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The study was partially presented as a poster in the 53rd Annual Meeting of the European Association for the Study of Diabetes, Lisbon 2017., The role of insulin-degrading enzyme (IDE), a metalloprotease with high affinity for insulin, in insulin clearance remains poorly understood. OBJECTIVE: This study aimed to clarify whether IDE is a major mediator of insulin clearance, and to define its role in the etiology of hepatic insulin resistance., [Methods] We generated mice with liver-specific deletion of Ide (L-IDE-KO) and assessed insulin clearance and action., [Results] L-IDE-KO mice exhibited higher (~20%) fasting and non-fasting plasma glucose levels, glucose intolerance and insulin resistance. This phenotype was associated with ~30% lower plasma membrane insulin receptor levels in liver, as well as ~55% reduction in insulin-stimulated phosphorylation of the insulin receptor, and its downstream signaling molecules, AKT1 and AKT2 (reduced by ~40%). In addition, FoxO1 was aberrantly distributed in cellular nuclei, in parallel with up-regulation of the gluconeogenic genes Pck1 and G6pc. Surprisingly, L-IDE-KO mice showed similar plasma insulin levels and hepatic insulin clearance as control mice, despite reduced phosphorylation of the carcinoembryonic antigen-related cell adhesion molecule 1, which upon its insulin-stimulated phosphorylation, promotes receptor-mediated insulin uptake to be degraded., [Conclusion] IDE is not a rate-limiting regulator of plasma insulin levels in vivo., This work was supported by grants from the Ministerio de Economía, Industria y Competitividad: SAF2014-58702-C2-1-R and SAF2016-77871-C2-1-R to ICC; SAF2014-58702-C2-2-R and SAF2016-77871-C2-2-R to GP; supported by the EFSD European Research Programme on New Targets for Type 2 Diabetes supported by an educational research grant from MSD to ICC and GP; the National Institutes of Health: R01-DK054254, R01-DK083850 and RO1-HL-112248 to SMN, and R01-GM115617 to MAL; and the American Diabetes Association: Career Development Award 7-11-CD-13 to MAL.

Insulin degrading enzyme (IDE) is a ubiquitously expressed zinc-metalloprotease that is one of the principal enzymes involved in the degradation and clearance of insulin, in addition to glucagon, amylin and the amyloid-β protein (Aβ). Several polymorphisms of the gen IDE are associated with risk for type 2 diabetes mellitus (T2DM) in humans, whereas hepatic insulin clearance is reduced in T2DM patients. Although strongly contributing to insulin clearance in vivo, the specific role of IDE in the liver -the primary site of plasma insulin clearance- remain unclear. The aim of our study is to decipher the role of IDE in hepatic insulin signaling and its impact on whole-body glucose metabolism and insulin clearance. To ablate IDE selectively in hepatocytes, mice homozygous for a floxed IDE allele were intercrossed with albumin-Cre mice (hereafter L-IDE-KO). Metabolic studies were performed in 1 and 3-months old mice to analyze plasma levels of fasting and non-fasting glucose, insulin, glucagon, triglycerides, cholesterol, amylin, Aβ-40 and their body weight. Likewise, intraperitoneal glucose, insulin and pyruvate tolerance test and hepatic plasma insulin clearance were assessed. To analyze hepatic insulin signaling, fasted mice were injected intraperitoneally with Insulin (0,75U/Kg) 10 minutes before were sacrificed. Afterwards, liver samples were examined by western blot for levels of total and phosphorilated insulin receptor (IR), protein kinase B (AKT1 and AKT2), glycogen synthase kinase 3 (GSK3), glycogen synthase (GS), forkhead box protein O1 (FoXO1) and IDE. L-IDE-KO male mice showed hyperglycaemia, glucose intolerance and insulin resistance. These changes were not associated with augmented food intake, body weight or plasma insulin levels. Surprisingly, hepatic plasma insulin clearance was similar between L-IDE-KO and WT mice. Hepatic insulin resistance in L-IDE-KO mice was associated with reduced plasma membrane IR levels (≈30%), as well as reduced phosphorylation levels (≈55%), which induced reduced phosphorylation levels of AKT1 and AKT2 by ≈45%. In summary, we have revealed a new role for IDE in the regulation of hepatic insulin signaling. We hypothesize that IDE regulates IR recycling to the plasma membrane. Finally, our data demonstrate that IDE is not essential for hepatic plasma insulin clearance., Departamento de Bioquímica y Biología Molecular y Fisiología, Doctorado en Investigación Biomédica

