Esther Fuentes

The Estrogenic Effect of Bisphenol A Disrupts Pancreatic β-Cell Function In Vivo and Induces Insulin Resistance

The function of the pancreatic β-cell is the storage and release of insulin, the main hormone involved in blood glucose homeostasis. The results in this article show that the widespread environmental contaminant bisphenol-A (BPA) imitates 17β-estradiol (E2) effects in vivo on blood glucose homeostasis through genomic and nongenomic pathways. The exposure of adult mice to a single low dose (10 μg/kg) of either E2 or BPA induces a rapid decrease in glycemia that correlates with a rise of plasma insulin. Longer exposures to E2 and BPA induce an increase in pancreatic β-cell insulin content in an estrogen-receptor–dependent manner. This effect is visible after 2 days of treatment and starting at doses as low as 10 μg/kg/day. After 4 days of treatment with either E2 or BPA, these mice developed chronic hyperinsulinemia, and their glucose and insulin tolerance tests were altered. These experiments unveil the link between environmental estrogens and insulin resistance. Therefore, either abnormal levels of endogenous estrogens or environmental estrogen exposure enhances the risk of developing type 2 diabetes mellitus, hypertension, and dyslipidemia.

Low Doses of Bisphenol A and Diethylstilbestrol Impair Ca2+ Signals in Pancreatic α-Cells through a Nonclassical Membrane Estrogen Receptor within Intact Islets of Langerhans

Glucagon, secreted from pancreatic α-cells integrated within the islets of Langerhans, is involved in the regulation of glucose metabolism by enhancing the synthesis and mobilization of glucose in the liver. In addition, it has other extrahepatic effects ranging from lipolysis in adipose tissue to the control of satiety in the central nervous system. In this article, we show that the endocrine disruptors bisphenol A (BPA) and diethylstilbestrol (DES), at a concentration of 10−9 M, suppressed low-glucose–induced intracellular calcium ion ([Ca2+]i) oscillations in α-cells, the signal that triggers glucagon secretion. This action has a rapid onset, and it is reproduced by the impermeable molecule estradiol (E2) conjugated to horseradish peroxidase (E-HRP). Competition studies using E-HRP binding in immunocytochemically identified α-cells indicate that 17β-E2, BPA, and DES share a common membrane-binding site whose pharmacologic profile differs from the classical ER. The effects triggered by BPA, DES, and E2 are blocked by the Gαi- and Gαo-protein inhibitor pertussis toxin, by the guanylate cyclase–specific inhibitor 1H-[1,2,4] oxadiazolo[4,3-a] quinoxalin-1-one, and by the nitric oxide synthase inhibitor N-nitro-l-arginine methyl ester. The effects are reproduced by 8-bromo-guanosine 3′,5′-cyclic monophosphate and suppressed in the presence of the cGMP-dependent protein kinase inhibitor KT-5823. The action of E2, BPA, and DES in pancreatic α-cells may explain some of the effects elicited by endocrine disruptors in the metabolism of glucose and lipid.

Low doses of the endocrine disruptor bisphenol-A and the native hormone 17beta-estradiol rapidly activate transcription factor CREB.

Endocrine‐disrupting chemicals (EDCs) are hormone‐like agents present in the environment that alter the endocrine system of wildlife and humans. Most EDCs have potencies far below those of the natural hormone 17β‐E2 when acting through the classic estrogen receptors (ERs). Here, we show that the environmental estrogen Bisphenol‐A and the native hormone 17β‐E2 activate the transcription factor, cAMP‐responsive element binding protein (CREB) with the same potency. Phosphorylated CREB (P‐CREB) was increased after only a 5‐minute application of either BPA or 17β‐E2 in a calcium‐dependent manner. The effect was reproduced by the membrane‐impermeable molecule E2 conjugated to horseradish peroxidase (E‐HRP). The increase in PCREB was not modified by the anti‐estrogen ICI 182,780. Therefore, low‐dose of BPA activates the transcription factor CREB via an alternative mechanism, involving a non‐classical membrane estrogen receptor. Because these effects are elicited at concentrations as low as 10–9 M, this observation is of environmental and public health relevance.

