Novel roles of mTORC2 in regulation of insulin secretion by actin filament remodeling.

Mammalian target of rapamycin (mTOR) kinase is an essential hub where nutrients and growth factors converge to control cellular metabolism. mTOR interacts with different accessory proteins to form complexes 1 and 2 (mTORC), and each complex has different intracellular targets. Although mTORC1's role in ß-cells has been extensively studied, less is known about mTORC2's function in ß-cells. Here, we show that mice with constitutive and inducible ß-cell-specific deletion of RICTOR (ßRicKO and ißRicKO mice, respectively) are glucose intolerant due to impaired insulin secretion when glucose is injected intraperitoneally. Decreased insulin secretion in ßRicKO islets was caused by abnormal actin polymerization. Interestingly, when glucose was administered orally, no difference in glucose homeostasis and insulin secretion were observed, suggesting that incretins are counteracting the mTORC2 deficiency. Mechanistically, glucagon-like peptide-1 (GLP-1), but not gastric inhibitory polypeptide (GIP), rescued insulin secretion in vivo and in vitro by improving actin polymerization in ßRicKO islets. In conclusion, mTORC2 regulates glucose-stimulated insulin secretion by promoting actin filament remodeling.NEW & NOTEWORTHY The current studies uncover a novel mechanism linking mTORC2 signaling to glucose-stimulated insulin secretion by modulation of the actin filaments. This work also underscores the important role of GLP-1 in rescuing defects in insulin secretion by modulating actin polymerization and suggests that this effect is independent of mTORC2 signaling.

Glucose stimulates microRNA-199 expression in murine pancreatic ß-cells.

MicroRNA 199 (miR-199) negatively impacts pancreatic ß-cell function and its expression is highly increased in islets from diabetic mice as well as in plasma of diabetic patients. Here we investigated how miR-199 expression is regulated in ß-cells by assessing expression of miR-199 precursors (primiR-199a1, primiR-199a2, and primiR-199b) and mature miR-199 (miR-199-3p and miR-199-5p) and promoter transcriptional activity assays in mouse islets and mouse insulinoma cells (MIN6) under different stimuli. We found that mouse islets equally express miR-199-3p and miR-199-5p. However, the primiRNA expression levels differed; although primiR-199a1 expression was about 30% greater than that of primiR-199a2, primiR-199b is barely detected in islets. We observed a 2-fold increase in primiR-199a1 and primiR-199a2 mRNA levels in mouse islets cultured in 10 mm glucose compared with 5.5 mm glucose. Similar responses to glucose were observed in MIN6 cells. Exposure to 30 mm KCl to induce membrane depolarization and calcium influx increased expression of primiR-199a2 but not of primiR-199a1 in MIN6 cells, indicating that calcium influx was involved. Transcriptional activity studies in MIN6 cells also revealed that primiR-199a2 promoter activity was enhanced by glucose and reduced by 2-deoxy-D-glucose-induced starvation. KCl and the potassium channel blocker tolbutamide also stimulated primiR-199a2 promoter activity. Calcium channel blockade by nifedipine reduced primiR-199a2 promoter activity in MIN6 cells, and diazoxide-mediated calcium influx inhibition blunted glucose up-regulation of miR-199-3p in islets. In conclusion, we uncover that glucose acutely up-regulates miR-199 family expression in ß-cells. Glucose metabolism and calcium influx are involved in primiR-199a2 expression but not primiR-199a1 expression.

3,5-Diiodothyronine protects against cardiac ischaemia-reperfusion injury in male rats.

