Membrane GLUT4 Research Articles

Elmo2 Is a Regulator of Insulin-dependent Glut4 Membrane Translocation

Elmo2, a member of the Elmo protein family, has been implicated in the regulation of Rac1 and Akt activation. Recently, we found that Elmo2 specifically interacts with ClipR-59. Because Akt and Rac1 have been implicated in insulin dependent Glut4 membrane translocation, we hypothesize here that Elmo2 may play a role in insulin-dependent Glut4 membrane translocation. Accordingly, we found that overexpression of Elmo2 enhanced, whereas its knockdown suppressed, insulin-dependent Glut4 membrane translocation in both 3T3-L1 adipocytes and L6 skeletal muscle cells. We also examined whether Elmo2 contributes to the insulin-mediated activation of Rac1 and Akt. We found that Elmo2 is required for insulin-induced Rac1 GTP loading, but not AKT activation, in L6 cells induced by insulin. Instead, Elmo2 is required to promote the insulin-induced membrane association of Akt. Together, our studies demonstrate that Elmo2 is a new regulator of insulin-dependent Glut4 membrane translocation through modulating Rac1 activity and Akt membrane compartmentalization.

Abstract 175: High Glucose Pre-treatment Suppresses Rankl-induced Macrophage Activation via Down-regulation of Glucose Uptake Through Decreased Insulin Receptor and Insulin Receptor Substrate 1 Expression

Background: Previous studies have suggested that the pathogenesis of abdominal aortic aneurysm (AAA) is associated with the local proteinases activation and the degradation of matrix proteins by matrix metalloproteinases (MMPs) produced from activated macrophages. One of the major features of diabetes mellitus (DM)-induced vascular pathology is severe arterial calcification. Although recent large epidemiological studies have shown that DM is an independent negative risk factor for AAA, the effects of hyperglycemia on macrophages are still controversial. We have hypothesized that hyperglycemia suppresses macrophage activation through altered glucose transportation. Methods and Results: RAW264.7 cells, a murine macrophage cell line were cultured under high glucose conditions (HG group, 15.5 mM glucose) or normal glucose conditions (NG group, 5.5 mM glucose) for 7days. Cells from both groups were then transferred to normal glucose condition and stimulated with recombinant murine sRANKL. Macrophage activation, confirmed by TRAP staining positive cells, and MMP-9 expression were induced in NG group but were significantly suppressed in HG group. Glucose uptake was increased during osteoclastogenesis in NG group but not in HG group. To elucidate the underlying mechanism for this observation, we studied glucose transporters (GLUTs). Although GLUT-1 and GLUT-3 expression were not affected in either groups, the membrane translocation of GLUT-1 was significantly increased in NG group during macrophage activation but not in HG group. Insulin receptor and insulin receptor substrate-1 (IRS-1) mRNA, known to stimulate membrane translocation of GLUT, were both decreased in HG group compared to NG group. Conclusions: Our results showed hyperglycemia suppresses macrophage activation. Our results also indicated that under normal conditions, recombinant murine sRANKL increases glucose uptake during macrophage activation. In contrast, this increase is impaired in high glucose pre-treated cells. We conclude that this impairment is due, in part, to suppressed GLUT-1 membrane translocation through down regulation of insulin receptor and IRS-1

Weekly enzyme replacement therapy: a French patient with infantile onset Pompe disease