This article belongs to the Special Issue Marine Compounds and Inflammation II, 2017., Type 2 diabetes (T2DM) is a complex disease linked to pancreatic beta-cell failure and insulin resistance. Current antidiabetic treatment regimens for T2DM include insulin sensitizers and insulin secretagogues. We have previously demonstrated that leptolide, a member of the furanocembranolides family, promotes pancreatic beta-cell proliferation in mice. Considering the beneficial effects of leptolide in diabetic mice, in this study, we aimed to address the capability of leptolide to improve insulin resistance associated with the pathology of obesity. To this end, we tested the hypothesis that leptolide should protect against fatty acid-induced insulin resistance in hepatocytes. In a time-dependent manner, leptolide (0.1 µM) augmented insulin-stimulated phosphorylation of protein kinase B (PKB) by two-fold above vehicle-treated HepG2 cells. In addition, leptolide (0.1 µM) counteracted palmitate-induced insulin resistance by augmenting by four-fold insulin-stimulated phosphorylation of PKB in HepG2 cells. In vivo, acute intraperitoneal administration of leptolide (0.1 mg/kg and 1 mg/kg) improved glucose tolerance and insulin sensitivity in lean mice. Likewise, prolonged leptolide treatment (0.1 mg/kg) in diet-induced obese mice improved insulin sensitivity. These effects were paralleled with an ~50% increased of insulin-stimulated phosphorylation of PKB in liver and skeletal muscle and reduced circulating pro-inflammatory cytokines in obese mice. We concluded that leptolide significantly improves insulin sensitivity in vitro and in obese mice, suggesting that leptolide may be another potential treatment for T2DM, This research has been funded by Sociedad Española de Diabetes (Ayudas Investigación Básica 2014), Salud Castilla y León (BIO/VA40/15) and Ministerio de Economía y Competitividad-Spain (SAF2014-58702-C2-1-R) to I.C. and Ministerio de Economía y Competitividad-Spain (SAF2014-58702-C2-2-R) to G.P., We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI).

Activation of pancreatic β-cell proliferation has been proposed as an approach to replace reduced functional β-cell mass in diabetes. Quiescent fibroblasts exit from G0 (quiescence) to G1 through pRb phosphorylation mediated by cyclin C/cdk3 complexes. Overexpression of cyclin D1, D2, D3, or cyclin E induces pancreatic β-cell proliferation. We hypothesized that cyclin C overexpression would induce β-cell proliferation through G0 exit, thus being a potential therapeutic target to recover functional β-cell mass. We used isolated rat and human islets transduced with adenovirus expressing cyclin C. We measured multiple markers of proliferation: [3H]thymidine incorporation, BrdU incorporation and staining, and Ki67 staining. Furthermore, we detected β-cell death by TUNEL, β-cell differentiation by RT-PCR, and β-cell function by glucose-stimulated insulin secretion. Interestingly, we have found that cyclin C increases rat and human β-cell proliferation. This augmented proliferation did not induce β-cell death, dedifferentiation, or dysfunction in rat or human islets. Our results indicate that cyclin C is a potential target for inducing β-cell regeneration., This work was supported by grants from ISCIII-Subdirección General de Evaluación y Fomento de la Investigación, Spain (PS09/00671); Programa Ramón y Cajal (RYC-2011-08101) Ministerio de Economía y Competitividad, Spain; and Europe-FP7 Marie Curie grant (IRG-247835) to I. Cózar-Castellano, by Europe-FP7 Marie Curie grant (IRG-256369) to G. Perdomo, by National Institute of Diabetes and Digestive and Kidney Diseases Grants (RO1-DK-073716;, DK-084236) and Juvenile Diabetes Research Foundation (17-2013-416) to E. Bernal-Mizrachi, and by a Minority Postdoctoral Fellowship from the American Diabetes Association to M. Jiménez-Palomares.

Diabetes is a consequence of a decrease on functional β-cell mass. We have recently demonstrated that epoxypukalide (Epoxy) is a natural compound with beneficial effects on primary cultures of rat islets. In this study, we extend our previous investigations to test the hypothesis that Epoxy protects β-cells and improves glucose metabolism in STZ-induced diabetic mice. We used 3-months old male mice that were treated with Epoxy at 200 μg/kg body weight. Glucose intolerance was induced by multiple intraperitoneal low-doses of streptozotocin (STZ) on 5 consecutive days. Glucose homeostasis was evaluated measuring plasma insulin levels and glucose tolerance. Histomorphometry was used to quantify the number of pancreatic β-cells per islet. β-cell proliferation was assessed by BrdU incorporation, and apoptosis by TUNEL staining. Epoxy treatment significantly improved glucose tolerance and plasma insulin levels. These metabolic changes were associated with increased β-cell numbers, as a result of a two-fold increase in β-cell proliferation and a 50% decrease in β-cell death. Our results demonstrate that Epoxy improves whole-body glucose homeostasis by preventing pancreatic β-cell death due to STZ-induced toxicity in STZ-treated mice., This work was supported by grants from: Programa Ramón y Cajal (RYC-2011-08101) Ministerio de Economía y Competitividad (Spain); Sociedad Española de Diabetes Basic Research Grant (2014) to IC.