NEW FINDINGS: What is the central question of this study? 3,5-Diiodothyronine (3,5-T2) administration increases resting metabolic rate, prevents or treats liver steatosis in rodent models, and ameliorates insulin resistance: what are its effects on cardiac electrical and contractile properties and autonomic regulation? What is the main finding and its importance? Chronic 3,5-T2 administration has no adverse effects on cardiac function. Remarkably, 3,5-T2 improves the autonomous control of the rat heart and protects against ischaemia-reperfusion injury. ABSTRACT: The use of 3,5,3'-triiodothyronine (T3) and thyroxine (T4) to treat metabolic diseases has been hindered by potential adverse effects on liver, lipid metabolism and cardiac electrical properties. It is recognized that 3,5-diiodothyronine (3,5-T2) administration increases resting metabolic rate, prevents or treats liver steatosis in rodent models and ameliorates insulin resistance, suggesting 3,5-T2 as a potential therapeutic tool. However, a comprehensive assessment of cardiac electrical and contractile properties has not been made so far. Three-month-old Wistar rats were daily administered vehicle, 3,5-T2 or 3,5-T2+T4 and no signs of atrial or ventricular arrhythmia were detected in non-anaesthetized rats during 90 days. Cardiac function was preserved as heart rate, left ventricle diameter and shortening fraction in 3,5-T2-treated rats compared to vehicle and 3,5-T2+T4 groups. Power spectral analysis indicated an amelioration of the heart rate variability only in 3,5-T2-treated rats. An increased baroreflex sensitivity at rest was observed in both 3,5-T2-treated groups. Finally, 3,5-T2 Langendorff-perfused hearts presented a significant recovery of left ventricular function and remarkably smaller infarction area after ischaemia-reperfusion injury. In conclusion, chronic 3,5-T2 administration ameliorates tonic cardiac autonomic control and confers cardioprotection against ischaemia-reperfusion injury in healthy male rats.

Islet pericytes convert into profibrotic myofibroblasts in a mouse model of islet vascular fibrosis.

AIMS/HYPOTHESIS: Islet vascular fibrosis may play an important role in the progression of type 2 diabetes, but there are no mouse models allowing detailed mechanistic studies to understand how a dysfunctional islet microvasculature contributes to diabetes pathogenesis. Here we report that the transgenic AktTg mouse, unlike other mouse strains, shows an increased deposition of extracellular matrix (ECM) proteins in perivascular regions, allowing us to study the cellular mechanisms that lead to islet vascular fibrosis. METHODS: Using immunohistochemistry, we labelled the islet microvasculature and ECM in pancreas sections of AktTg mice and human donors and performed lineage tracing to follow the fate of islet pericytes. We compared islet microvascular responses in living pancreas slices from wild-type and AktTg mice. RESULTS: We found that vascular pericytes proliferate extensively, convert into profibrotic myofibroblasts and substantially contribute to vascular fibrosis in the AktTg mouse model. The increased deposition of collagen I, fibronectin and periostin within the islet is associated with diminished islet perfusion as well as impaired capillary responses to noradrenaline (norepinephrine) and to high glucose in living pancreas slices. CONCLUSIONS/INTERPRETATION: Our study thus illustrates how the AktTg mouse serves to elucidate a cellular mechanism in the development of islet vascular fibrosis, namely a change in pericyte phenotype that leads to vascular dysfunction. Because beta cells in the AktTg mouse are more numerous and larger, and secrete more insulin, in future studies we will test the role beta cell secretory products play in determining the phenotype of pericytes and other cells residing in the islet microenvironment under physiological and pathophysiological conditions. Graphical abstract.

Sexual dimorphism in hypothalamic inflammation in the offspring of dams exposed to a diet rich in high fat and branched-chain amino acids.

Branched-chain amino acid (BCAAs: leucine, isoleucine, and valine) contribute to the development of obesity-associated insulin resistance in the context of consumption of a high-fat diet (HFD) in humans and rodents. Maternal diet is a major determinant of offspring health, and there is strong evidence that maternal HFD alters hypothalamic developmental programming and disrupts offspring energy homeostasis in rodents. In this study, we exposed pregnant and lactating C57BL/6JB female mice to either HFD, HFD with supplemented BCAA (HFD+BCAA), or standard diet (SC), and we studied offspring metabolic phenotypes. Both maternal HFD and HFD supplemented with BCAA had similar effect rendering the offspring metabolic imbalance and impairing their ability to cope with HFD when challenged during aging. The metabolic effects of HFD challenge were more profound in females, worsening female offspring ability to cope with an HFD challenge by activating hypothalamic inflammation in aging. Moreover, the sex differences in hypothalamic estrogen receptor α (ER-α) expression levels were lost in female offspring upon HFD challenge, supporting a link between ER-α levels and hypothalamic inflammation in offspring and highlighting the programming potential of hypothalamic inflammatory responses and maternal nutrition.