Perineuriomas are rare peripheral nerve sheath tumors arising from or differentiating along the lines of normal perineurial cells. They can be divided into intraneural and soft tissue types, with the latter category including a significant number of morphological variants. Herein, we further expand their morphological spectrum to include “pseudolipoblastic” perineuriomas. These lesions occurred in the tongue of a 30-year-old man and in the triceps of a 67-year-old woman and were characterized by bland, epithelioid cells with striking intracytoplasmic vacuolization. The architecture varied, with some areas showing a striking “net-like” or “microreticular” pattern and smaller areas having a more typical spindled and whorled appearance. Clinical follow-up (5 months and 52 months, respectively) showed no evidence of local recurrence or metastasis. Multiple perineurial markers, including epithelial membrane antigen, claudin-1, GLUT-1, and collagen IV, were diffusely positive. Both cases were submitted in consultation out of concern that they represented high-grade liposarcomas. To the best of our knowledge, this unusual morphological variant of perineurioma has not been reported. These tumors appear to be entirely benign and should be cured with simple excision. Pseudolipoblastic perineuriomas should be distinguished from round cell and epithelioid pleomorphic liposarcomas, as well as from other tumors that may show prominent intracytoplasmic vacuolization.

Evaluation of fish oil-rich in MUFAs for anti-diabetic and anti-inflammation potential in experimental type 2 diabetic rats.

The advantages of monounsaturated fatty acids (MUFAs) on insulin resistance and type 2 diabetes mellitus (T2DM) have been well established. However, the molecular mechanisms of the anti-diabetic action of MUFAs remain unclear. This study examined the anti-hyperglycemic effect and explored the molecular mechanisms involved in the actions of fish oil- rich in MUFAs that had been acquired from hybrid catfish (Pangasius larnaudii×Pangasianodon hypophthalmus) among experimental type 2 diabetic rats. Diabetic rats that were fed with fish oil (500 and 1,000 mg/kg BW) for 12 weeks significantly reduced the fasting plasma glucose levels without increasing the plasma insulin levels. The diminishing levels of plasma lipids and the muscle triglyceride accumulation as well as the plasma leptin levels were identified in T2DM rats, which had been administrated with fish oil. Notably, the plasma adiponectin levels increased among these rats. The fish oil supplementation also improved glucose tolerance, insulin sensitivity and pancreatic histological changes. Moreover, the supplementation of fish oil improved insulin signaling (p-AktSer473 and p-PKC-ζ/λThr410/403), p-AMPKThr172 and membrane GLUT4 protein expressions, whereas the protein expressions of pro-inflammatory cytokines (TNF-α and nuclear NF-κB) as well as p-PKC-θThr538 were down regulated in the skeletal muscle. These data indicate that the effects of fish oil-rich in MUFAs in these T2DM rats were partly due to the attenuation of insulin resistance and an improvement in the adipokine imbalance. The mechanisms of the anti-hyperglycemic effect are involved in the improvement of insulin signaling, AMPK activation, GLUT4 translocation and suppression of pro-inflammatory cytokine protein expressions.

Nanomolar Caffeic Acid Decreases Glucose Uptake and the Effects of High Glucose in Endothelial Cells

Epidemiological studies suggest that moderate and prolonged consumption of coffee is associated with a reduced risk of developing type 2 diabetes but the molecular mechanisms underlying this effect are not known. In this study, we report the effects of physiological concentrations of caffeic acid, easily achievable by normal dietary habits, in endothelial cells cultured in 25 mM of glucose (high glucose, HG). In HG, the presence of 10 nM caffeic acid was associated with a decrease of glucose uptake but not to changes of GLUT-1 membrane localization or mRNA levels. Moreover, caffeic acid countered HG-induced loss of barrier integrity, reducing actin rearrangement and FITC-dextran passage. The decreased flux of glucose associated to caffeic acid affected HG induced apoptosis by down-regulating the expression of initiator (caspase 8 and 9) and effector caspases (caspase 7 and 3) and by increasing the levels of phosphorylated Bcl-2. We also observed that caffeic acid in HG condition was associated to a reduction of p65 subunit nuclear levels with respect to HG alone. NF-κB activation has been shown to lead to apoptosis in HG treated cells and the analysis of the expression of a panel of about 90 genes related to NF-κB signaling pathway revealed that caffeic acid significantly influenced gene expression changes induced by HG. In conclusion, our results suggest that caffeic acid, decreasing the metabolic stress induced by HG, allows the activation of survival mechanisms mediated by a different modulation of NF-κB-related signaling pathways and to the activation of anti-apoptotic proteins.

