Exploring the Effects of Metabolism-Disrupting Chemicals on Pancreatic α-Cell Viability, Gene Expression and Function: A Screening Testing Approach.

Humans are constantly exposed to many environmental pollutants, some of which have been largely acknowledged as key factors in the development of metabolic disorders such as diabetes and obesity. These chemicals have been classified as endocrine-disrupting chemicals (EDCs) and, more recently, since they can interfere with metabolic functions, they have been renamed as metabolism-disrupting chemicals (MDCs). MDCs are present in many consumer products, including food packaging, personal care products, plastic bottles and containers, and detergents. The scientific literature has ever-increasingly focused on insulin-releasing pancreatic ß-cells as one of the main targets for MDCs. Evidence highlights that these substances may disrupt glucose homeostasis by altering pancreatic ß-cell physiology. However, their potential impact on glucagon-secreting pancreatic α-cells remains poorly known despite the essential role that this cellular type plays in controlling glucose metabolism. In the present study, we have selected seven paradigmatic MDCs representing major toxic classes, including bisphenols, phthalates, perfluorinated compounds, metals, and pesticides. By using an in vitro cell-based model, the pancreatic α-cell line αTC1-9, we have explored the effects of these compounds on pancreatic α-cell viability, gene expression, and secretion. We found that cell viability was moderately affected after bisphenol-A (BPA), bisphenol-F (BPF), and perfluorooctanesulfonic acid (PFOS) exposure, although cytotoxicity was relatively low. In addition, all bisphenols, as well as di(2-ethylhexyl) phthalate (DEHP) and cadmium chloride (CdCl2), promoted a marked decreased on glucagon secretion, together with changes in the expression of glucagon and/or transcription factors involved in cell function and identity, such as Foxo1 and Arx. Overall, our results indicated that most of the selected chemicals studied caused functional alterations in pancreatic α-cells. Moreover, we revealed, for the first time, their direct effects on key molecular aspects of pancreatic α-cell biology.

Screening of Relevant Metabolism-Disrupting Chemicals on Pancreatic ß-Cells: Evaluation of Murine and Human In Vitro Models.

Endocrine-disrupting chemicals (EDCs) are chemical substances that can interfere with the normal function of the endocrine system. EDCs are ubiquitous and can be found in a variety of consumer products such as food packaging materials, personal care and household products, plastic additives, and flame retardants. Over the last decade, the impact of EDCs on human health has been widely acknowledged as they have been associated with different endocrine diseases. Among them, a subset called metabolism-disrupting chemicals (MDCs) is able to promote metabolic changes that can lead to the development of metabolic disorders such as diabetes, obesity, hepatic steatosis, and metabolic syndrome, among others. Despite this, today, there are still no definitive and standardized in vitro tools to support the metabolic risk assessment of existing and emerging MDCs for regulatory purposes. Here, we evaluated the following two different pancreatic cell-based in vitro systems: the murine pancreatic ß-cell line MIN6 as well as the human pancreatic ß-cell line EndoC-ßH1. Both were challenged with the following range of relevant concentrations of seven well-known EDCs: (bisphenol-A (BPA), bisphenol-S (BPS), bisphenol-F (BPF), perfluorooctanesulfonic acid (PFOS), di(2-ethylhexyl) phthalate (DEHP), cadmium chloride (CdCl2), and dichlorodiphenyldichloroethylene (DDE)). The screening revealed that most of the tested chemicals have detectable, deleterious effects on glucose-stimulated insulin release, insulin content, electrical activity, gene expression, and/or viability. Our data provide new molecular information on the direct effects of the selected chemicals on key aspects of pancreatic ß-cell function, such as the stimulus-secretion coupling and ion channel activity. In addition, we found that, in general, the sensitivity and responses were comparable to those from other in vivo studies reported in the literature. Overall, our results suggest that both systems can serve as effective tools for the rapid screening of potential MDC effects on pancreatic ß-cell physiology as well as for deciphering and better understanding the molecular mechanisms that underlie their action.

Integrative Strategy of Testing Systems for Identification of Endocrine Disruptors Inducing Metabolic Disorders-An Introduction to the OBERON Project.