Coupling between Nutrient Availability and Thyroid Hormone Activation.

The activity of the thyroid gland is stimulated by food availability via leptin-induced thyrotropin-releasing hormone/thyroid-stimulating hormone expression. Here we show that food availability also stimulates thyroid hormone activation by accelerating the conversion of thyroxine to triiodothyronine via type 2 deiodinase in mouse skeletal muscle and in a cell model transitioning from 0.1 to 10% FBS. The underlying mechanism is transcriptional derepression of DIO2 through the mTORC2 pathway as defined in rictor knockdown cells. In cells kept in 0.1% FBS, there is DIO2 inhibition via FOXO1 binding to the DIO2 promoter. Repression of DIO2 by FOXO1 was confirmed using its specific inhibitor AS1842856 or adenoviral infection of constitutively active FOXO1. ChIP studies indicate that 4 h after 10% FBS-containing medium, FOXO1 binding markedly decreases, and the DIO2 promoter is activated. Studies in the insulin receptor FOXO1 KO mouse indicate that insulin is a key signaling molecule in this process. We conclude that FOXO1 represses DIO2 during fasting and that derepression occurs via nutritional activation of the PI3K-mTORC2-Akt pathway.

Role of the type 2 iodothyronine deiodinase (D2) in the control of thyroid hormone signaling.

BACKGROUND: Thyroid hormone signaling is critical for development, growth and metabolic control in vertebrates. Although serum concentration of thyroid hormone is remarkable stable, deiodinases modulate thyroid hormone signaling on a time- and cell-specific fashion by controlling the activation and inactivation of thyroid hormone. SCOPE OF THE REVIEW: This review covers the recent advances in D2 biology, a member of the iodothyronine deiodinase family, thioredoxin fold-containing selenoenzymes that modify thyroid hormone signaling in a time- and cell-specific manner. MAJOR CONCLUSIONS: D2-catalyzed T3 production increases thyroid hormone signaling whereas blocking D2 activity or disruption of the Dio2 gene leads to a state of localized hypothyroidism. D2 expression is regulated by different developmental, metabolic or environmental cues such as the hedgehog pathway, the adrenergic- and the TGR5-activated cAMP pathway, by xenobiotic molecules such as flavonols and by stress in the endoplasmic reticulum, which specifically reduces de novo synthesis of D2 via an eIF2a-mediated mechanism. Thus, D2 plays a central role in important physiological processes such as determining T3 content in developing tissues and in the adult brain, and promoting adaptive thermogenesis in brown adipose tissue. Notably, D2 is critical in the T4-mediated negative feed-back at the pituitary and hypothalamic levels, whereby T4 inhibits TSH and TRH expression, respectively. Notably, ubiquitination is a major step in the control of D2 activity, whereby T4 binding to and/or T4 catalysis triggers D2 inactivation by ubiquitination that is mediated by the E3 ubiquitin ligases WSB-1 and/or TEB4. Ubiquitinated D2 can be either targeted to proteasomal degradation or reactivated by deubiquitination, a process that is mediated by the deubiquitinases USP20/33 and is important in adaptive thermogenesis. GENERAL SIGNIFICANCE: Here we review the recent advances in the understanding of D2 biology focusing on the mechanisms that regulate its expression and their biological significance in metabolically relevant tissues. This article is part of a Special Issue entitled Thyroid hormone signalling.

Type 2 iodothyronine deiodinase is upregulated in rat slow- and fast-twitch skeletal muscle during cold exposure.