Loss of Toll-Like Receptor 4 Function Partially Protects against Peripheral and Cardiac Glucose Metabolic Derangements During a Long-Term High-Fat Diet

Diabetes is a chronic inflammatory disease that carries a high risk of cardiovascular disease. However, the pathophysiological link between these disorders is not well known. We hypothesize that TLR4 signaling mediates high fat diet (HFD)-induced peripheral and cardiac glucose metabolic derangements. Mice with a loss-of-function mutation in TLR4 (C3H/HeJ) and age-matched control (C57BL/6) mice were fed either a high-fat diet or normal diet for 16 weeks. Glucose tolerance and plasma insulin were measured. Protein expression of glucose transporters (GLUT), AKT (phosphorylated and total), and proinflammatory cytokines (IL-6, TNF-α and SOCS-3) were quantified in the heart using Western Blotting. Both groups fed a long-term HFD had increased body weight, blood glucose and insulin levels, as well as impaired glucose tolerance compared to mice fed a normal diet. TLR4-mutant mice were partially protected against long-term HFD-induced insulin resistance. In control mice, feeding a HFD decreased cardiac crude membrane GLUT4 protein content, which was partially rescued in TLR4-mutant mice. TLR4-mutant mice fed a HFD also had increased expression of GLUT8, a novel isoform, compared to mice fed a normal diet. GLUT8 content was positively correlated with SOCS-3 and IL-6 expression in the heart. No significant differences in cytokine expression were observed between groups, suggesting a lack of inflammation in the heart following a HFD. Loss of TLR4 function partially restored a healthy metabolic phenotype, suggesting that TLR4 signaling is a key mechanism in HFD-induced peripheral and cardiac insulin resistance. Our data further suggest that TLR4 exerts its detrimental metabolic effects in the myocardium through a cytokine-independent pathway.

α1A-Adrenergic receptor prevents cardiac ischemic damage through PKCδ/GLUT1/4-mediated glucose uptake

While α1-adrenergic receptors (ARs) have been previously shown to limit ischemic cardiac damage, the mechanisms remain unclear. Most previous studies utilized low oxygen conditions in addition to ischemic buffers with glucose deficiencies, but we discovered profound differences if the two conditions are separated. We assessed both mouse neonatal and adult myocytes and HL-1 cells in a series of assays assessing ischemic damage under hypoxic or low glucose conditions. We found that α1-AR stimulation protected against increased lactate dehydrogenase release or Annexin V+ apoptosis under conditions that were due to low glucose concentration not to hypoxia. The α1-AR antagonist prazosin or nonselective protein kinase C (PKC) inhibitors blocked the protective effect. α1-AR stimulation increased 3H-deoxyglucose uptake that was blocked with either an inhibitor to glucose transporter 1 or 4 (GLUT1 or GLUT4) or small interfering RNA (siRNA) against PKCδ. GLUT1/4 inhibition also blocked α1-AR-mediated protection from apoptosis. The PKC inhibitor rottlerin or siRNA against PKCδ blocked α1-AR stimulated GLUT1 or GLUT4 plasma membrane translocation. α1-AR stimulation increased plasma membrane concentration of either GLUT1 or GLUT4 in a time-dependent fashion. Transgenic mice overexpressing the α1A-AR but not α1B-AR mice displayed increased glucose uptake and increased GLUT1 and GLUT4 plasma membrane translocation in the adult heart while α1A-AR but not α1B-AR knockout mice displayed lowered glucose uptake and GLUT translocation. Our results suggest that α1-AR activation is anti-apoptotic and protective during cardiac ischemia due to glucose deprivation and not hypoxia by enhancing glucose uptake into the heart via PKCδ-mediated GLUT translocation that may be specific to the α1A-AR subtype.

