The normal β-cell response to obesity-associated insulin resistance is hypersecretion of insulin. Type 2 diabetes develops in subjects with β-cells that are susceptible to failure. Here, we investigated the time-dependent gene expression changes in islets of diabetes-prone db/db and diabetes-resistant ob/ob mice. The expressions of adaptive unfolded protein response (UPR) genes were progressively induced in islets of ob/ob mice, whereas they declined in diabetic db/db mice. Genes important for β-cell function and maintenance of the islet phenotype were reduced with time in db/db mice, whereas they were preserved in ob/ob mice. Inflammation and antioxidant genes displayed time-dependent upregulation in db/db islets but were unchanged in ob/ob islets. Treatment of db/db mouse islets with the chemical chaperone 4-phenylbutyric acid partially restored the changes in several β-cell function genes and transcription factors but did not affect inflammation or antioxidant gene expression. These data suggest that the maintenance (or suppression) of the adaptive UPR is associated with β-cell compensation (or failure) in obese mice. Inflammation, oxidative stress, and a progressive loss of β-cell differentiation accompany diabetes progression. The ability to maintain the adaptive UPR in islets may protect against the gene expression changes that underlie diabetes development in obese mice.
Abstract Insulin secretion from pancreatic β-cells is critical for maintaining glucose homeostasis and deregulation of circulating insulin levels is associated with the development of metabolic diseases. While many factors have been implicated in the stimulation of insulin secretion, the mechanisms that subsequently reduce insulin secretion remain largely unexplored. Here we demonstrate that mice with β-cell specific ablation of the Y1 receptor exhibit significantly upregulated serum insulin levels associated with increased body weight and adiposity. Interestingly, when challenged with a high fat diet these β-cell specific Y1-deficient mice also develop hyperglycaemia and impaired glucose tolerance. This is most likely due to enhanced hepatic lipid synthesis, resulting in an increase of lipid accumulation in the liver. Together, our study demonstrates that Y1 receptor signaling negatively regulates insulin release, and pharmacological inhibition of Y1 receptor signalling for the treatment of non-insulin dependent diabetes should be taken into careful consideration.
The skeleton has recently emerged as an additional player in the control of whole-body glucose metabolism; however, the mechanism behind this is not clear.Here we employ mice lacking neuropeptide Y, Y1 receptors solely in cells of the early osteoblastic lineage (Y1f3.6Cre), to examine the role of osteoblastic Y1 signalling in glycaemic control.Y1f3.6Cre mice not only have a high bone mass phenotype, but importantly also display altered glucose homeostasis; significantly decreased pancreas weight, islet number and pancreatic insulin content leading to elevated glucose levels and reduced glucose tolerance, but with no effect on insulin induced glucose clearance. The reduced glucose tolerance and elevated bone mass was corrected in Y1f3.6Cre mice by bone marrow transplant from wildtype animals, reinforcing the osteoblastic nature of this pathway. Importantly, when fed a high fat diet, Y1f3.6Cre mice, while equally gaining body weight and fat mass compared to controls, showed significantly improved glucose and insulin tolerance. Conditioned media from Y1f3.6Cre osteoblastic cultures was unable to stimulate insulin expression in MIN6 cells compared to conditioned media from wildtype osteoblast, indicating a direct signalling pathway. Importantly, osteocalcin a secreted osteoblastic factor previously identified as a modulator of insulin secretion was not altered in the Y1f3.6Cre model.This study identifies the existence of other osteoblast-derived regulators of pancreas function and insulin secretion and illustrates a mechanism by which NPY signalling in bone tissue is capable of regulating pancreatic function and glucose homeostasis.