During cold acclimation, shivering is progressively replaced by nonshivering thermogenesis. Brown adipose tissue (BAT) and skeletal muscle are relevant for nonshivering thermogenesis, which depends largely on thyroid hormone. Since the skeletal muscle fibers progressively adapt to cold exposure through poorly defined mechanisms, our intent was to determine whether skeletal muscle type 2 deiodinase (D2) induction could be implicated in the long-term skeletal muscle cold acclimation. We demonstrate that in the red oxidative soleus muscle, D2 activity increased 2.3-fold after 3 days at 4°C together with the brown adipose tissue D2 activity, which increased 10-fold. Soleus muscle and BAT D2 activities returned to the control levels after 10 days of cold exposure, when an increase of 2.8-fold in D2 activity was detected in white glycolytic gastrocnemius but not in red oxidative gastrocnemius fibers. Propranolol did not prevent muscle D2 induction, but it impaired the decrease of D2 in BAT and soleus after 10 days at 4°C. Cold exposure is accompanied by increased oxygen consumption, UCP3, and PGC-1α genes expression in skeletal muscles, which were partialy prevented by propranolol in soleus and gastrocnemius. Serum total and free T3 is increased during cold exposure in rats, even after 10 days, when BAT D2 is already normalized, suggesting that skeletal muscle D2 activity contributes significantly to circulating T3 under this adaptive condition. In conclusion, cold exposure is accompanied by concerted changes in the metabolism of BAT and oxidative and glycolytic skeletal muscles that are paralleled by type 2 deiodinase activation.

Defining the In Vivo Role of mTORC1 in Thyrocytes by Studying the TSC2 Conditional Knockout Mouse Model.

Background: The thyroid gland is susceptible to abnormal epithelial cell growth, often resulting in thyroid dysfunction. The serine-threonine protein kinase mechanistic target of rapamycin (mTOR) regulates cellular metabolism, proliferation, and growth through two different protein complexes, mTORC1 and mTORC2. The PI3K-Akt-mTORC1 pathway's overactivity is well associated with heightened aggressiveness in thyroid cancer, but recent studies indicate the involvement of mTORC2 as well. Methods: To elucidate mTORC1's role in thyrocytes, we developed a novel mouse model with mTORC1 gain of function in thyrocytes by deleting tuberous sclerosis complex 2 (TSC2), an intracellular inhibitor of mTORC1. Results: The resulting TPO-TSC2KO mice exhibited a 70-80% reduction in TSC2 levels, leading to a sixfold increase in mTORC1 activity. Thyroid glands of both male and female TPO-TSC2KO mice displayed rapid enlargement and continued growth throughout life, with larger follicles and increased colloid and epithelium areas. We observed elevated thyrocyte proliferation as indicated by Ki67 staining and elevated cyclin D3 expression in the TPO-TSC2KO mice. mTORC1 activation resulted in a progressive downregulation of key genes involved in thyroid hormone biosynthesis, including thyroglobulin (Tg), thyroid peroxidase (Tpo), and sodium-iodide symporter (Nis), while Tff1, Pax8, and Mct8 mRNA levels remained unaffected. NIS protein expression was also diminished in TPO-TSC2KO mice. Treatment with the mTORC1 inhibitor rapamycin prevented thyroid mass expansion and restored the gene expression alterations in TPO-TSC2KO mice. Although total thyroxine (T4), total triiodothyronine (T3), and TSH plasma levels were normal at 2 months of age, a slight decrease in T4 and an increase in TSH levels were observed at 6 and 12 months of age while T3 remained similar in TPO-TSC2KO compared with littermate control mice. Conclusions: Our thyrocyte-specific mouse model reveals that mTORC1 activation inhibits thyroid hormone (TH) biosynthesis, suppresses thyrocyte gene expression, and promotes growth and proliferation.

Raptor levels are critical for ß-cell adaptation to a high-fat diet in male mice.