Extracellular polysaccharide from Bordetella species reduces high glucose-induced macrophage apoptosis via regulating interaction between caveolin-1 and TLR4

Microphage apoptosis is a critical event in atherosclerotic lesions in patients with diabetes. In the present investigation, high glucose treatment inhibited Akt phosphorylation and activated caspase 3 in primary peritoneal macrophage, leading to cell apoptosis. Hypoxia prolonged macrophage survival in high glucose condition. Extracellular polysaccharide from Bordetella species (EPS) further decreased cell apoptosis in response to high glucose during hypoxia. Under high glucose and hypoxic condition, EPS treatment promoted caveolin-1 phosphorylation by recognizing TLR4. Caveolin-1 phosphorylation elevated membrane Glut1 level to accelerate glucose consumption, which should be the reason for protective effect of EPS on macrophage exposed to high glucose. Further investigation demonstrated that TLR4-dependent caveolin-1 phosphorylation induced by EPS promoted association of caveolin-1 with TLR4, which should be critical for activation of TLR4 signaling pathway.

Advanced glycation end-products and insulin signaling in granulosa cells.

Advanced glycation end-products (AGEs) may interfere with insulin intracellular signaling and glucose transport in human granulosa cells, potentially affecting ovarian function, follicular growth, linked with diminished fertility. The potential interaction of AGEs with insulin signaling pathways and glucose transport was investigated in human granulosa KGN cells. KGN cells were cultured with variable concentrations of human glycated albumin (HGA, 50-200 µg/mL) or insulin (100 ng/mL). Combined treatments of KGN cells with insulin (100 ng/mL) and HGA (200 µg/mL) were also performed. p-AKT levels and glucose transporter type 4 (Glut-4) translocation analysis were performed by Western blot. Phosphatidylinositol-3-kinase (PI3K)-specific signaling was checked by using the PI3K-inhibitor, LY294002. p-AKT levels were significantly increased following insulin treatment compared to basal levels or HGA exposure. This insulin-mediated AKT-phosphorylation was PI3K-specific and it was inhibited after combined treatment of insulin and HGA. Furthermore, Glut-4 translocation from the cytoplasm to the membrane compartments of KGN cells was remarkably reduced after the combined treatment of insulin and HGA. The present findings support that AGEs interfere with insulin signaling in granulosa cells and prevent Glut-4 membrane translocation suggesting that intra ovarian AGEs accumulation, from endogenous or exogenous sources, may contribute to the pathophysiology of states characterized with anovulation and insulin resistance such as polycystic ovary syndrome.

Differential Role of Insulin/IGF-1 Receptor Signaling in Muscle Growth and Glucose Homeostasis.

Insulin and insulin-like growth factor 1 (IGF-1) are major regulators of muscle protein and glucose homeostasis. To determine how these pathways interact, wegenerated mice with muscle-specific knockout of IGF-1 receptor (IGF1R) and insulin receptor (IR). These MIGIRKO mice showed >60% decrease in muscle mass. Despite a complete lack of insulin/IGF-1 signaling in muscle, MIGIRKO mice displayed normal glucose and insulin tolerance. Indeed, MIGIRKO mice showed fasting hypoglycemia and increased basal glucose uptake. This was secondary to decreased TBC1D1 resulting in increased Glut4 and Glut1 membrane localization. Interestingly, overexpression of a dominant-negative IGF1R in muscle induced glucose intolerance in MIGIRKO animals. Thus, loss of insulin/IGF-1 signaling impairs muscle growth, but not whole-body glucose tolerance due to increased membrane localization of glucose transporters. Nonetheless, presence of a dominant-negative receptor, even in the absence of functional IR/IGF1R, induces glucose intolerance, indicating that interactions between these receptors and other proteins in muscle can impair glucose homeostasis.