Exposure to chemical substances that can produce endocrine disrupting effects represents one of the most critical public health threats nowadays. In line with the regulatory framework implemented within the European Union (EU) to reduce the levels of endocrine disruptors (EDs) for consumers, new and effective methods for ED testing are needed. The OBERON project will build an integrated testing strategy (ITS) to detect ED-related metabolic disorders by developing, improving and validating a battery of test systems. It will be based on the concept of an integrated approach for testing and assessment (IATA). OBERON will combine (1) experimental methods (in vitro, e.g., using 2D and 3D human-derived cells and tissues, and in vivo, i.e., using zebrafish at different stages), (2) high throughput omics technologies, (3) epidemiology and human biomonitoring studies and (4) advanced computational models (in silico and systems biology) on functional endpoints related to metabolism. Such interdisciplinary framework will help in deciphering EDs based on a mechanistic understanding of toxicity by providing and making available more effective alternative test methods relevant for human health that are in line with regulatory needs. Data generated in OBERON will also allow the development of novel adverse outcome pathways (AOPs). The assays will be pre-validated in order to select the test systems that will show acceptable performance in terms of relevance for the second step of the validation process, i.e., the inter-laboratory validation as ring tests. Therefore, the aim of the OBERON project is to support the organization for economic co-operation and development (OECD) conceptual framework for testing and assessment of single and/or mixture of EDs by developing specific assays not covered by the current tests, and to propose an IATA for ED-related metabolic disorder detection, which will be submitted to the Joint Research Center (JRC) and OECD community.

Oestrogen receptor ß mediates the actions of bisphenol-A on ion channel expression in mouse pancreatic beta cells.

AIMS/HYPOTHESIS: Bisphenol-A (BPA) is a widespread endocrine-disrupting chemical that has been associated with type 2 diabetes development. Low doses of BPA modify pancreatic beta cell function and induce insulin resistance; some of these effects are mediated via activation of oestrogen receptors α (ERα) and ß (ERß). Here we investigated whether low doses of BPA regulate the expression and function of ion channel subunits involved in beta cell function. METHODS: Microarray gene profiling of isolated islets from vehicle- and BPA-treated (100 µg/kg per day for 4 days) mice was performed using Affymetrix GeneChip Mouse Genome 430.2 Array. Expression level analysis was performed using the normalisation method based on the processing algorithm 'robust multi-array average'. Whole islets or dispersed islets from C57BL/6J or oestrogen receptor ß (ERß) knockout (Erß-/-) mice were treated with vehicle or BPA (1 nmol/l) for 48 h. Whole-cell patch-clamp recordings were used to measure Na+ and K+ currents. mRNA expression was evaluated by quantitative real-time PCR. RESULTS: Microarray analysis showed that BPA modulated the expression of 1440 probe sets (1192 upregulated and 248 downregulated genes). Of these, more than 50 genes, including Scn9a, Kcnb2, Kcnma1 and Kcnip1, encoded important Na+ and K+ channel subunits. These findings were confirmed by quantitative RT-PCR in islets from C57BL/6J BPA-treated mice or whole islets treated ex vivo. Electrophysiological measurements showed a decrease in both Na+ and K+ currents in BPA-treated islets. The pharmacological profile indicated that BPA reduced currents mediated by voltage-activated K+ channels (Kv2.1/2.2 channels) and large-conductance Ca2+-activated K+ channels (KCa1.1 channels), which agrees with BPA's effects on gene expression. Beta cells from ERß-/- mice did not present BPA-induced changes, suggesting that ERß mediates BPA's effects in pancreatic islets. Finally, BPA increased burst duration, reduced the amplitude of the action potential and enlarged the action potential half-width, leading to alteration in beta cell electrical activity. CONCLUSIONS/INTERPRETATION: Our data suggest that BPA modulates the expression and function of Na+ and K+ channels via ERß in mouse pancreatic islets. Furthermore, BPA alters beta cell electrical activity. Altogether, these BPA-induced changes in beta cells might play a role in the diabetogenic action of BPA described in animal models.

Inhibition of connexin 36 hemichannels by glucose contributes to the stimulation of insulin secretion.

