Selenomethionine modulates insulin secretion in the MIN6-K8 mouse insulinoma cell line.

Selenium is an essential trace element of interest for its potential role in glucose homeostasis. The present study investigated the impact of selenium supplementation as selenomethionine (SeMet) on insulin secretion in MIN6-K8 cells, a pancreatic ß-cell model. We found that SeMet enhanced percent glucose-induced insulin secretion, while also increasing tolbutamide- and KCl-induced percent insulin secretion. RNA-sequencing showed that SeMet supplementation altered expression of several selenoproteins, including glutathione peroxidase 3 (Gpx3) and selenoprotein P (SelP). Targeted knockdown of Gpx3 increased both percent and total insulin release, while SelP knockdown increased insulin content and insulin release. Collectively, these studies support a putative role for selenium and selenoproteins in the regulation of insulin secretion, glucose homeostasis, and diabetes risk.

Dietary Selenium Deficiency Partially Mimics the Metabolic Effects of Arsenic.

Chronic arsenic exposure via drinking water is associated with diabetes in human pop-ulations throughout the world. Arsenic is believed to exert its diabetogenic effects via multiple mechanisms, including alterations to insulin secretion and insulin sensitivity. In the past, acute arsenicosis has been thought to be partially treatable with selenium supplementation, though a potential interaction between selenium and arsenic had not been evaluated under longer-term exposure models. The purpose of the present study was to explore whether selenium status may augment arsenic's effects during chronic arsenic exposure. To test this possibility, mice were exposed to arsenic in their drinking water and provided ad libitum access to either a diet replete with selenium (Control) or deficient in selenium (SelD). Arsenic significantly improved glucose tolerance and decreased insulin secretion and ß-cell function in vivo. Dietary selenium deficiency resulted in similar effects on glucose tolerance and insulin secretion, with significant interactions between arsenic and dietary conditions in select insulin-related parameters. The findings of this study highlight the complexity of arsenic's metabolic effects and suggest that selenium deficiency may interact with arsenic exposure on ß-cell-related physiological parameters.

Dimethyl sulfoxide acutely enhances regulated insulin secretion in the MIN6-K8 mouse insulinoma cell line.

Diabetes mellitus is a metabolic disorder projected to afflict 700 million people globally by 2045. Fundamental to the progression of diabetes is an insufficient supply of insulin to meet metabolic demand. The MIN6-K8 cell line is a mouse insulinoma model of pancreatic ß-cells frequently used to study the mechanisms of insulin secretion. Here, we evaluated the effects of short-term exposure to dimethyl sulfoxide (DMSO), a polar aprotic solvent commonly used in drug screening, on physiological characteristics of MIN6-K8 cells. Short-term exposure of MIN6-K8 cells to DMSO enhanced glucose-induced and tolbutamide-stimulated insulin secretion without significant effects on basal secretion or potassium responsiveness. Calcium influx was enhanced during glucose and tolbutamide treatments, suggesting that DMSO's mechanism of action is upstream of calcium-dependent insulin granule exocytosis. Based on these studies, investigators should use caution when conducting experiments with DMSO in the MIN6-K8 cell line and should report all DMSO concentrations when used as a solvent.

Arsenic Exposure Decreases Adiposity During High-Fat Feeding.