OBJECTIVE: The essential role of raptor/mTORC1 signaling in ß-cell survival and insulin processing has been recently demonstrated using raptor knock-out models. Our aim was to evaluate the role of mTORC1 function in adaptation of ß-cells to insulin resistant state. METHOD: Here, we use mice with heterozygous deletion of raptor in ß-cells (ßraHet) to assess whether reduced mTORC1 function is critical for ß-cell function in normal conditions or during ß-cell adaptation to high-fat diet (HFD). RESULTS: Deletion of a raptor allele in ß-cells showed no differences at the metabolic level, islets morphology, or ß-cell function in mice fed regular chow. Surprisingly, deletion of only one allele of raptor increases apoptosis without altering proliferation rate and is sufficient to impair insulin secretion when fed a HFD. This is accompanied by reduced levels of critical ß-cell genes like Ins1, MafA, Ucn3, Glut2, Glp1r, and specially PDX1 suggesting an improper ß-cell adaptation to HFD. CONCLUSION: This study identifies that raptor levels play a key role in maintaining PDX1 levels and ß-cell function during the adaptation of ß-cell to HFD. Finally, we identified that Raptor levels regulate PDX1 levels and ß-cell function during ß-cell adaptation to HFD by reduction of the mTORC1-mediated negative feedback and activation of the AKT/FOXA2/PDX1 axis. We suggest that Raptor levels are critical to maintaining PDX1 levels and ß-cell function in conditions of insulin resistance in male mice.

The Transcription Factor YY1 Is Essential for Normal DNA Repair and Cell Cycle in Human and Mouse ß-Cells.

Identifying the mechanisms behind the ß-cell adaptation to failure is important to develop strategies to manage type 2 diabetes (T2D). Using db/db mice at early stages of the disease process, we took advantage of unbiased RNA sequencing to identify genes/pathways regulated by insulin resistance in ß-cells. We demonstrate herein that islets from 4-week-old nonobese and nondiabetic leptin receptor-deficient db/db mice exhibited downregulation of several genes involved in cell cycle regulation and DNA repair. We identified the transcription factor Yin Yang 1 (YY1) as a common gene between both pathways. The expression of YY1 and its targeted genes was decreased in the db/db islets. We confirmed the reduction in YY1 expression in ß-cells from diabetic db/db mice, mice fed a high-fat diet (HFD), and individuals with T2D. Chromatin immunoprecipitation sequencing profiling in EndoC-ßH1 cells, a human pancreatic ß-cell line, indicated that YY1 binding regions regulate cell cycle control and DNA damage recognition and repair. We then generated mouse models with constitutive and inducible YY1 deficiency in ß-cells. YY1-deficient mice developed diabetes early in life due to ß-cell loss. ß-Cells from these mice exhibited higher DNA damage, cell cycle arrest, and cell death as well as decreased maturation markers. Tamoxifen-induced YY1 deficiency in mature ß-cells impaired ß-cell function and induced DNA damage. In summary, we identified YY1 as a critical factor for ß-cell DNA repair and cell cycle progression.

Physical exercise improves mitochondrial function in ovariectomized rats.

Estrogen deficiency causes metabolic disorders in humans and rodents, including in part due to changes in energy expenditure. We have shown previously that skeletal muscle mitochondrial function is compromised in ovariectomized (Ovx) rats. Since physical exercise is a powerful strategy to improve skeletal muscle mitochondrial content and function, we hypothesize that exercise training would counteract the deficiency-induced skeletal muscle mitochondrial dysfunction in Ovx rats. We report that exercised Ovx rats, at 60-65% of maximal exercise capacity for 8 weeks, exhibited less fat accumulation and body weight gain compared with sedentary controls. Treadmill exercise training decreased muscle lactate production, indicating a shift to mitochondrial oxidative metabolism. Furthermore, reduced soleus muscle mitochondrial oxygen consumption confirmed that estrogen deficiency is detrimental to mitochondrial function. However, exercise restored mitochondrial oxygen consumption in Ovx rats, achieving similar levels as in exercised control rats. Exercise-induced skeletal muscle peroxisome proliferator-activated receptor-γ coactivator-1α expression was similar in both groups. Therefore, the mechanisms by which exercise improves mitochondrial oxygen consumption appears to be different in Ovx-exercised and sham-exercised rats. While there was an increase in mitochondrial content in sham-exercised rats, demonstrated by a greater citrate synthase activity, no induction was observed in Ovx-exercised rats. Normalizing mitochondrial respiratory capacity by citrate synthase activity indicates a better oxidative phosphorylation efficiency in the Ovx-exercised group. In conclusion, physical exercise sustains mitochondrial function in ovarian hormone-deficient rats through a non-conventional mitochondrial content-independent manner.