The existence of functional connexin36 (Cx36) hemichannels in ß-cells was investigated in pancreatic islets of rat and wild-type (Cx36(+/+)), monoallelic (Cx36(+/-)), and biallelic (Cx36(-/-)) knockout mice. Hemichannel opening by KCl depolarization was studied by measuring ATP release and changes of intracellular ATP (ADP). Cx36(+/+) islets lost ATP after depolarization with 70 mM KCl at 5 mM glucose; ATP loss was prevented by 8 and 20 mM glucose or 50 µM mefloquine (connexin inhibitor). ATP content was higher in Cx36(-/-) than Cx36(+/+) islets and was not decreased by KCl depolarization; Cx36(+/-) islets showed values between that of control and homozygous islets. Five minimolar extracellular ATP increased ATP content and ATP/ADP ratio and induced a biphasic insulin secretion in depolarized Cx36(+/+) and Cx36(+/-) but not Cx36(-/-) islets. Cx36 hemichannels expressed in oocytes opened upon depolarization of membrane potential, and their activation was inhibited by mefloquine and glucose (IC50 ∼8 mM). It is postulated that glucose-induced inhibition of Cx36 hemichannels in islet ß-cells might avoid depolarization-induced ATP loss, allowing an optimum increase of the ATP/ADP ratio by sugar metabolism and a biphasic stimulation of insulin secretion. Gradual suppression of glucose-induced insulin release in Cx36(+/-) and Cx36(-/-) islets confirms that Cx36 gap junction channels are necessary for a full secretory stimulation and might account for the glucose intolerance observed in mice with defective Cx36 expression. Mefloquine targeting of Cx36 on both gap junctions and hemichannels also suppresses glucose-stimulated secretion. By contrast, glucose stimulation of insulin secretion requires Cx36 hemichannels' closure but keeping gap junction channels opened.

Nutrient regulation of glucagon secretion: involvement in metabolism and diabetes.

Glucose homeostasis is precisely regulated by glucagon and insulin, which are released by pancreatic α- and ß-cells, respectively. While ß-cells have been the focus of intense research, less is known about α-cell function and the actions of glucagon. In recent years, the study of this endocrine cell type has experienced a renewed drive. The present review contains a summary of established concepts as well as new information about the regulation of α-cells by glucose, amino acids, fatty acids and other nutrients, focusing especially on glucagon release, glucagon synthesis and α-cell survival. We have also discussed the role of glucagon in glucose homeostasis and in energy and lipid metabolism as well as its potential as a modulator of food intake and body weight. In addition to the well-established action on the liver, we discuss the effects of glucagon in other organs, where the glucagon receptor is expressed. These tissues include the heart, kidneys, adipose tissue, brain, small intestine and the gustatory epithelium. Alterations in α-cell function and abnormal glucagon concentrations are present in diabetes and are thought to aggravate the hyperglycaemic state of diabetic patients. In this respect, several experimental approaches in diabetic models have shown important beneficial results in improving hyperglycaemia after the modulation of glucagon secretion or action. Moreover, glucagon receptor agonism has also been used as a therapeutic strategy to treat obesity.

Estrogen receptors alpha and beta in male and female gerbil prostates.

The Mongolian gerbil (Meriones unguiculatus, Gerbilinae: Muridae) is useful for prostate studies, because both males and females spontaneously develop prostatic disorders with age. Estrogens regulate prostate homeostasis via two estrogen receptors, ER alpha (ESR1) and ER beta (ESR2), but the cellular distribution and regulation of these receptors in the gerbil prostate has not been described. Both receptors were localized by immunohistochemistry in the ventral prostate of intact male and female gerbils, in males 7 and 21 days after castration, and in females treated with testosterone for 7 and 21 days. In male and female adult gerbils, ER alpha was detected mainly in prostatic stromal cells, whereas ER beta was present mostly in secretory and basal cells. More ER alpha-positive stromal cells were found in females than in males, as was a reduction toward the male value in females treated with testosterone. Castration did not alter ER alpha expression. Testosterone was necessary for maintenance of ER beta in the male prostate epithelium: ER beta expression declined markedly in prostates of males older than 1 yr, and castration of 4-mo-old males caused a reduction in ER beta to levels seen in 1-yr-old males. Because ER beta is an antiproliferative receptor, its loss with age may predispose the aging gerbil to proliferative diseases of the prostate.

ReadEDTest: A tool to assess the readiness of in vitro test methods under development for identifying endocrine disruptors.