Bisphenol-A acts as a potent estrogen via non-classical estrogen triggered pathways

Bisphenol-A (BPA) is an estrogenic monomer commonly used in the manufacture of numerous consumer products such as food and beverage containers. Widespread human exposure to significant doses of this compound has been reported. Traditionally, BPA has been considered a weak estrogen, based on its lower binding affinity to the nuclear estrogen receptors (ERs) compared to 17-β estradiol (E2) as well as its low transcriptional activity after ERs activation. However, in vivo animal studies have demonstrated that it can interfere with endocrine signaling pathways at low doses during fetal, neonatal or perinatal periods as well as in adulthood. In addition, mounting evidence suggests a variety of pathways through which BPA can elicit cellular responses at very low concentrations with the same or even higher efficiency than E2. Thus, the purpose of the present review is to analyze with substantiated scientific evidence the strong estrogenic activity of BPA when it acts through alternative mechanisms of action at least in certain cell types.

Presence of functional cannabinoid receptors in human endocrine pancreas

Aims/hypothesisWe examined the presence of functional cannabinoid receptors 1 and 2 (CB1, CB2) in isolated human islets, phenotyped the cells producing cannabinoid receptors and analysed the actions of selective cannabinoid receptor agonists on insulin, glucagon and somatostatin secretion in vitro. We also described the localisation on islet cells of: (1) the endocannabinoid-producing enzymes N-acyl-phosphatidyl ethanolamine-hydrolysing phospholipase D and diacylglycerol lipase; and (2) the endocannabinoid-degrading enzymes fatty acid amidohydrolase and monoacyl glycerol lipase.MethodsReal-time PCR, western blotting and immunocytochemistry were used to analyse the presence of endocannabinoid-related proteins and genes. Static secretion experiments were used to examine the effects of activating CB1 or CB2 on insulin, glucagon and somatostatin secretion and to measure changes in 2-arachidonoylglycerol (2-AG) levels within islets. Analyses were performed in isolated human islets and in paraffin-embedded sections of human pancreas.ResultsHuman islets of Langerhans expressed CB1 and CB2 (also known as CNR1 and CNR2) mRNA and CB1 and CB2 proteins, and also the machinery involved in synthesis and degradation of 2-AG (the most abundant endocannabinoid, levels of which were modulated by glucose). Immunofluorescence revealed that CB1 was densely located in glucagon-secreting alpha cells and less so in insulin-secreting beta cells. CB2 was densely present in somatostatin-secreting delta cells, but absent in alpha and beta cells. In vitro experiments revealed that CB1 stimulation enhanced insulin and glucagon secretion, while CB2 agonism lowered glucose-dependent insulin secretion, showing these cannabinoid receptors to be functional.Conclusions/interpretationTogether, these results suggest a role for endogenous endocannabinoid signalling in regulation of endocrine secretion in the human pancreas.

Non‐genomic actions of 17β‐oestradiol in mouse pancreatic β‐cells are mediated by a cGMP‐dependent protein kinase

1 Intracellular calcium concentration ([Ca2+]i) was measured in mouse whole islets of Langerhans using the calcium‐sensitive fluorescent dye Indo‐1. 2 Application of physiological concentrations of 17β‐oestradiol in the presence of a stimulatory glucose concentration (8 mm) potentiated the [Ca2+]i signal in 83 % of islets tested. Potentiation was manifested as either an increase in the frequency or duration of [Ca2+]i oscillations. 3 The effects caused by 17β‐oestradiol were mimicked by the cyclic nucleotide analogues 8‐bromoguanosine‐3′,5′‐cyclic monophosphate (8‐Br‐cGMP) and 8‐bromoadenosine‐3′,5′‐cyclic monophosphate (8‐Br‐cAMP). 4 Direct measurements of both cyclic nucleotides demonstrated that nanomolar concentrations of 17β‐oestradiol in the presence of 8 mm glucose increased cGMP levels, yet cAMP levels were unchanged. The increment in cGMP was similar to that induced by 11 mm glucose. 5 Patch‐clamp recording in intact cells showed that 8‐Br‐cGMP reproduced the inhibitory action of 17β‐oestradiol on ATP‐sensitive K+ (KATP) channel activity. This was not a membrane‐bound effect since it could not be observed in excised patches. 6 The action of 17β‐oestradiol on KATP channel activity was not modified by the specific inhibitor of soluble guanylate cyclase (sGC) LY 83583. This result indicates a likely involvement of a membrane guanylate cyclase (mGC). 7 The rapid decrease in KATP channel activity elicited by 17β‐oestradiol was greatly reduced using Rp‐8‐pCPT‐cGMPS, a specific blocker of cGMP‐dependent protein kinase (PKG). Conversely, Rp‐cAMPS, which inhibits cAMP‐dependent protein kinase (PKA), had little effect. 8 The results presented here indicate that rapid, non‐genomic effects of 17β‐oestradiol after interaction with its binding site at the plasma membrane of pancreatic β‐cells is a cGMP‐dependent phosphorylation process.