OBJECTIVE: Arsenic is an endocrine-disrupting chemical associated with diabetes risk. Increased adiposity is a significant risk factor for diabetes and its comorbidities. Here, the impact of chronic arsenic exposure on adiposity and metabolic health was assessed in mice. METHODS: Male C57BL/6J mice were provided ad libitum access to a normal or high-fat diet and water +/- 50 mg/L of sodium arsenite. Changes in body weight, body composition, insulin sensitivity, energy expenditure, and locomotor activity were measured. Measures of adiposity were compared with accumulated arsenic in the liver. RESULTS: Despite uniform arsenic exposure, internal arsenic levels varied significantly among arsenic-exposed mice. Hepatic arsenic levels in exposed mice negatively correlated with overall weight gain, individual adipose depot masses, and hepatic triglyceride accumulation. No effects were observed in mice on a normal diet. For mice on a high-fat diet, arsenic exposure reduced fasting insulin levels, homeostatic model assessment of insulin resistance and ß-cell function, and systemic insulin resistance. Arsenic exposure did not alter energy expenditure or activity. CONCLUSIONS: Collectively, these data indicate that arsenic is antiobesogenic and that concentration at the source poorly predicts arsenic accumulation and phenotypic outcomes. In future studies, investigators should consider internal accumulation of arsenic rather than source concentration when assessing the outcomes of arsenic exposure.

Braving the Element: Pancreatic ß-Cell Dysfunction and Adaptation in Response to Arsenic Exposure.

Type 2 diabetes mellitus (T2DM) is a serious global health problem, currently affecting an estimated 451 million people worldwide. T2DM is characterized by hyperglycemia and low insulin relative to the metabolic demand. The precise contributing factors for a given individual vary, but generally include a combination of insulin resistance and insufficient insulin secretion. Ultimately, the progression to diabetes occurs only after ß-cells fail to meet the needs of the individual. The stresses placed upon ß-cells in this context manifest as increased oxidative damage, local inflammation, and ER stress, often inciting a destructive spiral of ß-cell death, increased metabolic stress due to further insufficiency, and additional ß-cell death. Several pathways controlling insulin resistance and ß-cell adaptation/survival are affected by a class of exogenous bioactive compounds deemed endocrine disrupting chemicals (EDCs). Epidemiological studies have shown that, in several regions throughout the world, exposure to the EDC inorganic arsenic (iAs) correlates significantly with T2DM. It has been proposed that a lifetime of exposure to iAs may exacerbate problems with both insulin sensitivity as well as ß-cell function/survival, promoting the development of T2DM. This review focuses on the mechanisms of iAs action as they relate to known adaptive and maladaptive pathways in pancreatic ß-cells.

Arsenic modifies serotonin metabolism through glucuronidation in pancreatic ß-cells

In arsenic-endemic regions of the world, arsenic exposure correlates with diabetes mellitus. Multiple animal models of inorganic arsenic (iAs, as As3+) exposure have revealed that iAs-induced glucose intolerance manifests as a result of pancreatic ß-cell dysfunction. To define the mechanisms responsible for this ß-cell defect, the MIN6-K8 mouse ß-cell line was exposed to environmentally relevant doses of iAs. Exposure to 0.1-1 µM iAs for 3 days significantly decreased glucose-induced insulin secretion (GIIS). Serotonin and its precursor, 5-hydroxytryptophan (5-HTP), were both decreased. Supplementation with 5-HTP, which loads the system with bioavailable 5-HTP and serotonin, rescued GIIS, suggesting that recovery of this pathway was sufficient to restore function. Exposure to iAs was accompanied by an increase in mRNA expression of UDP-glucuronosyltransferase 1 family, polypeptide a6a (Ugt1a6a), a phase-II detoxification enzyme that facilitates the disposal of cyclic amines, including serotonin, via glucuronidation. Elevated Ugt1a6a and UGT1A6 expression levels were observed in mouse and human islets, respectively, following 3 days of iAs exposure. Consistent with this finding, the enzymatic rate of serotonin glucuronidation was increased in iAs-exposed cells. Knockdown by siRNA of Ugt1a6a during iAs exposure restored GIIS in MIN6-K8 cells. This effect was prevented by blockade of serotonin biosynthesis, suggesting that the observed iAs-induced increase in Ugt1a6a affects GIIS by targeting serotonin or serotonin-related metabolites. Although it is not yet clear exactly which element(s) of the serotonin pathway is/are most responsible for iAs-induced GIIS dysfunction, this study provides evidence that UGT1A6A, acting on the serotonin pathway, regulates GIIS under both normal and pathological conditions.