Glut4 palmitoylation at Cys223 plays a critical role in Glut4 membrane trafficking

Recently, we identified Glut4 as a palmitoylated protein in adipocytes. To understand the role of Glut4 palmitoylation in Glut4 membrane trafficking, a process that is essential for maintenance of whole body glucose homeostasis, we have characterized Glut4 palmitoylation. We found that Glut4 is palmitoylated at Cys223 and Glut4 palmitoylation at Cys223 is essential for insulin dependent Glut4 membrane translocation as substitution of Cys223 with a serine residue in Glut4 (C223S Glut4) diminished Glut4 responsiveness to insulin in membrane translocation in both adipocytes and CHO-IR cells. We have examined C223S Glut4 subcellular localization and observed that it was absence from tubular-vesicle structure, where insulin responsive Glut4 vesicles were presented. Together, our studies uncover a novel mechanism under which Glut4 palmitoylation regulates Glut4 sorting to insulin responsive vesicles, thereby insulin-dependent Glut4 membrane translocation.

Transmembrane tumor necrosis factor-alpha sensitizes adipocytes to insulin

Transmembrane TNF-α (tmTNF-α) acts both as a ligand, delivering ‘forward signaling’ via TNFR, and as a receptor, transducing ‘reverse signaling’. The contradiction of available data regarding the effect of tmTNF-α on insulin resistance may be due to imbalance in both signals. Here, we demonstrated that high glucose-induced impairment of insulin-stimulated glucose uptake by 3T3-L1 adipocytes was concomitant with decreased tmTNF-α expression and increased soluble TNF-α (sTNF-α) secretion. However, when TACE was inhibited, preventing the conversion of tmTNF-α to sTNF-α, this insulin resistance was partially reversed, indicating a salutary role of tmTNF-α. Treatment of 3T3-L1 adipocytes with exogenous tmTNF-α promoted insulin-induced phosphorylation of IRS-1 and Akt, facilitated GLUT4 expression and membrane translocation, and increased glucose uptake while addition of sTNF-α resulted in the opposite effect. Furthermore, tmTNF-α downregulated the production of IL-6 and MCP-1 via NF-κB inactivation, as silencing of A20, an inhibitor for NF-κB, by siRNA, abolished this effect of tmTNF-α. However, tmTNF-α upregulated adiponectin expression through the PPAR-γ pathway, as inhibition of PPAR-γ by GW9662 abrogated both tmTNF-α-induced adiponectin transcription and glucose uptake. Our data suggest that tmTNF-α functions as an insulin sensitizer via forward signaling.

Tropomodulin3 is a novel Akt2 effector regulating insulin-stimulated GLUT4 exocytosis through cortical actin remodeling.

Akt2 and its downstream effectors mediate insulin-stimulated GLUT4-storage vesicle (GSV) translocation and fusion with the plasma membrane (PM). Using mass spectrometry, we identify actin-capping protein Tropomodulin 3 (Tmod3) as an Akt2-interacting partner in 3T3-L1 adipocytes. We demonstrate that Tmod3 is phosphorylated at Ser71 on insulin-stimulated Akt2 activation, and Ser71 phosphorylation is required for insulin-stimulated GLUT4 PM insertion and glucose uptake. Phosphorylated Tmod3 regulates insulin-induced actin remodelling, an essential step for GSV fusion with the PM. Furthermore, the interaction of Tmod3 with its cognate tropomyosin partner, Tm5NM1 is necessary for GSV exocytosis and glucose uptake. Together these results establish Tmod3 as a novel Akt2 effector that mediates insulin-induced cortical actin remodelling and subsequent GLUT4 membrane insertion. Our findings suggest that defects in cytoskeletal remodelling may contribute to impaired GLUT4 exocytosis and glucose uptake.