Growing evidence shows that endocrine disruptors (EDs), known to affect the reproductive system, may also disturb other hormone-regulated functions leading to cancers, neurodevelopmental defects, metabolic and immune diseases. To reduce exposure to EDs and limit their health effects, development of screening and mechanism-based assays to identify EDs is encouraged. Nevertheless, the crucial validation step of test methods by regulatory bodies is a time- and resource-consuming process. One of the main raisons of this long duration process is that method developers, mainly researchers, are not fully aware of the regulatory needs to validate a test. We propose an online self-assessment questionnaire (SAQ) called ReadEDTest easy to be used by all researchers. The aim of ReadEDTest is to speed up the validation process by assessing readiness criteria of in vitro and fish embryo ED test methods under development. The SAQ is divided into 7 sections and 13 sub-sections containing essential information requested by the validating bodies. The readiness of the tests can be assessed by specific score limits for each sub-section. Results are displayed via a graphical representation to help identification of the sub-sections having sufficient or insufficient information. The relevance of the proposed innovative tool was supported using two test methods already validated by the OECD and four under development test methods.

A role for epithelial-mesenchymal transition in the etiology of benign prostatic hyperplasia.

Benign prostatic hyperplasia (BPH) is usually described as a pathological proliferation of prostatic fibroblasts/myofibroblasts and epithelial cells. In the present study of BPH samples, we have made a morphological and immunohistochemical study of BPH prostatic sections using markers of proliferation, apoptosis, hormone receptors, and TGF-beta signaling. We found no evidence of proliferation in the stroma but in the epithelium of some ducts; 0.7% of the basal and 0.4% of luminal cells were positive for Ki67 and PCNA. Androgen receptor and estrogen receptor beta (ERbeta)1 and ERbetacx were abundant in both stromal and epithelial compartments but cells expressing ERalpha were very rare. What was very common in all BPH samples was the following: (i) regions of the ductal epithelium where the epithelial cells did not express E-cadherin, had lost their polarization, and become spindle shaped (the nuclei of these cells were strongly positive for pSmad 3 and Snail); and (ii) regions where the walls of the blood vessels were extremely thick and there was loss of endothelial layer. Loss of E-cadherin, increased pSmad 3, and high expression of Snail are all characteristic of epithelial-mesenchymal transition (EMT). We conclude that BPH is not a disease of prostatic stromal proliferation but rather of accumulation of mesenchymal-like cells derived from the prostatic epithelium and the endothelium. TGF-beta is thought to play a key role in EMT. Our data suggests that TGF-beta/Smad should be considered as targets for treatment of BPH.

The pancreatic ß-cell in ageing: Implications in age-related diabetes.

The prevalence of type 2 diabetes (T2D) and impaired glucose tolerance (IGT) increases with ageing. T2D generally results from progressive impairment of the pancreatic islets to adapt ß-cell mass and function in the setting of insulin resistance and increased insulin demand. Several studies have shown an age-related decline in peripheral insulin sensitivity. However, a precise understanding of the pancreatic ß-cell response in ageing is still lacking. In this review, we summarize the age-related alterations, adaptations and/or failures of ß-cells at the molecular, morphological and functional levels in mouse and human. Age-associated alterations include processes such as ß-cell proliferation, apoptosis and cell identity that can influence ß-cell mass. Age-related changes also affect ß-cell function at distinct steps including electrical activity, Ca2+ signaling and insulin secretion, among others. We will consider the potential impact of these alterations and those mediated by senescence pathways on ß-cells and their implications in age-related T2D. Finally, given the great diversity of results in the field of ß-cell ageing, we will discuss the sources of this heterogeneity. A better understanding of ß-cell biology during ageing, particularly at older ages, will improve our insight into the contribution of ß-cells to age-associated T2D and may boost new therapeutic strategies.

The Effects of Aging on Male Mouse Pancreatic ß-Cell Function Involve Multiple Events in the Regulation of Secretion: Influence of Insulin Sensitivity.