Arsenic exposure induces glucose intolerance and alters global energy metabolism.

Environmental pollutants acting as endocrine-disrupting chemicals (EDCs) are recognized as potential contributors to metabolic disease pathogenesis. One such pollutant, arsenic, contaminates the drinking water of ~100 million people globally and has been associated with insulin resistance and diabetes in epidemiological studies. Despite these observations, the precise metabolic derangements induced by arsenic remain incompletely characterized. In the present study, the impact of arsenic on in vivo metabolic physiology was examined in 8-wk-old male C57BL/6J mice exposed to 50 mg/l inorganic arsenite in their drinking water for 8 wk. Glucose metabolism was assessed via in vivo metabolic testing, and feeding behavior was analyzed using indirect calorimetry in metabolic cages. Pancreatic islet composition was assessed via immunofluorescence microscopy. Arsenic-exposed mice exhibited impaired glucose tolerance compared with controls; however, no difference in peripheral insulin resistance was noted between groups. Instead, early insulin release during glucose challenge was attenuated relative to the rise in glycemia. Despite decreased insulin secretion, pancreatic ß-cell mass was not altered, suggesting that arsenic primarily disrupts ß-cell function. Finally, metabolic cage analyses revealed that arsenic exposure induced novel alterations in the diurnal rhythm of food intake and energy metabolism. Taken together, these data suggest that arsenic exposure impairs glucose tolerance through functional impairments in insulin secretion from ß-cells rather than by augmenting peripheral insulin resistance. Further elucidation of the mechanisms underlying arsenic-induced behavioral and ß-cell-specific metabolic disruptions will inform future intervention strategies to address this ubiquitous environmental contaminant and novel diabetes risk factor.

Glycogen Repletion in Brown Adipose Tissue upon Refeeding Is Primarily Driven by Phosphorylation-Independent Mechanisms.

Glycogen storage in brown adipose tissue (BAT) is generally thought to take place through passive, substrate-driven activation of glycogenesis rather than programmatic shifts favoring or opposing the storage and/or retention of glycogen. This perception exists despite a growing body of evidence suggesting that BAT glycogen storage is actively regulated by covalent modification of key glycogen-metabolic enzymes, protein turnover, and endocrine hormone signaling. Members of one such class of covalent-modification regulators, glycogen-binding Phosphoprotein Phosphatase-1 (PP1)-regulatory subunits (PPP1Rs), targeting PP1 to glycogen-metabolic enzymes, were dynamically regulated in response to 24 hr of starvation and/or 24 hr of starvation followed by ad libitum refeeding. Over-expression of the PPP1R Protein Targeting to Glycogen (PTG), under the control of the aP2 promoter in mice, inactivated glycogen phosphorylase (GP) and enhanced basal- and starvation-state glycogen storage. Total interscapular BAT glycogen synthase and the constitutive activity of GS were conditionally affected. During starvation, glucose-6-phosphate (G-6-P) levels and the relative phosphorylation of Akt (p-Ser-473-Akt) were both increased in PTG-overexpressing (Tg) mice, suggesting that elevated glycogen storage during starvation modifies broader cellular metabolic pathways. During refeeding, Tg and WT mice reaccumulated glycogen similarly despite altered GS and GP activities. All observations during refeeding suggest that the phosphorylation states of GS and GP are not physiologically rate-controlling, despite there being a clear balance of endogenous kinase- and phosphatase activities. The studies presented here reveal IBAT glycogen storage to be a tightly-regulated process at all levels, with potential effects on nutrient sensing in vivo.

Systemic regulation of adipose metabolism.