Dengue Virus Serotype-2 Interferes with the Formation of Neutrophil Extracellular Traps

Objectives: Neutrophils play an important role in the control of pathogens through several mechanisms, including phagocytosis and the formation of neutrophil extracellular traps (NETs). The latter consists of DNA as a backbone with embedded antimicrobial peptides, histones, and proteases, providing a matrix to entrap and in some cases to kill microbes. Some metabolic requirements for NET formation have recently been described. The virus-induced formation of NETs and the role of these traps in viral infections remain scarcely reported. Here, we analyzed whether dengue virus serotype-2 (DENV-2) induces NET formation and the DENV-2 effect on phorbol myristate acetate (PMA)-induced NETs. Methods: Peripheral blood-derived neutrophils were exposed in vitro to DENV-2 or exposed to DENV-2 and then stimulated with PMA. NET formation was assessed by fluorescence microscopy. Cell membrane Glut-1, glucose uptake, and reactive oxygen species (ROS) production were assessed. Results: DENV-2 does not induce the formation of NETs. Moreover, DENV-2 inhibits PMA-induced formation of NETs by about 80%. This effect is not related to the production of ROS. The mechanism seemingly accountable for this inhibitory effect is the DENV-2-mediated inhibition of PMA-induced glucose uptake by neutrophils. Conclusion: Our results suggest that DENV-2 inhibits glucose uptake as a metabolism-based way to avoid the formation of NETs.

The cholesterol-lowering agent methyl-β-cyclodextrin promotes glucose uptake via GLUT4 in adult muscle fibers and reduces insulin resistance in obese mice.

Insulin stimulates glucose uptake in adult skeletal muscle by promoting the translocation of GLUT4 glucose transporters to the transverse tubule (T-tubule) membranes, which have particularly high cholesterol levels. We investigated whether T-tubule cholesterol content affects insulin-induced glucose transport. Feeding mice a high-fat diet (HFD) for 8 wk increased by 30% the T-tubule cholesterol content of triad-enriched vesicular fractions from muscle tissue compared with triads from control mice. Additionally, isolated muscle fibers (flexor digitorum brevis) from HFD-fed mice showed a 40% decrease in insulin-stimulated glucose uptake rates compared with fibers from control mice. In HFD-fed mice, four subcutaneous injections of MβCD, an agent reported to extract membrane cholesterol, improved their defective glucose tolerance test and normalized their high fasting glucose levels. The preincubation of isolated muscle fibers with relatively low concentrations of MβCD increased both basal and insulin-induced glucose uptake in fibers from controls or HFD-fed mice and decreased Akt phosphorylation without altering AMPK-mediated signaling. In fibers from HFD-fed mice, MβCD improved insulin sensitivity even after Akt or CaMK II inhibition and increased membrane GLUT4 content. Indinavir, a GLUT4 antagonist, prevented the stimulatory effects of MβCD on glucose uptake. Addition of MβCD elicited ryanodine receptor-mediated calcium signals in isolated fibers, which were essential for glucose uptake. Our findings suggest that T-tubule cholesterol content exerts a critical regulatory role on insulin-stimulated GLUT4 translocation and glucose transport and that partial cholesterol removal from muscle fibers may represent a useful strategy to counteract insulin resistance.

Exercise ameliorates insulin resistance via Ca2+ signals distinct from those of insulin for GLUT4 translocation in skeletal muscles.

Muscle contraction and insulin induce glucose uptake in skeletal muscle through GLUT4 membrane translocation. Beneficial effects of exercise on glucose homeostasis in insulin-resistant individuals are known to be due to their distinct mechanism between contraction and insulin action on glucose uptake in skeletal muscle. However, the underlying mechanisms are not clear. Here we show that in skeletal muscle, distinct Ca(2+) second messengers regulate GLUT4 translocation by contraction and insulin treatment; d-myo-inositol 1,4,5-trisphosphate/nicotinic acid adenine dinucleotide phosphate (NAADP) and cyclic ADP-ribose/NAADP are main players for insulin- and contraction-induced glucose uptake, respectively. Different patterns of phosphorylation of AMPK and Ca(2+)/calmodulin-dependent protein kinase II were shown in electrical stimuli (ES)- and insulin-induced glucose uptake pathways. ES-induced Ca(2+) signals and glucose uptake are dependent on glycolysis, which influences formation of NAD(P)-derived signaling messengers, whereas insulin-induced signals are not. High-fat diet (HFD) induced a defect in only insulin-mediated, but not ES-mediated, Ca(2+) signaling for glucose uptake, which is related to a specifically lower NAADP formation. Exercise decreases blood glucose levels in HFD-induced insulin resistance mice via NAADP formation. Thus we conclude that different usage of Ca(2+) signaling in contraction/insulin-stimulated glucose uptake in skeletal muscle may account for the mechanism by which exercise ameliorates glucose homeostasis in individuals with type 2 diabetes.