Aging is associated with a decline in peripheral insulin sensitivity and an increased risk of impaired glucose tolerance and type 2 diabetes. During conditions of reduced insulin sensitivity, pancreatic ß cells undergo adaptive responses to increase insulin secretion and maintain euglycemia. However, the existence and nature of ß-cell adaptations and/or alterations during aging are still a matter of debate. In this study, we investigated the effects of aging on ß-cell function from control (3-month-old) and aged (20-month-old) mice. Aged animals were further categorized into 2 groups: high insulin sensitive (aged-HIS) and low insulin sensitive (aged-LIS). Aged-LIS mice were hyperinsulinemic, glucose intolerant, and displayed impaired glucose-stimulated insulin and C-peptide secretion, whereas aged-HIS animals showed characteristics in glucose homeostasis similar to controls. In isolated ß cells, we observed that glucose-induced inhibition of KATP channel activity was reduced with aging, particularly in the aged-LIS group. Glucose-induced islet NAD(P)H production was decreased in aged mice, suggesting impaired mitochondrial function. In contrast, voltage-gated Ca2+ currents were higher in aged-LIS ß cells, and pancreatic islets of both aged groups displayed increased glucose-induced Ca2+ signaling and augmented insulin secretion compared with controls. Morphological analysis of pancreas sections also revealed augmented ß-cell mass with aging, especially in the aged-LIS group, as well as ultrastructural ß-cell changes. Altogether, these findings indicate that aged mouse ß cells compensate for the aging-induced alterations in the stimulus-secretion coupling, particularly by adjusting their Ca2+ influx to ensure insulin secretion. These results also suggest that decreased peripheral insulin sensitivity exacerbates the effects of aging on ß cells.

Combinatorial pathway disruption is a powerful approach to delineate metabolic impacts of endocrine disruptors.

The prevalence of metabolic diseases, such as obesity, diabetes, metabolic syndrome and chronic liver diseases among others, has been rising for several years. Epidemiology and mechanistic (in vivo, in vitro and in silico) toxicology have recently provided compelling evidence implicating the chemical environment in the pathogenesis of these diseases. In this review, we will describe the biological processes that contribute to the development of metabolic diseases targeted by metabolic disruptors, and will propose an integrated pathophysiological vision of their effects on several organs. With regard to these pathomechanisms, we will discuss the needs, and the stakes of evolving the testing and assessment of endocrine disruptors to improve the prevention and management of metabolic diseases that have become a global epidemic since the end of last century.

Bisphenol-A: a new diabetogenic factor?

The aim of this review was to analyze the potential effects of environmental chemicals on homeostatic control related to glycemia and energy balance. Many of the environmental chemicals can mimic or interfere with the action of hormones and are generally referred to as "endocrine disruptors". Among these compounds, polychlorinated biphenyls, dioxins, phthalates and bisphenol-A have been correlated with alterations in blood glucose homeostasis in humans. In rodents it has been demonstrated that small doses of bisphenol-A have profound effects on glucose metabolism. Therefore, this altered blood glucose homeostasis may enhance the development of type 2 diabetes.

Morphological and functional adaptations of pancreatic alpha-cells during late pregnancy in the mouse.

BACKGROUND: Pregnancy represents a major metabolic challenge for the mother, and involves a compensatory response of the pancreatic beta-cell to maintain normoglycemia. However, although pancreatic alpha-cells play a key role in glucose homeostasis and seem to be involved in gestational diabetes, there is no information about their potential adaptations or changes during pregnancy. MATERIAL AND METHODS: Non-pregnant (controls) and pregnant C57BL/6 mice at gestational day 18.5 (G18.5) and their isolated pancreatic islets were used for in vivo and ex vivo studies, respectively. The effect of pregnancy hormones was tested in glucagon-secreting α-TC1.9 cells. Immunohistochemical analysis was performed in pancreatic slices. Glucagon gene expression was monitored by RT-qPCR. Glucagon secretion and plasma hormones were measured by ELISA. RESULTS: Pregnant mice on G18.5 exhibited alpha-cell hypertrophy as well as augmented alpha-cell area and mass. This alpha-cell mass expansion was mainly due to increased proliferation. No changes in alpha-cell apoptosis, ductal neogenesis, or alpha-to-beta transdifferentiation were found compared with controls. Pregnant mice on G18.5 exhibited hypoglucagonemia. Additionally, in vitro glucagon secretion at low glucose levels was decreased in isolated islets from pregnant animals. Glucagon content was also reduced. Experiments in α-TC1.9 cells indicated that, unlike estradiol and progesterone, placental lactogens and prolactin stimulated alpha-cell proliferation. Placental lactogens, prolactin and estradiol also inhibited glucagon release from α-TC1.9 cells at low glucose levels. CONCLUSIONS: The pancreatic alpha-cell in mice undergoes several morphofunctional changes during late pregnancy, which may contribute to proper glucose homeostasis. Gestational hormones are likely involved in these processes.