White adipose tissue serves as a critical energy storage depot and endocrine organ. Adipocytes are subject to numerous levels of regulation, including neuronal, endocrine and metabolic. While insulin is the classical endocrine regulator of lipid metabolism in adipose tissue, other important endocrine hormones also control adipose tissue physiology. In this review, we will focus on the contribution of the pituitary in the modulation of adipocyte function, through the direct release of growth hormone as well as via the regulation of the thyroid gland and release of thyroid hormone. This article is part of a Special Issue entitled: Modulation of Adipose Tissue in Health and Disease.

Refeeding-induced brown adipose tissue glycogen hyper-accumulation in mice is mediated by insulin and catecholamines.

Brown adipose tissue (BAT) generates heat during adaptive thermogenesis through a combination of oxidative metabolism and uncoupling protein 1-mediated electron transport chain uncoupling, using both free-fatty acids and glucose as substrate. Previous rat-based work in 1942 showed that prolonged partial fasting followed by refeeding led to a dramatic, transient increase in glycogen stores in multiple fat depots. In the present study, the protocol was replicated in male CD1 mice, resulting in a 2000-fold increase in interscapular BAT (IBAT) glycogen levels within 4-12 hours (hr) of refeeding, with IBAT glycogen stores reaching levels comparable to fed liver glycogen. Lesser effects occurred in white adipose tissues (WAT). Over the next 36 hr, glycogen levels dissipated and histological analysis revealed an over-accumulation of lipid droplets, suggesting a potential metabolic connection between glycogenolysis and lipid synthesis. 24 hr of total starvation followed by refeeding induced a robust and consistent glycogen over-accumulation similar in magnitude and time course to the prolonged partial fast. Experimentation demonstrated that hyperglycemia was not sufficient to drive glycogen accumulation in IBAT, but that elevated circulating insulin was sufficient. Additionally, pharmacological inhibition of catecholamine production reduced refeeding-induced IBAT glycogen storage, providing evidence of a contribution from the central nervous system. These findings highlight IBAT as a tissue that integrates both canonically-anabolic and catabolic stimulation for the promotion of glycogen storage during recovery from caloric deficit. The preservation of this robust response through many generations of animals not subjected to food deprivation suggests that the over-accumulation phenomenon plays a critical role in IBAT physiology.

Chronic social isolation is associated with metabolic gene expression changes specific to mammary adipose tissue.

Chronic social isolation is linked to increased mammary tumor growth in rodent models of breast cancer. In the C3(1)/SV40 T-antigen FVB/N (TAg) mouse model of "triple-negative" breast cancer, the heightened stress response elicited by social isolation has been associated with increased expression of metabolic genes in the mammary gland before invasive tumors develop (i.e., during the in situ carcinoma stage). To further understand the mechanisms underlying how accelerated mammary tumor growth is associated with social isolation, we separated the mammary gland adipose tissue from adjacent ductal epithelial cells and analyzed individual cell types for changes in metabolic gene expression. Specifically, increased expression of the key metabolic genes Acaca, Hk2, and Acly was found in the adipocyte, rather than the epithelial fraction. Surprisingly, metabolic gene expression was not significantly increased in visceral adipose depots of socially isolated female mice. As expected, increased metabolic gene expression in the mammary adipocytes of socially isolated mice coincided with increased glucose metabolism, lipid synthesis, and leptin secretion from this adipose depot. Furthermore, application of media that had been cultured with isolated mouse mammary adipose tissue (conditioned media) resulted in increased proliferation of mammary cancer cells relative to group-housed-conditioned media. These results suggest that exposure to a chronic stressor (social isolation) results in specific metabolic reprogramming in mammary gland adipocytes that in turn contributes to increased proliferation of adjacent preinvasive malignant epithelial cells. Metabolites and/or tumor growth-promoting proteins secreted from adipose tissue could identify biomarkers and/or targets for preventive intervention in breast cancer.

PCB 126 and other dioxin-like PCBs specifically suppress hepatic PEPCK expression via the aryl hydrocarbon receptor.