Nutrient Sensor mTORC1 Regulates Insulin Secretion by Modulating ß-Cell Autophagy.

The dynamic regulation of autophagy in ß-cells by cycles of fasting-feeding and its effects on insulin secretion are unknown. In ß-cells, mechanistic target of rapamycin complex 1 (mTORC1) is inhibited while fasting and is rapidly stimulated during refeeding by a single amino acid, leucine, and glucose. Stimulation of mTORC1 by nutrients inhibited the autophagy initiator ULK1 and the transcription factor TFEB, thereby preventing autophagy when ß-cells were continuously exposed to nutrients. Inhibition of mTORC1 by Raptor knockout mimicked the effects of fasting and stimulated autophagy while inhibiting insulin secretion, whereas moderate inhibition of autophagy under these conditions rescued insulin secretion. These results show that mTORC1 regulates insulin secretion through modulation of autophagy under different nutritional situations. In the fasting state, autophagy is regulated in an mTORC1-dependent manner, and its stimulation is required to keep insulin levels low, thereby preventing hypoglycemia. Reciprocally, stimulation of mTORC1 by elevated leucine and glucose, which is common in obesity, may promote hyperinsulinemia by inhibiting autophagy.

The RNA-binding protein LARP1 is dispensable for pancreatic ß-cell function and mass.

Mechanistic target of rapamycin complex 1 (mTORC1) deficiency or chronic hyperactivation in pancreatic ß-cells leads to diabetes. mTORC1 complexes with La-related protein 1 (LARP1) to specifically regulate the expression of 5' terminal oligopyrimidine tract (5'TOP) mRNAs which encode proteins of the translation machinery and ribosome biogenesis. Here we show that LARP1 is the most expressed LARP in mouse islets and human ß-cells, being 2-4-fold more abundant than LARP1B, a member of the family that also interacts with mTORC1. Interestingly, ß-cells from diabetic patients have higher LARP1 and LARP1B expression. However, specific deletion of Larp1 gene in ß-cells (ß-Larp1KO mice) did not impair insulin secretion and glucose metabolism in male and female mice. High fat or high branched-chain amino acid (BCAA) diets did not disturb glucose homeostasis compared to control littermates up to 8 weeks; BCAA diet slightly impaired glucose tolerance in the ß-Larp1KO mice at 16 weeks. However, no differences in plasma insulin levels, non-fasting glycemia and ß-cell mass were observed in the ß-Larp1KO mice. In conclusion, LARP1 is the most abundant LARP in mouse islets and human ß-cells, and it is upregulated in diabetic subjects. However, genetically disruption of Larp1 gene did not impact glucose homeostasis in basal and diabetogenic conditions, suggesting no major role for LARP1 in ß-cells.

Exercise-Stimulated ROS Sensitive Signaling Pathways in Skeletal Muscle.

