Caffeine Inhibits Both Basal and Insulin-Activated Urate Transport.

OBJECTIVE: Caffeine, an adenosine receptor antagonist, is a potent central nervous system stimulant that also impairs insulin signaling. Recent studies have suggested that coffee consumption lowers serum urate (SU) and protects against gout by unknown mechanisms. We hypothesized that caffeine lowers SU by affecting activity of urate transporters. METHODS: We examined the effect of caffeine and adenosine on basal and insulin stimulation of net 14C-urate uptake in the human renal proximal tubule cell line PTC-05 and on individual urate transporters expressed in Xenopus laevis oocytes. RESULTS: We found that caffeine and adenosine efficiently inhibited both basal and insulin stimulation of net 14C-urate uptake mediated by endogenous urate transporters in PTC-05 cells. In oocytes expressing individual urate transporters, caffeine (>0.2 mM) more efficiently inhibited the basal urate transport activity of GLUT9 isoforms, OAT4, OAT1, OAT3, NPT1, ABCG2, and ABCC4 than did adenosine without significantly affecting URAT1 and OAT10. However, unlike adenosine, caffeine at lower concentrations (<0.2 mM) very effectively inhibited insulin activation of urate transport activity of GLUT9, OAT10, OAT1, OAT3, NPT1, ABCG2, and ABCC4 by blocking activation of Akt and extracellular signal-regulated kinase. CONCLUSION: We postulate that inhibition of urate transport activity of the re-absorptive transporters GLUT9, OAT10, and OAT4 by caffeine is a key mechanism in its urate-lowering effects. Additionally, the ability of caffeine to block insulin-activated urate transport by GLUT9a and OAT10 suggests greater relative inhibition of these transporters in hyperinsulinemia.

Monoethanolamine-induced glucose deprivation promotes apoptosis through metabolic rewiring in prostate cancer.

Rationale: Cancer cells rely on glucose metabolism for fulfilling their high energy demands. We previously reported that monoethanolamine (Etn), an orally deliverable lipid formulation, reduced intracellular glucose and glutamine levels in prostate cancer (PCa). Glucose deprivation upon Etn treatment exacerbated metabolic stress in PCa, thereby enhancing cell death. Moreover, Etn was potent in inhibiting tumor growth in a PCa xenograft model. However, the precise mechanisms underlying Etn-induced metabolic stress in PCa remain elusive. The purpose of the present study was to elucidate the mechanisms contributing to Etn-mediated metabolic rewiring in PCa. Methods: Glucose transporters (GLUTs) facilitate glucose transport across the plasma membrane. Thus, we assessed the expression of GLUTs and the internalization of GLUT1 in PCa. We also evaluated the effects of Etn on membrane dynamics, mitochondrial structure and function, lipid droplet density, autophagy, and apoptosis in PCa cells. Results: Compared to other GLUTs, GLUT1 was highly upregulated in PCa. We observed enhanced GLUT1 internalization, altered membrane dynamics, and perturbed mitochondrial structure and function upon Etn treatment. Etn-induced bioenergetic stress enhanced lipolysis, decreased lipid droplet density, promoted accumulation of autophagosomes, and increased apoptosis. Conclusion: We provide the first evidence that Etn alters GLUT1 trafficking leading to metabolic stress in PCa. By upregulating phosphatidylethanolamine (PE), Etn modulates membrane fluidity and affects mitochondrial structure and function. Etn also induces autophagy in PCa cells, thereby promoting apoptosis. These data strongly suggest that Etn rewires cellular bioenergetics and could serve as a promising anticancer agent for PCa.

Quercetin­conjugated superparamagnetic iron oxide nanoparticles modulate glucose metabolism-related genes and miR-29 family in the hippocampus of diabetic rats.