Bisphenol-A exposure during pregnancy alters pancreatic ß-cell division and mass in male mice offspring: A role for ERß.

Bisphenol-A (BPA) is a widespread endocrine disrupting chemical that constitutes a risk factor for type 2 diabetes mellitus (T2DM). Data from animal and human studies have demonstrated that early exposure to BPA results in adverse metabolic outcomes in adult life. In the present work, we exposed pregnant heterozygous estrogen receptor ß (ERß) knock out (BERKO) mice to 10 µg/kg/day BPA, during days 9-16 of pregnancy, and measured ß-cell mass and proliferation in wildtype (WT) and BERKO male offspring at postnatal day 30. We observed increased pancreatic ß-cell proliferation and mass in WT, yet no effect was produced in BERKO mice. Dispersed islet cells in primary culture treated with 1 nM BPA showed an enhanced pancreatic ß-cell replication rate, which was blunted in pancreatic ß-cells from BERKO mice and mimicked by the selective ERß agonist WAY200070. This increased ß-cell proliferation was found in male adult as well as in neonate pancreatic ß-cells, suggesting that BPA directly impacts ß-cell division at earliest stages of life. These findings strongly indicate that BPA during pregnancy upregulates pancreatic ß-cell division and mass in an ERß-dependent manner. Thus, other natural or artificial chemicals may use this ERß-mediated pathway to promote similar effects.

The role of oestrogens in the adaptation of islets to insulin resistance.

Pregnancy is characterized by peripheral insulin resistance, which is developed in parallel with a plasma increase of maternal hormones; these include prolactin, placental lactogens, progesterone and oestradiol among others. Maternal insulin resistance is counteracted by the adaptation of the islets of Langerhans to the higher insulin demand. If this adjustment is not produced, gestational diabetes may be developed. The adaptation process of islets is characterized by an increase of insulin biosynthesis, an enhanced glucose-stimulated insulin secretion (GSIS) and an increase of beta-cell mass. It is not completely understood why, in some individuals, beta-cell mass and function fail to adapt to the metabolic demands of pregnancy, yet a disruption of the beta-cell response to maternal hormones may play a key part. The role of the maternal hormone 17beta-oestradiol (E2) in this adaptation process has been largely unknown. However, in recent years, it has been demonstrated that E2 acts directly on beta-cells to increase insulin biosynthesis and to enhance GSIS through different molecular mechanisms. E2 does not increase beta-cell proliferation but it is involved in beta-cell survival. Classical oestrogen receptors ERalpha and ERbeta, as well as the G protein-coupled oestrogen receptor (GPER) seem to be involved in these adaptation changes. In addition, as the main production of E2 in post-menopausal women comes from the adipose tissue, E2 may act as a messenger between adipocytes and islets in obesity.

Bisphenol A Regulates Sodium Ramp Currents in Mouse Dorsal Root Ganglion Neurons and Increases Nociception.

17ß-Estradiol mediates the sensitivity to pain and is involved in sex differences in nociception. The widespread environmental disrupting chemical bisphenol A (BPA) has estrogenic activity, but its implications in pain are mostly unknown. Here we show that treatment of male mice with BPA (50 µg/kg/day) during 8 days, decreases the latency to pain behavior in response to heat, suggesting increased pain sensitivity. We demonstrate that incubation of dissociated dorsal root ganglia (DRG) nociceptors with 1 nM BPA increases the frequency of action potential firing. SCN9A encodes the voltage-gated sodium channel Nav1.7, which is present in DRG nociceptors and is essential in pain signaling. Nav1.7 and other voltage-gated sodium channels in mouse DRG are considered threshold channels because they produce ramp currents, amplifying small depolarizations and enhancing electrical activity. BPA increased Nav-mediated ramp currents elicited with slow depolarizations. Experiments using pharmacological tools as well as DRG from ERß-/- mice indicate that this BPA effect involves ERα and phosphoinositide 3-kinase. The mRNA expression and biophysical properties other than ramp currents of Nav channels, were unchanged by BPA. Our data suggest that BPA at environmentally relevant doses affects the ability to detect noxious stimuli and therefore should be considered when studying the etiology of pain conditions.

