microRNA-375 regulates glucose metabolism-related signaling for insulin secretion.

Enhanced beta cell glycolytic and oxidative metabolism are necessary for glucose-induced insulin secretion. While several microRNAs modulate beta cell homeostasis, miR-375 stands out as it is highly expressed in beta cells where it regulates beta cell function, proliferation and differentiation. As glucose metabolism is central in all aspects of beta cell functioning, we investigated the role of miR-375 in this process using human and rat islets; the latter being an appropriate model for in-depth investigation. We used forced expression and repression of mR-375 in rat and human primary islet cells followed by analysis of insulin secretion and metabolism. Additionally, miR-375 expression and glucose-induced insulin secretion were compared in islets from rats at different developmental ages. We found that overexpressing of miR-375 in rat and human islet cells blunted insulin secretion in response to glucose but not to α-ketoisocaproate or KCl. Further, miR-375 reduced O2 consumption related to glycolysis and pyruvate metabolism, but not in response to α-ketoisocaproate. Concomitantly, lactate production was augmented suggesting that glucose-derived pyruvate is shifted away from mitochondria. Forced miR-375 expression in rat or human islets increased mRNA levels of pyruvate dehydrogenase kinase-4, but decreased those of pyruvate carboxylase and malate dehydrogenase1. Finally, reduced miR-375 expression was associated with maturation of fetal rat beta cells and acquisition of glucose-induced insulin secretion function. Altogether our findings identify miR-375 as an efficacious regulator of beta cell glucose metabolism and of insulin secretion, and could be determinant to functional beta cell developmental maturation.

Amino acid-induced regulation of hepatocyte growth: possible role of Drosha.

In an adult healthy liver, hepatocytes are in a quiescent stage unless a physical injury, such as ablation, or a toxic attack occur. Indeed, to maintain their crucial organismal homeostatic role, the damaged or remaining hepatocytes will start proliferating to restore their functional mass. One of the limiting conditions for cell proliferation is amino-acid availability, necessary both for the synthesis of proteins important for cell growth and division, and for the activation of the mTOR pathway, known for its considerable role in the regulation of cell proliferation. The overarching aim of our present work was to investigate the role of amino acids in the regulation of the switch between quiescence and growth of adult hepatocytes. To do so we used non-confluent primary adult rat hepatocytes as a model of partially ablated liver. We discovered that the absence of amino acids induces in primary rat hepatocytes the entrance in a quiescence state together with an increase in Drosha protein, which does not involve the mTOR pathway. Conversely, Drosha knockdown allows the hepatocytes, quiescent after amino-acid deprivation, to proliferate again. Further, hepatocyte proliferation appears to be independent of miRNAs, the canonical downstream partners of Drosha. Taken together, our observations reveal an intriguing non-canonical action of Drosha in the control of growth regulation of adult hepatocytes responding to a nutritional strain, and they may help to design novel preventive and/or therapeutic approaches for hepatic failure.

Argonaute-2 is associated to brown adipose tissue activation.

MicroRNAs (miRNAs) are important modulators of thermogenic brown adipose tissue (BAT). They have been involved in its differentiation and hence its functioning. While different regulators of the miRNA machinery have been shown to be essential for BAT differentiation, little is known about their implication in BAT activation. The aim of this work was to evaluate the role of AGO2, the chief miRNA mediator, in BAT activation. We took advantage of two non-genetic models of BAT activation to analyze the miRNA machinery and miRNA expression in BAT. We used principal component analysis (PCA) to obtain an overview of miRNA expression according to the BAT activation state. In vitro, we examined AGO2 expression during brown adipocyte differentiation and activation. Finally, we downregulated AGO2 to reveal its potential role in the thermogenic function of brown adipocytes. PCA analysis allowed to cluster animals on their miRNA signature in active BAT. Moreover, hierarchical clustering showed a positive correlation between global upregulation of miRNA expression and active BAT. Consistently, the miRNA machinery, particularly AGO2, was upregulated in vivo in active BAT and in vitro in mature brown adipocytes. Finally, the partial loss-of-function of AGO2 in mature brown adipocytes is sufficient to lead to a diminished expression of UCP1 associated to a decreased uncoupled respiration. Therefore, our study shows the potential contribution of AGO2 in BAT activation. Since BAT is a calorie-burning tissue these data have a translational potential in terms of therapeutic target in the field of altered fuel homeostasis associated to obesity and diabetes.

