed mice, twice-daily administration of YAGglucagon for 14 days reduced plasma glucose (40% reduction; p<0.01) and increased plasma insulin concentrations (1.8-fold; p<0.05). Glycaemic responses were markedly improved (19-48% reduction; p<0.05) and insulin secretion enhanced (1.5-fold; p<0.05) after a glucose load, which were independent of changes in insulin sensitivity, food intake and body weight.