Physical exercise represents a major challenge to whole-body homeostasis, provoking acute and adaptative responses at the cellular and systemic levels. Different sources of reactive oxygen species (ROS) have been described in skeletal muscle (e.g., NADPH oxidases, xanthine oxidase, and mitochondria) and are closely related to the physiological changes induced by physical exercise through the modulation of several signaling pathways. Many signaling pathways that are regulated by exercise-induced ROS generation, such as adenosine monophosphate-activated protein kinase (AMPK), mitogen activated protein kinase (MAPK), nuclear respiratory factor2 (NRF2), and PGC-1α are involved in skeletal muscle responses to physical exercise, such as increased glucose uptake, mitochondriogenesis, and hypertrophy, among others. Most of these adaptations are blunted by antioxidants, revealing the crucial role played by ROS during and after physical exercise. When ROS generation is either insufficient or exacerbated, ROS-mediated signaling is disrupted, as well as physical exercise adaptations. Thus, an understanding the limit between "ROS that can promote beneficial effects" and "ROS that can promote harmful effects" is a challenging question in exercise biology. The identification of new mediators that cause reductive stress and thereby disrupt exercise-stimulated ROS signaling is a trending on this topic and are covered in this current review.

Pancreatic and duodenal homeobox-1 (PDX1) contributes to ß-cell mass expansion and proliferation induced by Akt/PKB pathway.

Maintenance of pancreatic ß-cell mass and function is fundamental to glucose homeostasis and to prevent diabetes. The PI3 K-Akt-mTORC1 pathway is critical for ß-cells mass and function, while PDX1 has been implicated in ß-cell development, maturation, and function. Here we tested whether Akt signaling requires PDX1 expression to regulate ß-cell mass, proliferation, and glucose homeostasis. In order to address that, we crossed a mouse model overexpressing constitutively active Akt mutant in ß-cells (ß-caAkt) with mice lacking one allele of PDX1gene (ß-caAkt/pdx1+/-). While the ß-caAkt mice exhibit higher plasma insulin levels, greater ß-cell mass and improved glucose tolerance compared to control mice, the ß-caAkt/pdx1+/- mice are hyperglycemic and intolerant to glucose. The changes in glucose homeostasis in ß-caAkt/pdx1+/- were associated with a 60% reduction in ß-cell mass compared to ß-caAkt mice. The impaired ß-cell mass in the ß-caAkt/pdx1+/- mice can be explained by a lesser ß-cell proliferation measured by the number of Ki67 positive ß-cells. We did not observe any differences in apoptosis between ß-caAkt/pdx1+/- and ß-caAkt mice. In conclusion, PDX1 contributes to ß-cell mass expansion and glucose metabolism induced by activation of Akt signaling.

Redox Signaling in Widespread Health Benefits of Exercise.

Significance: Exercise-induced reactive oxygen species (ROS) production activates multiple intracellular signaling pathways through genomic and nongenomic mechanisms that are responsible for the beneficial effects of exercise in muscle. Beyond the positive effect of exercise on skeletal muscle cells, other tissues such as white and brown adipose, liver, central nervous system, endothelial, heart, and endocrine organ tissues are also responsive to exercise. Recent Advances: Crosstalk between different cells is essential to achieve homeostasis and to promote the benefits of exercise through paracrine or endocrine signaling. This crosstalk can be mediated by different effectors that include the secretion of metabolites of muscle contraction, myokines, and exosomes. During the past 20 years, it has been demonstrated that contracting muscle cells produce and secrete different classes of myokines, which functionally link muscle with nearly all other cell types. Critical Issues: The redox signaling behind this exercise-induced crosstalk is now being decoded. Many of these widespread beneficial effects of exercise require not only a complex ROS-dependent intramuscular signaling cascade but simultaneously, an integrated network with many remote tissues. Future Directions: Strong evidence suggests that the powerful beneficial effect of regular physical activity for preventing (or treating) a large range of disorders might also rely on ROS-mediated signaling. Within a contracting muscle, ROS signaling may control exosomes and myokines secretion. In remote tissues, exercise generates regular and synchronized ROS waves, creating a transient pro-oxidative environment in many cells. These new concepts integrate exercise, ROS-mediated signaling, and the widespread health benefits of exercise.

Type 2 deiodinase polymorphism causes ER stress and hypothyroidism in the brain.

