Expression Levels Of Glucose Transporter Research Articles

Abstract 448: Alterations in glycolytic pathway enzymes and metabolic plasticity in prostate cancer progression

Abstract Background: Although oxidative phosphorylation (OXPHOS) is the main ATP source in primary prostate cancer, as the tumor progresses to castration resistance and/or metastatic disease, glycolysis becomes increasingly prevalent. Dependence on glycolysis can further increase under hypoxic conditions found in advanced cancers. We previously demonstrated that a novel small molecule, BKIDC-1553, inhibits proliferation and tumor growth of prostate cancer cell lines and xenografts through a hexokinase-2 (HK2) dependent inhibition of glycolysis. HK2 is a glucose 6-phosphorylating enzyme that works in conjunction with glucose transporter-1 (GLUT1) to drive glycolysis in cells. While our previous work suggests that BKIDC-1553 works primarily through a HK2 dependent mechanism, the objective of this current project is to further examine how expression and function of HK2 and GLUT1 are altered in response to BKIDC-1553 treatment. Methods: Hypoxic conditions are common in late-stage prostate cancer. Effects of hypoxia on cell proliferation and expression levels of HK1, HK2, and GLUT1 were examined with and without BKIDC-1553 treatment. To examine role of HK1, HK2, and GLUT1 in response to BKIDC-1553 treatment, we created CRISPR knockouts and shRNA knockdowns of HK1 and HK2 in the LNCaP prostate cancer cell line. GLUT1 was inhibited using a commercially available inhibitor, BAY-876. PCR and Western blots were used to confirm expression levels of HK1, HK2, and GLUT1. MTS proliferation assays were used to examine effect of KO and KD on response to BKIDC-1553 and BAY-876. Results: GLUT1 levels, but not HK1 or 2, increased at 72 hrs under hypoxic conditions. HK1, HK2, and GLUT1 levels all increased in cells in response to 144 hrs of BKIDC-1553 compared with 72 hrs treatment in both normoxic and hypoxic conditions. The KO and KD studies were all conducted in normoxic conditions. Westerns blots demonstrated that HK2 KO, but not HK1 KO, results in increased levels of GLUT1. HK1 or HK2 KD, however, did not alter GLUT1 levels. HK2 KD cells, with GLUT1 levels remaining the same, displayed less growth inhibition than control cells in response to BKIDC-1553 treatment, while HK2 KO cells, with increased levels of GLUT1, remained sensitive to BKIDC-1553. Because GLUT1 levels increase in response to HK2 KO, we wanted to see if inhibiting GLUT1 in these cells could decrease their proliferation. We found that KO cells were indeed more sensitive to growth inhibition of BAY-876. Conclusion/Summary: These studies indicate that KO of HK2 increases levels of GLUT1. These cells remain sensitive to BKIDC-1553 treatment, which could in part be due to the cells' continued reliance on glycolysis through increased GLUT1. A continued reliance on glycolysis allows BKIDC-1553 to continue to inhibit cell growth. Future studies examining how combination treatment with BKIDC-1553 and BAY-876 affects growth of advanced prostate cancer cells is underway. Citation Format: Cynthia Sprenger, Mika Munari, Shihua Sun, Kathryn Soriano Epilepsia, Stephen Plymate. Alterations in glycolytic pathway enzymes and metabolic plasticity in prostate cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 448.

Detection of glucose transporter 1 in living cells for assessment of tumor development and therapy using an electrochemical biosensor

The expression level of glucose transporter 1 (GLUT-1) is highly correlated with tumor malignancy, making it a promising therapeutic target for cancer treatment. The detection of GLUT-1 expression level is significant for cancer discovery and valuating the efficacy of drug treatments. However, current methods for GLUT-1 detection primarily rely on traditional techniques. Therefore, the development of anon-destructive in vivo monitoring system would be invaluable for assessing GLUT-1 expression and tumor responses to various drugs. In this study, an electrochemical platform for detection of GLUT-1 on living cells was established using reduced graphene oxide-multi-wall carbon nanotube composite (rGO-MWCNT) as a conductive coating and toluidine blue O (TBO)-graphene-gold nanoparticle-GLUT-1 antibody as the electrochemical probe. The sensor demonstrated excellent performance in detecting GLUT-1 on cells with a linear range of 10 - 105 cells/mL, good stability and selectivity. The sensor successfully detected GLUT-1 expressions in multiple tumor cell types, including those treated with siRNA or drugs, and the results were consistent with those obtained from traditional methods such as flow cytometry, western blotting, and immunofluorescence. The sensor is promising in evaluating the malignant level of tumor cells, distinguishing glucose uptake pathways in tumor cells, reducing medical costs, and facilitating the translation of electrochemical sensing technology to the clinical settings.