Quercetin (QC) is a dietary bioflavonoid that can be conjugated with nanoparticles to facilitate its brain bioavailability. We previously showed that quercetin-conjugated superparamagnetic iron oxide nanoparticles (QCSPIONs) reduced the level of blood glucose in diabetic rats. Glucose transporters (GLUTs), insulin-like growth factor-1 (IGF-1), and microRNA-29 (miR-29) play a critical role in brain glucose homeostasis. In the current study, we examined the effects of QCSPION on the expression of glucose metabolism-related genes, and the miR-29 family as a candidate regulator of glucose handling in the hippocampus of diabetic rats. Our in silico analyses introduce the miR-29 family as potential regulators of glucose transporters and IGF-1 genes. The expression level of the miR-29 family, IGF-1, GLUT1, GLUT2, GLUT3, and GLUT4 were measured by qPCR. Our results indicate that diabetes significantly results in upregulation of the miR-29 family and downregulation of the GLUT1, 2, 3, 4, and IGF-1 genes. Interestingly, QCSPIONs reduced miR-29 family expression and subsequently enhanced GLUT1, 2, 3, 4, and IGF-1expression. In conclusion, our findings suggest that QCSPION could regulate the expression of the miR-29 family, which in turn increases the expression of glucose transporters and IGF-1, thereby reducing diabetic complications.

Placental Inositol Reduced in Gestational Diabetes as Glucose Alters Inositol Transporters and IMPA1 Enzyme Expression.

CONTEXT: Perturbed inositol physiology in insulin-resistant conditions has led to proposals of inositol supplementation for gestational diabetes (GDM) prevention, but placental inositol biology is poorly understood. OBJECTIVE: Investigate associations of maternal glycemia with placental inositol content, determine glucose effects on placental expression of inositol enzymes and transporters, and examine relations with birthweight. DESIGN AND PARTICIPANTS: Case-control study of placentae from term singleton pregnancies (GDM n = 24, non-GDM n = 26), and culture of another 9 placentae in different concentrations of glucose and myo-inositol for 48 hours. MAIN OUTCOME MEASURES: Placental inositol was quantified by the Megazyme assay. Relative expression of enzymes involved in myo-inositol metabolism and plasma membrane inositol transport was determined by quantitative RT-PCR and immunoblotting. Linear regression analyses were adjusted for maternal age, body mass index, ethnicity, gestational age, and sex. RESULTS: Placental inositol content was 17% lower in GDM compared with non-GDM. Higher maternal mid-gestation glycemia were associated with lower placental inositol. Increasing fasting glycemia was associated with lower protein levels of the myo-inositol synthesis enzyme, IMPA1, and the inositol transporters, SLC5A11 and SLC2A13, the expression of which also correlated with placental inositol content. In vitro, higher glucose concentrations reduced IMPA1 and SLC5A11 mRNA expression. Increasing fasting glycemia positively associated with customized birthweight percentile as expected in cases with low placental inositol, but this association was attenuated with high placental inositol. CONCLUSION: Glycemia-induced dysregulation of placental inositol synthesis and transport may be implicated in reduced placental inositol content in GDM, and this may in turn be permissive to accelerated fetal growth.

Arsenite-induced transgenerational glycometabolism is associated with up-regulation of H3K4me2 via inhibiting spr-5 in caenorhabditis elegans.

Arsenic (As) is a toxic element that is highly abundant in the environment. However, there has not been sufficient research into the mechanisms of arsenic-induced transgenerational effects. In biomedical and environmental toxicology research field, C. elegans are often used as the ideal model. In this study, F0 generation animals were cultured with arsenite, while subsequent generations animals (F1 - F6) were cultured in the absence of arsenic. Experiments were performed to examine the transgenerational glycometabolism and the associated mechanisms in all seven generations (F0 - F6) of C. elegans. Results show that arsenite exposure increased total glucose content but reduced glucose metabolites in F0 generation C. elegans. The total glucose content was also elevated in subsequent generations probably due to transgenerational downregulation of fgt-1. In addition, arsenite exposure induced transgenerational downregulation of histone demethyltransferase spr-5 and elevated histone dimethylation in F0 generation. This study highlights that single generation exposure to arsenite causes transgenerational changes in glycometabolism in C. elegans, which may be caused by downregulation of spr-5 and elevation of H3K4me2.

Evaluation of the possibility of selective modulation of retinal glucose transporters in diabetic complications: An experimental study.