In utero exposure to bisphenol-A disrupts key elements of retinoid system in male mice offspring.

The retinoid system controls essential cellular processes including mitosis, differentiation and metabolism among others. Although the retinoid-signalling pathway is a potential target for the action of several endocrine disrupting chemicals (EDCs), the information about the developmental effects of bisphenol-A (BPA) on the hepatic retinoid system is scarce. Herein, male mice were in utero exposed to BPA following maternal subcutaneous doses of 0, 10 and 100 µg/kg bw/day from gestational day 9-16 and they were sacrificed at post-natal day 30. Retinoid concentrations and gene expression of key elements involved in the retinoid system were determined in liver. BPA increased all-trans-retinoic acid concentration and expression of Adh1, Aox1 and Cyp1a2 (biosynthesis of retinoic acid), while reduced Mrp3 (efflux from hepatocyte to blood), increased Bcrp expression (biliary excretion) and changed the retinoid-dependent signalling system after reducing expression of Rxrß and increasing that of Fgf21. Furthermore, we found bivariate associations of Rarγ and Rxrγ expressions with all-trans-retinoic acid concentrations and of Fgf21 expression with that of Rarγ. Those findings occurred in animals which showed altered pancreatic function and impaired glucose metabolism during adulthood. The present information should be useful for enhancing testing methods for the identification of EDCs.

Pancreatic alpha-cell mass in the early-onset and advanced stage of a mouse model of experimental autoimmune diabetes.

Most studies in type 1 diabetes (T1D) have focused on the loss of the pancreatic beta-cell population. However, despite the involvement of the alpha-cell in the aetiology and complications of T1D, little is known about the regulation of the pancreatic alpha-cell mass in this disease. The need for a better understanding of this process is further emphasized by recent findings suggesting that alpha-cells may constitute a potential reservoir for beta-cell regeneration. In this study, we characterized the pancreatic alpha-cell mass and its regulatory processes in the transgenic RIP-B7.1 mice model of experimental autoimmune diabetes (EAD). Diabetic mice presented insulitis, hyperglycaemia, hypoinsulinemia and hyperglucagonemia along with lower pancreatic insulin content. While alpha-cell mass and pancreatic glucagon content were preserved at the early-onset of EAD, both parameters were reduced in the advanced phase. At both stages, alpha-cell size, proliferation and ductal neogenesis were up-regulated, whereas apoptosis was almost negligible. Interestingly, we found an increase in the proportion of glucagon-containing cells positive for insulin or the beta-cell transcription factor PDX1. Our findings suggest that pancreatic alpha-cell renewal mechanisms are boosted during the natural course of EAD, possibly as an attempt to maintain the alpha-cell population and/or to increase beta-cell regeneration via alpha-cell transdifferentiation.

Cortistatin regulates glucose-induced electrical activity and insulin secretion in mouse pancreatic beta-cells.

Although there is growing evidence that cortistatin regulates several functions in different tissues, its role in the endocrine pancreas is not totally known. Here, we aim to study the effect of cortistatin on pancreatic beta-cells and glucose-stimulated insulin secretion (GSIS). Exposure of isolated mouse islets to cortistatin inhibited GSIS. This effect was prevented using a somatostatin receptor antagonist. Additionally, cortistatin hyperpolarized the membrane potential and reduced glucose-induced action potentials in isolated pancreatic beta-cells. Cortistatin did not modify ATP-dependent K+ (KATP) channel activity. In contrast, cortistatin increased the activity of a small conductance channel with characteristics of G protein-coupled inwardly rectifying K+ (GIRK) channels. The cortistatin effects on membrane potential and GSIS were largely reduced in the presence of a GIRK channel antagonist and by down-regulation of GIRK2 with small interfering RNA. Thus, cortistatin acts as an inhibitory signal for glucose-induced electrical activity and insulin secretion in the mouse pancreatic beta-cell.