Dioxins and dioxin-like compounds encompass a group of structurally related heterocyclic compounds that bind to and activate the aryl hydrocarbon receptor (AhR). The prototypical dioxin is 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a highly toxic industrial byproduct that incites numerous adverse physiological effects. Global commercial production of the structurally similar polychlorinated biphenyls (PCBs), however, commenced early in the 20(th) century and continued for decades; dioxin-like PCBs therefore contribute significantly to total dioxin-associated toxicity. In this study, PCB 126, the most potent dioxin-like PCB, was evaluated with respect to its direct effects on hepatic glucose metabolism using primary mouse hepatocytes. Overnight treatment with PCB 126 reduced hepatic glycogen stores in a dose-dependent manner. Additionally, PCB 126 suppressed forskolin-stimulated gluconeogenesis from lactate. These effects were independent of acute toxicity, as PCB 126 did not increase lactate dehydrogenase release nor affect lipid metabolism or total intracellular ATP. Interestingly, provision of cells with glycerol instead of lactate as the carbon source completely restored hepatic glucose production, indicating specific impairment in the distal arm of gluconeogenesis. In concordance with this finding, PCB 126 blunted the forskolin-stimulated increase in phosphoenolpyruvate carboxykinase (PEPCK) mRNA levels without affecting glucose-6-phosphatase expression. Myricetin, a putative competitive AhR antagonist, reversed the suppression of PEPCK induction by PCB 126. Furthermore, other dioxin-like PCBs demonstrated similar effects on PEPCK expression in parallel with their ability to activate AhR. It therefore appears that AhR activation mediates the suppression of PEPCK expression by dioxin-like PCBs, suggesting a role for these pollutants as disruptors of energy metabolism.

Deletion of the mammalian INDY homolog mimics aspects of dietary restriction and protects against adiposity and insulin resistance in mice.

Reduced expression of the Indy (I'm Not Dead, Yet) gene in D. melanogaster and its homolog in C. elegans prolongs life span and in D. melanogaster augments mitochondrial biogenesis in a manner akin to caloric restriction. However, the cellular mechanism by which Indy does this is unknown. Here, we report on the knockout mouse model of the mammalian Indy (mIndy) homolog, SLC13A5. Deletion of mIndy in mice (mINDY(-/-) mice) reduces hepatocellular ATP/ADP ratio, activates hepatic AMPK, induces PGC-1α, inhibits ACC-2, and reduces SREBP-1c levels. This signaling network promotes hepatic mitochondrial biogenesis, lipid oxidation, and energy expenditure and attenuates hepatic de novo lipogenesis. Together, these traits protect mINDY(-/-) mice from the adiposity and insulin resistance that evolve with high-fat feeding and aging. Our studies demonstrate a profound effect of mIndy on mammalian energy metabolism and suggest that mINDY might be a therapeutic target for the treatment of obesity and type 2 diabetes.

Prevention of hepatic steatosis and hepatic insulin resistance by knockdown of cAMP response element-binding protein.

In patients with poorly controlled type 2 diabetes mellitus (T2DM), hepatic insulin resistance and increased gluconeogenesis contribute to fasting and postprandial hyperglycemia. Since cAMP response element-binding protein (CREB) is a key regulator of gluconeogenic gene expression, we hypothesized that decreasing hepatic CREB expression would reduce fasting hyperglycemia in rodent models of T2DM. In order to test this hypothesis, we used a CREB-specific antisense oligonucleotide (ASO) to knock down CREB expression in liver. CREB ASO treatment dramatically reduced fasting plasma glucose concentrations in ZDF rats, ob/ob mice, and an STZ-treated, high-fat-fed rat model of T2DM. Surprisingly, CREB ASO treatment also decreased plasma cholesterol and triglyceride concentrations, as well as hepatic triglyceride content, due to decreases in hepatic lipogenesis. These results suggest that CREB is an attractive therapeutic target for correcting both hepatic insulin resistance and dyslipidemia associated with nonalcoholic fatty liver disease (NAFLD) and T2DM.