Levothyroxine (LT4) is a form of thyroid hormone used to treat hypothyroidism. In the brain, T4 is converted to the active form T3 by type 2 deiodinase (D2). Thus, it is intriguing that carriers of the Thr92Ala polymorphism in the D2 gene (DIO2) exhibit clinical improvement when liothyronine (LT3) is added to LT4 therapy. Here, we report that D2 is a cargo protein in ER Golgi intermediary compartment (ERGIC) vesicles, recycling between ER and Golgi. The Thr92-to-Ala substitution (Ala92-D2) caused ER stress and activated the unfolded protein response (UPR). Ala92-D2 accumulated in the trans-Golgi and generated less T3, which was restored by eliminating ER stress with the chemical chaperone 4-phenyl butyric acid (4-PBA). An Ala92-Dio2 polymorphism-carrying mouse exhibited UPR and hypothyroidism in distinct brain areas. The mouse refrained from physical activity, slept more, and required additional time to memorize objects. Enhancing T3 signaling in the brain with LT3 improved cognition, whereas restoring proteostasis with 4-PBA eliminated the Ala92-Dio2 phenotype. In contrast, primary hypothyroidism intensified the Ala92-Dio2 phenotype, with only partial response to LT4 therapy. Disruption of cellular proteostasis and reduced Ala92-D2 activity may explain the failure of LT4 therapy in carriers of Thr92Ala-DIO2.

A Single Dose of Beetroot Gel Rich in Nitrate Does Not Improve Performance but Lowers Blood Glucose in Physically Active Individuals.

Background. Beetroot consumption has been proposed to improve exercise performance, since the nitrate content of this food is able to stimulate the synthesis of nitric oxide. Objective. The acute effect of 100 g of a beetroot gel containing ~10 mmol of nitrate was tested on the nitric oxide synthesis, on metabolic and biochemical parameters, and on performance in physically active individuals. Methods. Through a double blind, crossover, placebo-controlled study, 25 healthy runners ingested a single dose of beetroot and placebo gels. Participants performed an aerobic exercise protocol on a treadmill (3 min warm-up of 40% peak oxygen consumption, 4 min at 90% of gas exchange threshold I and 70% (Δ) maximal end speed until volitional fatigue). Results. Urinary levels of nitrite and nitrate increased after 90 min of beetroot gel ingestion. Plasma glucose concentrations lowered after the exercise and the decrease was maintained for 20 min. Systolic and diastolic blood pressures, serum cortisol, and blood lactate were not altered after the beetroot gel ingestion compared to a placebo gel. Conclusion. The single dose of beetroot gel provoked an increase of nitric oxide synthesis although no improvement on the physical performance of athletes during aerobic submaximal exercise was observed.

Loss of mTORC1 signalling impairs ß-cell homeostasis and insulin processing.

Deregulation of mTOR complex 1 (mTORC1) signalling increases the risk for metabolic diseases, including type 2 diabetes. Here we show that ß-cell-specific loss of mTORC1 causes diabetes and ß-cell failure due to defects in proliferation, autophagy, apoptosis and insulin secretion by using mice with conditional (ßraKO) and inducible (MIP-ßraKOf/f) raptor deletion. Through genetic reconstitution of mTORC1 downstream targets, we identify mTORC1/S6K pathway as the mechanism by which mTORC1 regulates ß-cell apoptosis, size and autophagy, whereas mTORC1/4E-BP2-eIF4E pathway regulates ß-cell proliferation. Restoration of both pathways partially recovers ß-cell mass and hyperglycaemia. This study also demonstrates a central role of mTORC1 in controlling insulin processing by regulating cap-dependent translation of carboxypeptidase E in a 4EBP2/eIF4E-dependent manner. Rapamycin treatment decreases CPE expression and insulin secretion in mice and human islets. We suggest an important role of mTORC1 in ß-cells and identify downstream pathways driving ß-cell mass, function and insulin processing.