17-β estradiol attenuates ovariectomy-induced changes in cardiomyocyte contractile function via activation of AMP-activated protein kinase

Menopause increases the risk of cardiometabolic diseases in women. This circumstance is usually attributed to a deficiency in circulating estrogen levels although the underlying mechanism remains elusive. Given the pivotal role of AMP-activated protein kinase (AMPK) in the regulation of energy metabolism and cardiac function, this study was designed to examine the role of AMPK in estrogen deficiency and replacement-exerted cardiomyocyte responses. Adult female WT and AMPK kinase dead (KD) mice were subjected to bilateral ovariectomy (OVX) or sham operation. A cohort of ovariectomized mice received 17β-estradiol (E2) (40μg/kg/day, i.p.) for 6 weeks. Mechanical and intracellular Ca2+ properties were evaluated including peak shortening (PS), time-to-PS (TPS), time-to-90%-relengthening (TR90), and maximal velocity of shortening/relengthening (±dL/dt). Levels of AMPK, Akt JNK, ACC, SERCA, membrane Glut4, AS160 and PGC-1α were assessed using Western blot. OVX significantly decreased PS, ±dL/dt and intracellular Ca2+ rise in responsible to electric stimulus, prolonged TR90 and intracellular Ca2+ decay without affecting TPS and resting intracellular Ca2+, the effects of which were reconciled by E2 replacement. Western blot analysis depicted that OVX suppressed phosphorylation of Akt AMPK and ACC although it promoted JNK phosphorylation, the effects of which were mitigated or significantly attenuated by E2 treatment in WT but not KD mice. Moreover, OVX procedure downregulated SERCA2a and membrane Glut4 while inhibiting AS160 phosphorylation without affecting PGC-1α levels. In vitro study revealed that E2 corrected cardiomyocyte contractile dysfunction elicited by OVX in cardiomyocytes from WT but not the AMPK kinase dead mice. Taken together, these data suggest that E2 treatment ameliorates estrogen deficiency-induced changes in cardiac contractile function possibly through an AMPK-dependent mechanism.

Chromium chloride increases insulin-stimulated glucose uptake in the perfused rat hindlimb.

To determine the effect of chromium chloride (CrCl3 ) on healthy skeletal muscle glucose uptake in the absence and presence of submaximal insulin using the rat hindlimb perfusion technique. Sprague-Dawley rats were randomly assigned to an experimental group: basal (Bas), chromium chloride (Cr), submaximal insulin (sIns) or chromium chloride plus submaximal insulin (Cr-sIns). Insulin significantly increased [H(3)]-2 deoxyglucose (2-DG) uptake in the gastrocnemius muscles. Additionally, Cr-sIns displayed greater rates of 2-DG uptake than sIns (Cr-sIns 6.86 ± 0.74 μmol g h(-1) vs. sIns 4.83 ± 0.42 μmol g h(-1)). There was no difference between Cr and Bas treatment groups. It has been speculated that chromium works to increase glucose uptake by increasing insulin signalling. We found that Akt and AS160 phosphorylation was increased in the sINS treatment group, while chromium treatment had no additional effect on Akt or AS160 phosphorylation in the absence or presence of insulin. Cr-sIns significantly increased plasma membrane GLUT4 concentration above that of sIns (Cr-sIns 72.22 ± 12.7%, sIns 53.4 ± 6.1%), but in the absence of insulin, chromium had no effect. Exposure of healthy skeletal muscle to chromium may increase skeletal muscle insulin-stimulated GLUT4 translocation and glucose uptake. However, these effects do not appear to result from enhanced insulin signalling proximal to AS160.