Contribution of the Long Noncoding RNA H19 to ß-Cell Mass Expansion in Neonatal and Adult Rodents.

Pancreatic ß-cell expansion throughout the neonatal period is essential to generate the appropriate mass of insulin-secreting cells required to maintain blood glucose homeostasis later in life. Hence, defects in this process can predispose to diabetes development during adulthood. Global profiling of transcripts in pancreatic islets of newborn and adult rats revealed that the transcription factor E2F1 controls expression of the long noncoding RNA H19, which is profoundly downregulated during the postnatal period. H19 silencing decreased ß-cell expansion in newborns, whereas its re-expression promoted proliferation of ß-cells in adults via a mechanism involving the microRNA let-7 and the activation of Akt. The offspring of rats fed a low-protein diet during gestation and lactation display a small ß-cell mass and an increased risk of developing diabetes during adulthood. We found that the islets of newborn rats born to dams fed a low-protein diet express lower levels of H19 than those born to dams that did not eat a low-protein diet. Moreover, we observed that H19 expression increases in islets of obese mice under conditions of increased insulin demand. Our data suggest that the long noncoding RNA H19 plays an important role in postnatal ß-cell mass expansion in rats and contributes to the mechanisms compensating for insulin resistance in obesity.

Age-Dependent Control of Energy Homeostasis by Brown Adipose Tissue in Progeny Subjected to Maternal Diet-Induced Fetal Programming.

Epidemiological and animal studies show that deleterious maternal environments predispose aging offspring to metabolic disorders and type 2 diabetes. Young progenies in a rat model of maternal low-protein (LP) diet are normoglycemic despite collapsed insulin secretion. However, without further worsening of the insulin secretion defect, glucose homeostasis deteriorates in aging LP descendants. Here we report that normoglycemic and insulinopenic 3-month-old LP progeny shows increased body temperature and energy dissipation in association with enhanced brown adipose tissue (BAT) activity. In addition, it is protected against a cold challenge and high-fat diet (HFD)-induced obesity with associated insulin resistance and hyperglycemia. Surgical BAT ablation in 3-month-old LP offspring normalizes body temperature and causes postprandial hyperglycemia. At 10 months, BAT activity declines in LP progeny with the appearance of reduced protection to HFD-induced obesity; at 18 months, LP progeny displays a BAT activity comparable to control offspring and insulin resistance and hyperglycemia occur. Together our findings identify BAT as a decisive physiological determinant of the onset of metabolic dysregulation in offspring predisposed to altered ß-cell function and hyperglycemia and place it as a critical regulator of fetal programming of adult metabolic disease.

A beta cell ATGL-lipolysis/adipose tissue axis controls energy homeostasis and body weight via insulin secretion in mice.

AIMS/HYPOTHESIS: To directly assess the role of beta cell lipolysis in insulin secretion and whole-body energy homeostasis, inducible beta cell-specific adipose triglyceride lipase (ATGL)-deficient (B-Atgl-KO) mice were studied under normal diet (ND) and high-fat diet (HFD) conditions. METHODS: Atgl flox/flox mice were cross-bred with Mip-Cre-ERT mice to generate Mip-Cre-ERT/+;Atgl flox/flox mice. At 8 weeks of age, these mice were injected with tamoxifen to induce deletion of beta cell-specific Atgl (also known as Pnpla2), and the mice were fed an ND or HFD. RESULTS: ND-fed male B-Atgl-KO mice showed decreased insulinaemia and glucose-induced insulin secretion (GSIS) in vivo. Changes in GSIS correlated with the islet content of long-chain saturated monoacylglycerol (MAG) species that have been proposed to be metabolic coupling factors for insulin secretion. Exogenous MAGs restored GSIS in B-Atgl-KO islets. B-Atgl-KO male mice fed an HFD showed reduced insulinaemia, glycaemia in the fasted and fed states and after glucose challenge, as well as enhanced insulin sensitivity. Moreover, decreased insulinaemia in B-Atgl-KO mice was associated with increased energy expenditure, and lipid metabolism in brown (BAT) and white (WAT) adipose tissues, leading to reduced fat mass and body weight. CONCLUSIONS/INTERPRETATION: ATGL in beta cells regulates insulin secretion via the production of signalling MAGs. Decreased insulinaemia due to lowered GSIS protects B-Atgl-KO mice from diet-induced obesity, improves insulin sensitivity, increases lipid mobilisation from WAT and causes BAT activation. The results support the concept that fuel excess can drive obesity and diabetes via hyperinsulinaemia, and that an islet beta cell ATGL-lipolysis/adipose tissue axis controls energy homeostasis and body weight via insulin secretion.