PURPOSE: To identify the possibility of modulating retinal glucose transporters in diabetic conditions to prevent retinal complications of diabetic retinopathy. MATERIALS AND METHODS: In silico and in vitro binding assays were performed to assess the effect of genistein and positive controls (pioglitazone and estradiol) on nuclear receptor estrogen receptor beta and peroxisome proliferator-activated receptor gamma (PPARγ). In vivo effects of compounds were tested on diabetic rats. Structural and functional analysis of retina was performed at 28th day followed by gene expression analysis of glucose transporters and nuclear receptors. Pioglitazone and genistein levels were analyzed by liquid chromatography with tandem mass spectrometry. RESULTS: Genistein showed equi-affinity toward PPARγ in in silico experiments contrary to in vitro findings. In multidose study, their therapeutic effects were observed by analyzing the retinal function. Retinal gene expression studies revealed that both test agents significantly up regulated PPARγ, GLUT4, and down regulated GLUT1. Genistein showed significant up regulation of GLUT4 and down regulation of GLUT1 as compared to PGZ which has been well correlated with the Electroretinography (ERG) outcome. CONCLUSION: This study showed the possibility of selective upregulation of GLUT4 (independent of PPARγ activation) in the retina of diabetic rats using genistein. Selective modulation of retinal glucose transporters as therapeutic target in ocular diabetic complications can be possibly explored.

Aged garlic extract and S-allylcysteine increase the GLUT3 and GCLC expression levels in cerebral ischemia.

BACKGROUND: During cerebral ischemia, energy restoration through the regulation of glucose transporters and antioxidant defense mechanisms is essential to maintain cell viability. Antioxidant therapy has been considered effective to attenuate brain damage; moreover, the regulation of transcription factors that positively regulate the expression of glucose transporters is associated with this therapy. Recently, it has been reported that the use of antioxidants such as S-allylcysteine (SAC), a component of aged garlic extract (AGE), improves survival in experimental models of cerebral ischemia. OBJECTIVES: The aim of this study was to determine the effect of AGE and SAC on the level of mRNA expression of the main neuronal glucose transporter (GLUT3) and the glutamate cysteine ligase catalytic subunit (GCLC) in rats with transient focal cerebral ischemia. MATERIAL AND METHODS: Cerebral ischemia was induced in male Wistar rats by middle cerebral artery occlusion (MCAO) for 2 h. The animals were sacrificed after different reperfusion times (0-48 h). Animals injected with AGE (360 mg/kg, intraperitoneally (i.p.)) and SAC (300 mg/kg, i.p.) at the beginning of reperfusion were sacrificed after 2 h. The mRNA expression level was analyzed in the fronto-parietal cortex using quantitative polymerase chain reaction (qPCR). RESULTS: Two major increases in GLUT3 expression at 1 h and 24 h of reperfusion were found. Both treatments increased GLUT3 and GCLC mRNA levels in control and under ischemic/reperfusion injury animals. CONCLUSIONS: This data suggests that SAC and AGE might induce neuroprotection, while controlling reactive oxygen species (ROS) levels, as indicated by the increase in GCLC expression, and regulating the energy content of the cell by increasing glucose transport mediated by GLUT3.

Fluoxetine induces glucose uptake and modifies glucose transporter palmitoylation in human peripheral blood mononuclear cells.

Introduction: In line with the monoamine hypothesis of major depressive disorder (MDD), the clinical efficacy of the selective serotonin reuptake inhibitor fluoxetine has classically been ascribed to central serotonin enhancing properties. Current research described disturbances in brain energy metabolism in MDD. Additionally, fluoxetine showed beneficial effects in neuropsychiatric disorders associated with central energy imbalance. Areas covered: The effect of in vitro fluoxetine exposure on cellular glucose uptake and cerebral glucose transporter function was assessed in human peripheral blood mononuclear cells (PBMC) and murine neuroblastoma N2a cells. Fluoxetine augmented glucose uptake, measured by utilizing the radionuclide-labled glucose analog [18]F-fluorodeoxyglucose, in PBMC without affecting glucose transporter protein content. Analysis of protein palmitoylation using the acyl-biotinyl exchange method revealed GLUT3 to be palmitoylated in PBMC and N2a cells, while palmitoylation of GLUT1 was detected only in N2a cells. Treatment with fluoxetine significantly increased palmitoylation of GLUT3 in PBMC and strongly induced palmitoylation of GLUT1 in PBMC and N2a cells. Expert opinion: Our findings suggest a novel mechanism exerted by fluoxetine targeting glucose metabolism by regulating glucose transporter palmitoylation. Thus, fluoxetine might evoke its therapeutic effects in neuropsychiatric diseases characterized by disturbances in central energy metabolism at least partly by improving cerebral glucose uptake.