Regulation of human trophoblast GLUT1 glucose transporter by insulin-like growth factor I (IGF-I).

Glucose transport to the fetus across the placenta takes place via glucose transporters in the opposing faces of the barrier layer, the microvillous and basal membranes of the syncytiotrophoblast. While basal membrane content of the GLUT1 glucose transporter appears to be the rate-limiting step in transplacental transport, the factors regulating transporter expression and activity are largely unknown. In view of the many studies showing an association between IGF-I and fetal growth, we investigated the effects of IGF-I on placental glucose transport and GLUT1 transporter expression. Treatment of BeWo choriocarcinoma cells with IGF-I increased cellular GLUT1 protein. There was increased basolateral (but not microvillous) uptake of glucose and increased transepithelial transport of glucose across the BeWo monolayer. Primary syncytial cells treated with IGF-I also demonstrated an increase in GLUT1 protein. Term placental explants treated with IGF-I showed an increase in syncytial basal membrane GLUT1 but microvillous membrane GLUT1 was not affected. The placental dual perfusion model was used to assess the effects of fetally perfused IGF-I on transplacental glucose transport and syncytial GLUT1 content. In control perfusions there was a decrease in transplacental glucose transport over the course of the perfusion, whereas in tissues perfused with IGF-I through the fetal circulation there was no change. Syncytial basal membranes from IGF-I perfused tissues showed an increase in GLUT1 content. These results demonstrate that IGF-I, whether acting via microvillous or basal membrane receptors, increases the basal membrane content of GLUT1 and up-regulates basal membrane transport of glucose, leading to increased transepithelial glucose transport. These observations provide a partial explanation for the mechanism by which IGF-I controls nutrient supply in the regulation of fetal growth.

Regulation of insulin resistance and adiponectin signaling in adipose tissue by liver X receptor activation highlights a cross-talk with PPARγ.

Liver X receptors (LXRs) have been recognized as a promising therapeutic target for atherosclerosis; however, their role in insulin sensitivity is controversial. Adiponectin plays a unique role in maintaining insulin sensitivity. Currently, no systematic experiments elucidating the role of LXR activation in insulin function based on adiponectin signaling have been reported. Here, we investigated the role of LXR activation in insulin resistance based on adiponectin signaling, and possible mechanisms. C57BL/6 mice maintained on a regular chow received the LXR agonist, T0901317 (30 mg/kg.d) for 3 weeks by intraperitoneal injection, and differentiated 3T3-L1 adipocytes were treated with T0901317 or GW3965. T0901317 treatment induced significant insulin resistance in C57BL/6 mice. It decreased adiponectin gene transcription in epididymal fat, as well as serum adiponectin levels. Activity of AMPK, a key mediator of adiponectin signaling, was also decreased, resulting in decreased Glut-4 membrane translocation in epididymal fat. In contrast, adiponectin activity was not changed in the liver of T0901317 treated mice. In vitro, both T0901317 and GW3965 decreased adiponectin expression in adipocytes in a dose-dependent manner, an effect which was diminished by LXRα silencing. ChIP-qPCR studies demonstrated that T0901317 decreased the binding of PPARγ to the PPAR-responsive element (PPRE) of the adiponectin promoter in a dose-dependent manner. Furthermore, T0901317 exerted an antagonistic effect on the expression of adiponectin in adipocytes co-treated with 3 µM Pioglitazone. In luciferase reporter gene assays, T0901317 dose-dependently inhibited PPRE-Luc activity in HEK293 cells co-transfected with LXRα and PPARγ. These results suggest that LXR activation induces insulin resistance with decreased adiponectin signaling in epididymal fat, probably due to negative regulation of PPARγ signaling. These findings indicate that the potential of LXR activation as a therapeutic target for atherosclerosis may be limited by the possibility of exacerbating insulin resistance-related disease.