Visfatin expression analysis in association with recruitment and activation of human and rodent brown and brite adipocytes.

Human brown adipocytes are able to burn fat and glucose and are now considered as a potential strategy to treat obesity, type 2 diabetes and metabolic disorders. Besides their thermogenic function, brown adipocytes are able to secrete adipokines. One of these is visfatin, a nicotinamide phosphoribosyltransferase involved in nicotinamide dinucleotide synthesis, which is known to participate in the synthesis of insulin by pancreatic ß cells. In a therapeutic context, it is of interest to establish whether a potential correlation exists between brown adipocyte activation and/or brite adipocyte recruitment, and adipokine expression. We analyzed visfatin expression, as a pre-requisite to its secretion, in rodent and human biopsies and cell models of brown/brite adipocytes. We found that visfatin was preferentially expressed in mature adipocytes and that this expression was higher in brown adipose tissue of rodents compared to other fat depots. However, using various rodent models we were unable to find any correlation between visfatin expression and brown or brite adipocyte activation or recruitment. Interestingly, the situation is different in humans where visfatin expression was found to be equivalent between white and brown or brite adipocytes in vivo and in vitro. In conclusion, visfatin can be considered only as a rodent brown adipocyte biomarker, independently of tissue activation.

Circulating microRNAs and diabetes: potential applications in medical practice.

The explosive increase in the worldwide prevalence of diabetes over recent years has transformed the disease into a major public health concern. While diabetes can be screened for and diagnosed by reliable biological tests based on blood glucose levels, by and large there are no means of detecting at-risk patients or of following diabetic complications. The recent discovery that microRNAs are not only chief intracellular players in many biological processes, including insulin secretion and action, but are also circulating, has put them in the limelight as possible biological markers. Here we discuss the potential role of circulating microRNAs as biomarkers in the context of diabetes and its associated complications.

Maternal protein restriction leads to pancreatic failure in offspring: role of misexpressed microRNA-375.

The intrauterine environment of the fetus is a preeminent actor in long-term health. Indeed, mounting evidence shows that maternal malnutrition increases the risk of type 2 diabetes (T2D) in progeny. Although the consequences of a disturbed prenatal environment on the development of the pancreas are known, the underlying mechanisms are poorly defined. In rats, restriction of protein during gestation alters the development of the endocrine pancreas and favors the occurrence of T2D later in life. Here we evaluate the potential role of perturbed microRNA (miRNA) expression in the decreased ß-cell mass and insulin secretion characterizing progeny of pregnant dams fed a low-protein (LP) diet. miRNA profiling shows increased expression of several miRNAs, including miR-375, in the pancreas of fetuses of mothers fed an LP diet. The expression of miR-375 remains augmented in neoformed islets derived from fetuses and in islets from adult (3-month-old) progeny of mothers fed an LP diet. miR-375 regulates the proliferation and insulin secretion of dissociated islet cells, contributing to the reduced ß-cell mass and function of progeny of mothers fed an LP diet. Remarkably, miR-375 normalization in LP-derived islet cells restores ß-cell proliferation and insulin secretion. Our findings suggest the existence of a developmental memory in islets that registers intrauterine protein restriction. Hence, pancreatic failure after in utero malnutrition could result from transgenerational transmission of miRNA misexpression in ß-cells.

[microRNA and diabetes: tiny things causing huge effects]. / MicroARN et diabète - Petites structures - grands effets.