Discovery of Small-Molecule Selective mTORC1 Inhibitors via Direct Inhibition of Glucose Transporters.

The mechanistic target of rapamycin (mTOR) is a central regulator of cellular metabolic processes. Dysregulation of this kinase complex can result in a variety of human diseases. Rapamycin and its analogs target mTORC1 directly; however, chronic treatment in certain cell types and in vivo results in the inhibition of both mTORC1 and mTORC2. We have developed a high-throughput cell-based screen for the detection of phosphorylated forms of the mTORC1 (4E-BP1, S6K1) and mTORC2 (Akt) substrates and have identified and characterized a chemical scaffold that demonstrates a profile consistent with the selective inhibition of mTORC1. Stable isotope labeling of amino acids in cell culture-based proteomic target identification revealed that class I glucose transporters were the primary target for these compounds yielding potent inhibition of glucose uptake and, as a result, selective inhibition of mTORC1. The link between the glucose uptake and selective mTORC1 inhibition are discussed in the context of a yet-to-be discovered glucose sensor.

Effects of Lactobacillus Plantarum and Lactobacillus Helveticus on Renal Insulin Signaling, Inflammatory Markers, and Glucose Transporters in High-Fructose-Fed Rats.

Background and Objectives: The excess consumption of fructose in the diet may cause metabolic syndrome, which is associated with an increased risk of kidney disease. There is limited data on probiotic treatment in high-fructose-induced metabolic syndrome. The present study aims to investigate whether the supplementation of Lactobacillus plantarum (L. plantarum) and Lactobacillus helveticus (L. helveticus) could provide an improving effect on the renal insulin signaling effectors, inflammatory parameters, and glucose transporters in fructose-fed rats. Materials and Methods: The model of metabolic syndrome in male Wistar rats was produced by fructose, which was given as 20% solution in drinking water for 15 weeks. L. plantarum and L. helveticus supplementations were given by gastric gavage from 10 to 15 weeks of age. Results: High-fructose consumption in rats reduced renal protein expressions of insulin receptor substrate (IRS)-1, protein kinase B (AKT), and endothelial nitric oxide synthase (eNOS), which were improved by L. plantarum and partially by L. helveticus supplementations. Dietary fructose-induced elevations in renal tissue levels of tumor necrosis factor α (TNF-α), interleukin (IL)-1ß, IL-6, and IL-10, as well as expression of IL-6 mRNA, were attenuated, especially in L. plantarum treated rats. The increased renal expression of sodium-glucose cotransporter-2 (SGLT2), but not that of glucose transporter type-5 (GLUT5), was suppressed by the treatment with L. plantarum. Conclusion: Suppression in insulin signaling pathway together with the induction of inflammatory markers and upregulation of SGLT2 in fructose-fed rats were improved by L. plantarum supplementation. These findings may offer a new approach to the management of renal dysregulation induced by dietary high-fructose.

GLUT12 expression and regulation in murine small intestine and human Caco-2 cells.