The plasma membrane estrogen receptor: nuclear or unclear?

Abstract Although the existence of a plasma membrane estrogen receptor (pmER) is widely accepted, its molecular structure is still undetermined. Several studies have proposed a range of structures, from a unique protein identical to the classical nuclear estrogen receptor to a completely novel pmER.

Estrogen and xenoestrogen actions on endocrine pancreas : from ion channel modulation to activation of nuclear function

17beta-Estradiol elicits a rapid opposite effect on [Ca2+]i in alpha- and beta-cells within intact islets of Langerhans. In beta-cells, physiological concentrations of the gonadal hormone decreases KATP channel activity in synergy with glucose, leading to a membrane depolarization that opens voltage-gated Ca2+ channels, potentiating Ca2+ signals. As a consequence insulin release is enhanced and transcription factor CREB is activated in a Ca(2+)-dependent manner. In glucagon-containing alpha-cells, 17beta-estradiol provokes the abolishment of Ca2+ oscillations generated by low glucose, a situation that should decrease glucagon release. In both types of cells the second messenger involved is cGMP. The estrogen receptor involved is located in the plasma membrane and has a pharmacological profile unrelated to classical estrogen receptors ERalpha and ERbeta. For that reason, it has been named non-classical membrane estrogen receptor (ncmER). Although the physiological roles of this receptor are still unknown, it may be implicated in the responses of the endocrine pancreas to the physiological and pathological changes of 17beta-estradiol.

Rapid endocrine disruption: Environmental estrogen actions triggered outside the nucleus☆

An exogenous substance is defined as an endocrine disrupter chemical (EDC) if it alters the function of the endocrine system provoking adverse health effects. Environmental estrogens are the most studied EDCs. They follow the same mechanisms of action as the gonadal hormone 17beta-estradiol. Up to now, the estrogenicity of environmental estrogenic pollutants has been based on the property of these compounds to bind to estrogen receptors (ERs), either ERalpha or ERbeta, and to act subsequently as transcription factors when binding to the estrogen response element (ERE) in the DNA. All the estrogenic bioassays currently used are based on this mechanism of action. New evidence indicates that the definition of estrogenicity for a chemical should take into account other estrogen receptors as well as new signaling pathways. These include the activation of additional transcription factors as well as the action of xenoestrogens through estrogen receptors located outside the nucleus: in the plasma membrane, mitochondria and probably the cytosol. Therefore, new estrogenic bioassays should be developed to include the novel concept of rapid endocrine disruption.

ACTIONS OF SERUM AND PLASMA ALBUMIN ON INTRACELLULAR CA2+ IN HUMAN ENDOTHELIAL CELLS

1 The effects of serum and plasma albumin on [Ca2+]i in human endothelial cells were examined using single‐cell Ca2+ imaging. Two types of endothelial cell were used: human umbilical vein endothelial cells (HUVEC) in primary culture, and the endothelial‐derived cell line ECV304. 2 Serum albumin caused a large and transient rise in [Ca2+]i, due to Ca2+ release from an IP3‐sensitive internal store, followed by a maintained elevation in [Ca2+]i attributable to Ca2+ influx from the external medium. A half‐maximal rise in [Ca2+]i was produced by a concentration of serum albumin of about 1 μg ml−1. 3 The Ca2+‐releasing action of serum albumin is abolished by methanol extraction and is therefore attributable to an attached polar lipid. A possible candidate is lysophosphatidic acid, known to be released from platelets during blood coagulation, which produced similar effects to those of serum albumin. 4 In HUVEC, plasma albumin caused a sustained decrease in [Ca2+]i from the mean resting level of 114 nm to 58 nm. No effect of plasma albumin was observed in ECV304 cells. 5 The decrease in [Ca2+]i caused by plasma albumin is due to an uptake into intracellular stores. The store loading substantially potentiates the action of Ca2+‐releasing agonists such as histamine. 6 The results show that normal plasma albumin, which carries few lipids, lowers [Ca2+]i and potentiates the actions of Ca2+‐releasing agonists by promoting Ca2+ uptake into intracellular stores. When converted to the serum form, by binding lysophosphatidic acid released during blood coagulation, albumin has a potent effect in elevating [Ca2+]i. Blood coagulation may therefore play a role in regulating vascular tone and capillary permeability.