Comparative study of carbohydrate levels on growth, oxidative stress and glucolipid metabolism of hybrid fish between Megalobrama amblycephala (♀) × Culter alburnus (♂) and Culter alburnus

The present study used hybrid fish (Megalobrama amblycephala ♀ × Culter alburnus ♂, abbreviated as BT) and topmouth culter (Culter alburnus, abbreviated as TC) as research subjects to assess and compare the effects of carbohydrate on the growth, oxidative stress and glucolipid metabolism of the two fish. The TC (initial weight, 13.10 ± 0.10 g) and BT (initial weight, 12.85 ± 0.13 g) were fed isonitrogenous (43% crude protein) and isolipidic (6.5% crude lipid) diets with carbohydrate levels of 10% (LC diet) and 30% (HC diet) for 11 weeks, respectively. The results showed that the HC diet resulted in a significant decrease in specific growth rate (SGR) of both fish. However, the SGR in BT was significantly higher than in TC, regardless of diet. Furthermore, HC diet significantly increased serum alanine transaminase (GPT) and hepatic malondialdehyde (MDA) content, decreased hepatic catalase (CAT) activity and resulted in increased hepatocyte volume, nucleus deficiency and nucleus deviation from the center of cytoplasm in both fish. However, BT had significantly higher glutathione peroxidase (GPx) activity and lower serum GPT and aspartate transaminase (GOT) content compared to TC. The HC diet induced significant increases in hepatosomatic index (HSI), liver lipid droplet area, liver glycogen, serum glucose (GLU) and triglyceride (TG) content in both fish. However, HSI and lipid droplet area were significantly higher in BT than in TC, while serum GLU and TG content were significantly lower in BT than in TC. Both fish can adapt to the HC diet by up-regulating the expression of glycolysis-related gene (glucokinase (gk)) and lipogenesis-related genes (acetyl-CoA carboxylase a (acca), fatty acid synthase (fas)). However, the expression levels of glucose transporter 2 (glut2), glycolysis-related gene (phosphofructokinase liver b (pfklb)) and lipogenesis-related genes (sterol regulatory element-binding protein 1 (srebp1), ATP citrate lyase a (aclya)) were increased only in BT fed HC diet. In summary, the BT liver has stronger glucose transport, glycolysis, and lipid tolerance compared to TC, thus exhibiting better glucose homeostasis and growth performance. It indicates that hybridization may help to produce new strain of fish with high carbohydrate utilization capacity.

Neuregulin-1β increases glucose uptake and promotes GLUT4 translocation in palmitate-treated C2C12 myotubes by activating PI3K/AKT signaling pathway.

Insulin resistance (IR) is a feature of type 2 diabetes (T2DM) accompanied by reduced glucose uptake and glucose transporter 4 (GLUT4) translocation by skeletal muscle. Neuregulin-1β (NRG-1β) is essential for myogenesis and the regulation of skeletal muscle metabolism. Neuregulin-1β increases insulin sensitivity, promotes glucose uptake and glucose translocation in normal skeletal muscle. Here, we explored whether Neuregulin-1β increased glucose uptake and GLUT4 translocation in palmitate (PA)-treated C2C12 myotubes. After C2C12 myoblasts differentiated into myotubes, we used palmitate to induce cellular insulin resistance. Cells were incubated with or without Neuregulin-1β and glucose uptake was determined using the 2-NBDG assay. The expression level of glucose transporter 4 (GLUT4) was measured via immunofluorescence and Western blotting. MK2206, an inhibitor of AKT, was employed to reveal the important role played by AKT signaling in PA-treated C2C12 myotubes. We then established an animal model with T2DM and evaluated the effects of Neuregulin-1β on body weight and the blood glucose level. The GLUT4 level in the gastrocnemius of T2DM mice was also measured. NRG-1β not only increased glucose uptake by PA-treated myotubes but also promoted GLUT4 translocation to the plasma membrane. The effect of NRG-1β on PA-treated C2C12 myotubes was associated with AKT activation. In T2DM mice, Neuregulin-1β not only improved diabetes-induced weight loss and diabetes-induced hyperglycemia, but also promoted GLUT4 translocation in the gastrocnemius. In summary, Neuregulin-1β increased glucose uptake and promoted translocation of GLUT4 to the plasma membrane in PA-treated C2C12 myotubes by activating the PI3K/AKT signaling pathway.

Scavenging Reactive Oxygen Species Decreases Amyloid-β Levels via Activation of PI3K/Akt/GLUT1 Pathway in N2a/APP695swe Cells.