GLUT12 was cloned from the mammary cancer cell line MCF-7, but its physiological role still needs to be elucidated. To gain more knowledge of GLUT12 function in the intestine, we investigated GLUT12 subcellular localization in the small intestine and its regulation by sugars, hormones, and intracellular mediators in Caco-2 cells and mice. Immunohistochemical methods were used to determine GLUT12 subcellular localization in human and murine small intestine. Brush border membrane vesicles were isolated for western blot analyses. Functional studies were performed in Caco-2 cells by measuring α-methyl-d-glucose (αMG) uptake in the absence of sodium. GLUT12 is located in the apical cytoplasm, below the brush border membrane, and in the perinuclear region of murine and human enterocytes. In Caco-2 cells, GLUT12 translocation to the apical membrane and α-methyl- d-glucose uptake by the transporter are stimulated by protons, glucose, insulin, tumor necrosis factor-α (TNF-α), protein kinase C, and AMP-activated protein kinase. In contrast, hypoxia decreases GLUT12 expression in the apical membrane. Upregulation of TNF-α and hypoxia-inducible factor-1α ( HIF-1α) genes is found in the jejunal mucosa of diet-induced obese mice. In these animals, GLUT12 expression in the brush border membrane is slightly decreased compared with lean animals. Moreover, an intraperitoneal injection of insulin does not induce GLUT12 translocation to the membrane, as it occurs in lean animals. GLUT12 rapid translocation to the enterocytes' apical membrane in response to glucose and insulin could be related to GLUT12 participation in sugar absorption during postprandial periods. In obesity, in which insulin sensitivity is reduced, the contribution of GLUT12 to sugar absorption is affected.

A Study on Neonatal Intake of Oleanolic Acid and Metformin in Rats (Rattus norvegicus) with Metabolic Dysfunction: Implications on Lipid Metabolism and Glucose Transport.

Metabolic syndrome, a cluster of different disorders which include diabetes, obesity and cardiovascular diseases, is a global epidemic that is growing at an alarming rate. The origins of disease can be traced back to early developmental stages of life. This has increased mortalities and continues to reduce life expectancies of individuals across the globe. The aim of this study was to investigate the sub-acute and long term effects of neonatal oral administration of oleanolic acid and metformin on lipids (free fatty acids, FFAs) and genes associated with lipid metabolism and glucose transport using a neonatal rat experimental model. In the first study, seven days old pups were randomly grouped into control-distilled water (DW); oleanolic acid (60 mg/kg), metformin (500 mg/kg), high fructose diet (20% w/v, HF), oleanolic acid (OA) + high fructose diet (OA + HF), and Metformin + high fructose diet (MET + HF) groups. The pups were treated for 7 days, and then terminated on postnatal day (PD) 14. In the second study, rat pups were initially treated similarly to study 1 and weaned onto normal rat chow and plain drinking water on PD 21 till they reached adulthood (PD112). Tissue and blood samples were collected for further analyses. Measurement of the levels of free fatty acids (FFAs) was done using gas chromatography-mass spectrometry. Quantitative polymerase chain reaction (qPCR) was used to analyze the gene expression of glut-4, glut-5, fas, acc-1, nrf-1 and cpt-1 in the skeletal muscle. The results showed that HF accelerated accumulation of saturated FFAs within skeletal muscles. The HF fed neonatal rats had increased stearic acid, which was associated with decreased glucose, suppressed expression of glut-4, glut-5, nrf-1 and cpt-1 genes, and increased expression of acc-1 (p < 0.01) and fas. OA + HF and MET + HF treated groups had increased mono- and polyunsaturated FFAs; oleic, and octadecadienoic acids than the HF group. These unsaturated FFAs were associated with increased glut-4, glut-5 and nrf-1 (p < 0.01) and decreased acc-1 and fas (p < 0.05) in both OA + HF and MET + HF treated groups. CONCLUSIONS: The present study shows that neonatal oral administration of oleanolic acid and metformin potentially protects against the development of fructose-induced metabolic dysfunction in the rats in both short and long time periods.

Adverse placental effects of valproic acid: Studies in perfused human placentas.