Soon after their discovery microRNA (miRNA) emerged as central natural regulators of gene expression. Although the complex mechanisms of action and impact of miRNA on development, physiology and disease are still elusive, significant progress has been made in deciphering the roles of some miRNA in insulin secretion and action. Here we examine the close relationship existing between miRNA and glucose metabolism as well as their putative role in the pathogenesis of diabetes and their possible utility as biomarkers of this disease.

MicroRNAs and metabolism crosstalk in energy homeostasis.

In record time, microRNAs (miRNAs) have acquired the respected stature of important natural regulators of global gene expression. Multiple studies have demonstrated that a large number of miRNAs are under the control of various metabolic stimuli, including nutrients, hormones, and cytokines. Conversely, it is now well recognized that miRNAs control metabolism, thereby generating a bidirectional functional link, which perturbs energy homeostasis in case of disconnection in the miRNA-metabolism interplay. A challenging road lies ahead for defining the role of miRNAs in the pathogenesis of diseases such as diabetes and for establishing their usefulness as new medications and clinically reliable biomarkers.

Impairment of rat fetal beta-cell development by maternal exposure to dexamethasone during different time-windows.

AIM: Glucocorticoids (GCs) take part in the direct control of cell lineage during the late phase of pancreas development when endocrine and exocrine cell differentiation occurs. However, other tissues such as the vasculature exert a critical role before that phase. This study aims to investigate the consequences of overexposure to exogenous glucocorticoids during different time-windows of gestation for the development of the fetal endocrine pancreas. METHODS: Pregnant Wistar rats received dexamethasone acetate in their drinking water (1 µg/ml) during the last week or throughout gestation. Fetuses and their pancreases were analyzed at day 15 and 21 of gestation. Morphometrical analysis was performed on pancreatic sections after immunohistochemistry techniques and insulin secretion was evaluated on fetal islets collected in vitro. RESULTS: Dexamethasone given the last week or throughout gestation reduced the beta-cell mass in 21-day-old fetuses by respectively 18% or 62%. This was accompanied by a defect in insulin secretion. The alpha-cell mass was reduced similarly. Neither islet vascularization nor beta-cell proliferation was affected when dexamethasone was administered during the last week, which was however the case when given throughout gestation. When given from the beginning of gestation, dexamethasone reduced the number of cells expressing the early marker of endocrine lineage neurogenin-3 when analyzed at 15 days of fetal age. CONCLUSIONS: GCs reduce the beta- and alpha-cell mass by different mechanisms according to the stage of development during which the treatment was applied. In fetuses exposed to glucocorticoids the last week of gestation only, beta-cell mass is reduced due to impairment of beta-cell commitment, whereas in fetuses exposed throughout gestation, islet vascularization and lower beta-cell proliferation are involved as well, amplifying the reduction of the endocrine mass.

Maternal malnutrition programs the endocrine pancreas in progeny.

Type 2 diabetes arises when the endocrine pancreas fails to secrete sufficient insulin to cope with metabolic demands resulting from ß cell secretory dysfunction, decreased ß cell mass, or both. Epidemiologic studies have shown strong relations between poor fetal and early postnatal nutrition and susceptibility to diabetes later in life. Animal models have been established, and studies have shown that a reduction in the availability of nutrients during fetal development programs the endocrine pancreas and insulin-sensitive tissues. We investigated several modes of early malnutrition in rats. Regardless of the type of diet investigated, whether there was a deficit in calories or protein in food or even in the presence of a high-fat diet, malnourished pups were born with a defect in their ß cell population, with fewer ß cells that did not secrete enough insulin and that were more vulnerable to oxidative stress; such populations of ß cells will never completely recover. Despite the similar endpoint, the cellular and physiologic mechanisms that contribute to alterations in ß cell mass differ depending on the nature of the nutritional insult. Hormones that are operative during fetal life, such as insulin, insulin-like growth factors, and glucocorticoids; specific molecules, such as taurine; and islet vascularization have been implicated as possible factors in amplifying this defect. The molecular mechanisms responsible for intrauterine programming of ß cells are still elusive, but among them the programming of mitochondria may be a strong central candidate.