Sodium-glucose cotransporter 2 (SGLT2) inhibitors (SGLT2i), or gliflozins, are anti-diabetic drugs that lower glycemia by promoting glucosuria, but they also stimulate endogenous glucose and ketone body production. The likely causes of these metabolic responses are increased blood glucagon levels, and decreased blood insulin levels, but the mechanisms involved are hotly debated. This study verified whether or not SGLT2i affect glucagon and insulin secretion by a direct action on islet cells in three species, using multiple approaches.We tested the in vivo effects of two selective SGLT2i (dapagliflozin, empagliflozin) and a SGLT1/2i (sotagliflozin) on various biological parameters (glucosuria, glycemia, glucagonemia, insulinemia) in mice. mRNA expression of SGLT2 and other glucose transporters was assessed in rat, mouse, and human FACS-purified α- and β-cells, and by analysis of two human islet cell transcriptomic datasets. Immunodetection of SGLT2 in pancreatic tissues was performed with a validated antibody. The effects of dapagliflozin, empagliflozin, and sotagliflozin on glucagon and insulin secretion were assessed using isolated rat, mouse and human islets and the in situ perfused mouse pancreas. Finally, we tested the long-term effect of SGLT2i on glucagon gene expression.SGLT2 inhibition in mice increased the plasma glucagon/insulin ratio in the fasted state, an effect correlated with a decline in glycemia. Gene expression analyses and immunodetections showed no SGLT2 mRNA or protein expression in rodent and human islet cells, but moderate SGLT1 mRNA expression in human α-cells. However, functional experiments on rat, mouse, and human (29 donors) islets and the in situ perfused mouse pancreas did not identify any direct effect of dapagliflozin, empagliflozin or sotagliflozin on glucagon and insulin secretion. SGLT2i did not affect glucagon gene expression in rat and human islets.The data indicate that the SGLT2i-induced increase of the plasma glucagon/insulin ratio in vivo does not result from a direct action of the gliflozins on islet cells.
Background and aims: Hypoxia is implicated in the loss of functional beta cell mass in type 2 diabetes and with islet transplantation, although the mechanisms are unknown. The adaptive unfolded protein response (UPR) is required for endoplasmic reticulum (ER) homeostasis and beta cell integrity. Here we investigated the influence of hypoxia on the adaptive UPR and the role it plays in apoptosis. Materials and methods: Isolated mouse islets and MIN6 cells were exposed to various O2 tensions. Ddit3 (Chop) and Hif1α were inhibited using siRNA. Hspa5 (Bip) was overexpressed using a plasmid vector. JNK was inhibited using SP600125. UPR and hypoxia-response gene expression was assessed in islets from prediabetic and diabetic db/db mice and age-matched lean control mice. mRNA and protein levels were measured by real-time RT-PCR and western blot. Apoptosis was measured by DNA fragmentation ELISA. Results: Deprivation of O2 (1% vs 20%) for 4-24h markedly reduced the mRNA and protein levels of adaptive UPR genes, including Hspa5, Hsp90b1 and Fkbp11 as well as the activation of Xbp1 (splicing) and PERK (phosphorylation). Opposing effects were observed in MEF cells suggesting that hypoxia specifically inhibits the adaptive UPR in beta cells. This was accompanied by upregulation of integrated stress response (ISR) genes, including Ddit3, Atf3 and Trb3 along with increased phosphorylated EIF2A. Interestingly, Ddit3 knockdown significantly increased adaptive UPR gene expression in association with partial protection against hypoxia-induced apoptosis (p<0.05). Moreover, Hspa5 overexpression alone partially protected against hypoxia-induced apoptosis (p<0.01). JNK inhibition, but not Hif1α knockdown, partially prevented the hypoxia-mediated loss of adaptive UPR gene expression and protected against hypoxia-induced apoptosis (p<0.01). Finally, mRNA levels of hypoxia-response genes, including Hyou1, Tpi1, Gapdh and Eno1, were markedly upregulated in vivo in the islets of diabetic db/db mice, but not in prediabetic db/db mice, suggesting that islet hypoxia correlates with beta cell failure. Interestingly, the upregulation of hypoxia-response genes further correlate with downregulation of adaptive UPR gene expression in diabetic db/db islets. Conclusion: Hypoxia inhibits the adaptive UPR in beta cells partially via Ddit3 and JNK activation, but independently of Hif1α. Downregulation of the adaptive UPR contributes to hypoxia-induced beta cell apoptosis and may play a role in the loss of functional beta cell mass in type 2 diabetes.
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