OBJECTIVE: In utero exposure to valproic acid (VPA) has been associated with worse pregnancy outcomes compared to all other antiepileptic drugs. We have previously shown that VPA alters the expression of placental transporters for hormones and nutrients in vitro and in pregnant mice. Here, our aim was to characterize the effects of short exposure to VPA on the expression of carriers for compounds essential for fetal development in human placentas ex vivo, under controlled conditions. METHODS: Placentas were obtained from cesarean deliveries of women with no known epilepsy. Cotyledons were cannulated and perfused in the absence or the presence of VPA (42, 83, or 166 µg/mL; n = 6/group) in the maternal perfusate over 180 minutes. A customized gene panel array was used to analyze the expression of carrier genes in the perfused cotyledons. We additionally measured in the perfused placentas folic acid concentrations and histone acetylation. RESULTS: VPA significantly altered the mRNA levels of major carriers for folic acid, glucose, choline, thyroid hormones, and serotonin (P < .05) and reduced placental folate concentrations by 25%-35% (P = .059). The effects were observed at therapeutic concentrations sufficient to enhance placental histone acetylation, and some were concentration-dependent. SIGNIFICANCE: Our results point to the placenta as a novel target of VPA, implying potential involvement of the placenta in VPA's adverse fetal outcomes.

A glucose-targeted mixed micellar formulation outperforms Genexol in breast cancer cells.

Breast cancer represents the top cancer among women, accounting 521.000 deaths per year. Development of targeted nanomedicines to breast cancer tissues represents a milestone to reduce chemotherapy side effects. Taking advantage of the over-expression of glucose (Glu) membrane transporters in breast cancer cells, we aim to expand the potential of a paclitaxel (PTX)-loaded mixed micellar formulation based on polyvinyl caprolactam-polyvinylacetate-polyethylene glycol graft copolymer (Soluplus®) and D-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) by its surface decoration with Glu moieties. The glycopolymer (Soluplus(Glu)) was obtained by microwave-assisted ring opening reaction of δ-gluconolactone initiated by Soluplus®. The glycosylation was confirmed by 1H NMR and by agglutination assays employing Concanavalin A. The hydrodynamic diameter of Soluplus(Glu) micelles was characterized by dynamic light scattering (100.3±3.8nm) as well as the critical micellar concentration value (0.0151% w/v). Then, a mixed micelle formulation employing Soluplus®, Soluplus(Glu) and TPGS (3:1:1wt ratio) loaded with PTX (4mg/mL) was developed as a multifunctional nanocarrier. Its in vitro anticancer performance in MCF-7 (1.6-fold) and MDA-MB-231 (14.1-fold) was significantly enhanced (p<0.05) versus the unique commercially available micellar-based PTX-nanoformulation (Genexol®). Furthermore, the in vitro PTX cellular uptake assays revealed that the drug intracellular/cell content was significantly (p<0.05) higher for the Glu-containing mixed micelles versus Genexol® after 6h of incubation with MCF-7 (30.5-fold) and MDA-MB-231 (5-fold). Overall, results confirmed the potential of our Glu-decorated mixed colloidal formulation as an intelligent nanocarrier for PTX-targeted breast cancer chemotherapy.

Epigenetic regulation of placental glucose transporters mediates maternal cadmium-induced fetal growth restriction.

Cadmium (Cd) is one of the most toxic environmental pollutants that cause fetal malformation and growth restriction. However, the molecular mechanisms underlying maternal Cd toxicity on fetal growth remain largely unknown. Specifically, the role of placental nutrient transporters, including glucose transporters (GLUTs), has been poorly characterized in the etiology of Cd-induced fetal growth restriction (FGR). In the present study, we established a murine model of FGR induced by maternal Cd exposure, and examined the toxic effects of Cd on placental GLUTs. Our results showed that GLUT3 is significantly downregulated in Cd-exposed mouse placentas when compared to the normal ones. Data from bisulfite PCR demonstrated the hypermethylation of the promoter region of GLUT3. However, methylation levels remained unchanged in two major repetitive elements (LINE-1 and IAP) in Cd-exposed placentas. Moreover, DNA methyltransferase (DNMT) 3B and DNMT3L were significantly upregulated in Cd-exposed placentas, and there were no expression changes of DNMT1 and DNMT3A. Collectively, our results suggest that changes in DNMT3B and DNMT3L expressions and site-specific DNA methylation may be involved in the etiology of Cd-induced fetal growth restriction through downregulation of GLUT3.

[Inhibition of Sirolimus on the growth of pancreatic carcinoma and its effect on the expression of glucose transporter and hexokinase â ¡].

OBJECTIVE: To investigate whether Sirolimus could affect the glycolytic catabolism pathways of pancreatic carcinoma through the control of hypoxia induced factor (HIF-1α) to inhibit the growth of tumor, and explore the potential mechanism of targeting the signaling pathways of mTOR for the treatment of pancreatic carcinoma. METHODS: Sirolimus was applied to treat the pancreatic carcinoma in nude mice orthotopic transplantation model, its difference with the control group was compared; RT-PCR and Western blot were used to measure the mRNA and protein expression of mTOR, HIF-1α, Glucose carrier protein 1 (GLUT-1) and Hexokinase Ⅱ (HK-Ⅱ), respectively; the changes of activity of HK-Ⅱ in the tumor was determined. RESULTS: The tumor mass of the control group (1.97±0.21)g was significantly larger than that of the Sirolimus group (0.38±0.10)g (P<0.01), and the volume of the control group (1.40±0.15) mm(3) was significantly larger than that of the Sirolimus group (0.27±0.07) mm(3) (P<0.01). The expressions of mTOR, GLUT-1 and HK-Ⅱ mRNA in the control group were higher than those of the Sirolimus group (P<0.05), while no significant change was observed in the expression of HIF-1α (P>0.05); the expressions of p-mTOR, HIF-1α, GLUT-1 and HK-Ⅱ proteins in the control group were higher than those of the Sirolimus group (P<0.05). The activity of HK-Ⅱ in the control group was higher than that of the Sirolimus group (P<0.05). CONCLUSION: Sirolimus could affect the expression of GLUT-1 and HK-Ⅱ in pancreatic carcinoma through the effects of HIF-1α to inhibit tumor growth, indicating that blocking the mTOR pathway could control the glycolytic metabolism pathways of pancreatic carcinoma, which may become the new strategy for the treatment of pancreatic carcinoma.

Dose-dependent effects of caffeine in human Sertoli cells metabolism and oxidative profile: relevance for male fertility.

Caffeine is a widely consumed substance present in several beverages. There is an increasing consumption of energetic drinks, rich in caffeine, among young individuals in reproductive age. Caffeine has been described as a modulator of cellular metabolism. Hence, we hypothesized that it alters human Sertoli cells (hSCs) metabolism and oxidative profile, which are essential for spermatogenesis. For that purpose, hSCs were cultured with increasing doses of caffeine (5, 50, 500 µM). Caffeine at the lowest concentrations (5 and 50 µM) stimulated lactate production, but only hSCs exposed to 50 µM showed increased expression of glucose transporters (GLUTs). At the highest concentration (500 µM), caffeine stimulated LDH activity to sustain lactate production. Notably, the antioxidant capacity of hSCs decreased in a dose-dependent manner and SCs exposed to 500 µM caffeine presented a pro-oxidant potential, with a concurrent increase of protein oxidative damage. Hence, moderate consumption of caffeine appears to be safe to male reproductive health since it stimulates lactate production by SCs, which can promote germ cells survival. Nevertheless, caution should be taken by heavy consumers of energetic beverages and food supplemented with caffeine to avoid deleterious effects in hSCs functioning and thus, abnormal spermatogenesis.

Cryptotanshinone reverses ovarian insulin resistance in mice through activation of insulin signaling and the regulation of glucose transporters and hormone synthesizing enzymes.

OBJECTIVE: To investigate the effects of cryptotanshinone (CRY), an active component of Chinese medicine, on ovarian androgen production, insulin resistance (IR), and glucose metabolism in mice. DESIGN: Animal model and in vitro tissue model. SETTING: University-affiliated laboratory. ANIMAL(S): Mice. INTERVENTION(S): Ovarian IR was induced by dexamethasone (DEX) in vivo. Animals were randomized to receive CRY treatment for 3 days or not. Ovulation rates, serum steroid levels, and glucose uptake in ovaries were quantified, and proteins in the phosphatidylinositol 3-hydroxy kinase pathway were measured. In vitro ovarian IR was also induced by DEX for 3 days. Ovarian steroid hormone secretion and glucose uptake were measured, and the hormone-synthesizing enzymes were determined by semiquantitative reverse transcription-polymerase chain reaction. MAIN OUTCOME MEASURE(S): Ovarian glucose uptake, in vivo ovulation rate, serum and culture medium steroid level, and molecular expression of phosphatidylinositol 3-hydroxy kinase and steroidogenic enzymes. RESULT(S): Dexamethasone significantly increased ovulation rates in vivo and increased T and E2 production and decreased ovarian glucose uptake in vivo and in vitro. Cryptotanshinone significantly reduced ovulation rates in vivo and decreased T and estrogen production in vitro. Cryptotanshinone attenuated the inhibition of DEX on AKT2 and suppressed the up-regulation of CYP11 and CYP17 expression by DEX. CONCLUSION(S): Cryptotanshinone reversed DEX-induced androgen excess and ovarian IR in mice through activation of insulin signaling and the regulation of glucose transporters and hormone-synthesizing enzymes. This suggests a potential role for CRY in treating the ovulatory dysfunction associated with PCOS.

The essential role of endothelial nitric oxide synthase activation in insulin-mediated neuroprotection against ischemic stroke in diabetes.

BACKGROUND: Stroke patients with diabetes have a higher mortality rate, worse neurologic outcome, and more severe disability than those without diabetes. Results from clinical trials comparing the outcomes of stroke seen with intensive glycemic control in diabetic individuals are conflicting. Therefore, the present study was aimed to identify the key factor involved in the neuroprotective action of insulin beyond its hypoglycemic effects in streptozotocin-diabetic rats with ischemic stroke. METHODS: Long-Evans male rats were divided into three groups (control, diabetes, and diabetes treated with insulin) and subjected to focal cerebral ischemia-reperfusion (FC I/R) injury. RESULTS: Hyperglycemia aggravated FC I/R injuries with an increase in cerebral infarction and neurologic deficits, inhibition of glucose uptake and membrane-trafficking activity of glucose transporter 1, and reduction of Akt and endothelial nitric oxide synthase (eNOS) phosphorylation in the cerebrum. Insulin treatment alleviated hyperglycemia and the symptoms of diabetes in streptozotocin-diabetic rats. Insulin administration also significantly decreased cerebral infarction and neurologic deficits and increased phosphorylation of Akt and eNOS protein in the cerebrum of FC I/R-injured diabetic rats. However, the glucose uptake and membrane trafficking activity of glucose transporter 1 in the cerebrum were not restored by insulin treatment. Coadministration of the eNOS inhibitor, N-iminoethyl-L-ornithine, with insulin abrogated beneficial effects of insulin on cerebral infarct volume and neurologic deficits in FC I/R-injured diabetic rats without affecting the hypoglycemic action of insulin. CONCLUSIONS: These results suggest that eNOS activation is required for the neuroprotection of insulin against ischemic stroke in patients with diabetes.

Riluzole increases the rate of glucose transport in L6 myotubes and NSC-34 motor neuron-like cells via AMPK pathway activation.

Riluzole is the only approved ALS drug. Riluzole influences several cellular pathways, but its exact mechanism of action remains unclear. Our goal was to study the drug's influence on the glucose transport rate in two ALS relevant cell types, neurons and myotubes. Stably transfected wild-type or mutant G93A human SOD1 NSC-34 motor neuron-like cells and rat L6 myotubes were exposed to riluzole. The rate of glucose uptake, translocation of glucose transporters to the cell's plasma membrane and the main glucose transport regulatory proteins' phosphorylation levels were measured. We found that riluzole increases the glucose transport rate and up-regulates the translocation of glucose transporters to plasma membrane in both types of cells. Riluzole leads to AMPK phosphorylation and to the phosphorylation of its downstream target, AS-160. In conclusion, increasing the glucose transport rate in ALS affected cells might be one of the mechanisms of riluzole's therapeutic effect. These findings can be used to rationally design and synthesize novel anti-ALS drugs that modulate glucose transport in neurons and skeletal muscles.