Dysregulated glucose metabolism in the brain is considered to be one of the key causes of Alzheimer's disease (AD). Abnormal glucose uptake in AD is tightly associated with decreased levels of glucose transporter 1 (GLUT1) and GLUT3 in the brain, but the underlying mechanisms remain unclear. We aimed to explore the cause and mechanism of impaired glucose uptake in AD. N2a/WT and N2a/APP695swe cells were cultured in vitro, and cellular glucose uptake and ATP content, as well as the expression of GLUT1, GLUT3, and PI3K/Akt pathway members, were detected. Intracellular reactive oxygen species (ROS) levels were detected by flow cytometry. After treatment with the ROS scavenger N-acetyl-L-cysteine (NAC), the above indicators were detected again. GLUT1 expression was significantly decreased (p = 0.0138) in N2a/APP695swe cells, while GLUT3 expression was no statistical difference (p > 0.05). After NAC treatment, PI3K and Akt phosphorylation levels, GLUT1 expression, glucose uptake and ATP levels were remarkably increased (p = 0.0006, p = 0.0008, p = 0.0009, p = 0.0001, p = 0.0013), while Aβ levels were significantly decreased (p = 0.0058, p = 0.0066). After addition of the PI3K inhibitor LY29004, GLUT1 expression was reduced (p = 0.0008), and Aβ levels were increased (p = 0.0009, p = 0.0117). In addition, increases in glucose uptake and ATP levels induced by the Akt activator SC79 were hindered by the GLUT1 inhibitor WZB117 (p = 0.0002, p = 0.0005). Aβ levels were decreased after SC79 treatment and increased after WZB117 treatment (p = 0.0212, p = 0.0006). Taken together, scavenging of ROS prevents from Aβ deposition via activation of the PI3K/Akt/GLUT1 pathway, and improved the impaired glucose uptake in N2a/APP695swe cells.

Construction of the experimental rat model of gestational diabetes.

Numerous methods for modeling gestational diabetes mellitus (GDM) in rats exist. However, their repeatability and stability are unclear. This study aimed to compare the effects of high-fat and high-sugar (HFHS) diet, HFHS diet combined with streptozotocin (STZ) administration, and HFHS diet combined with movement restriction (MR) modeling methods on rat models to confirm the best method for constructing a rat model of GDM. Forty female Sprague-Dawley rats were randomly divided into four groups (n = 10): the normal control (NC), HFHS, HFHS+STZ, and HFHS+MR groups. The rats in the NC group were fed with a standard diet, and those in the remaining groups were fed with a HFHS diet. The rats in the HFHS+STZ group received 25 mg/kg STZ on their first day of pregnancy, and those in the HFHS+MR group were subjected to MR during pregnancy. Bodyweight, food intake, water intake, fasting blood glucose (FBG), fasting insulin (FINS), homeostasis model assessment of insulin resistance (HOMA-IR), homeostasis model assessment of insulin sensitivity (HOMA-IS), homeostasis model assessment of β-cell function, pancreatic and placental morphology, and the expression levels of glucose transporter 1 (GLUT1) and glucose transporter 3 (GLUT3) in placentas were then quantified. Moreover, iTRAQ was used to identify placental proteomics. During pregnancy, the rats in the HFHS+STZ group showed FBG levels that were kept stable in a state of moderate hyperglycemia; the typical GDM symptoms of polydipsia, polyphagia, polyuria, and increased body weight; and the modeling rate of 87.5%. On the first and 19th days of pregnancy, the rats in the HFHS group showed higher FBG than that of the NC group, increasing body weight and food intake and the modeling rate of 50%. On the 19th day of pregnancy, the FBG of the rats in the HFHS+MR group was higher than that of the rats in the NC group, and the modeling rate of 42.9%. Comparison with the NC group revealed that the three modeling groups exhibited increased FINS and HOMA-IR, decreased HOMA-IS, and different degrees of pathological changes in pancreases and placentas. Among the groups, the HFHS+STZ group displayed the greatest changes with significant reductions in the numbers of pancreatic and placental cells and appeared cavitation. The expression levels of GLUT1 and GLUT3 in the placentas of the HFHS+STZ and HFHS+MR groups were higher than those in the placentas of the NC and HFHS groups. The above results indicated that the rats in the HFHS+STZ group showed the best performance in terms of modeling indicators. After the changes in placental proteomics in the HFHS+STZ group were compared with those in the NC group, we found that in the HFHS+STZ group, five proteins were up-regulated and 18 were down-regulated; these proteins were enriched in estrogen signaling pathways. HFHS combined with the intraperitoneal injection of 25 mg/kg STZ was the best modeling method for the nonspontaneous model of experimentally induced GDM, and its modeling rate was high. The pathological characteristics of the constructed GDM rat model were similar to those of human patients with GDM. Moreover, the model was stable and reliable. The modeling method can provide a basis for constructing a GDM rat model for subsequent research on the prevention and treatment of GDM.

A mathematical model of GLUT1 modulation in rods and RPE and its differential impact in cell metabolism

We present a mathematical model of key glucose metabolic pathways in two cells of the human retina: the rods and the retinal pigmented epithelium (RPE). Computational simulations of glucose transporter 1 (GLUT1) inhibition in the model accurately reproduce experimental data from conditional knockout mice and reveal that modification of GLUT1 expression levels of both cells differentially impacts their metabolism. We hypothesize that, under glucose scarcity, the RPE’s energy producing pathways are altered in order to preserve its functionality, impacting the photoreceptors’ outer segment renewal. On the other hand, when glucose is limited in the rods, aerobic glycolysis is preserved, which maintains the lactate contribution to the RPE.

Inducible knockout of syncytin-a leads to poor placental glucose transport in mice

IntroductionCell-cell fusion of cytotrophoblasts into the syncytiotrophoblast layer is a key process in placental development. Syncytin, an endogenous retroviral envelope protein, is expressed in placental trophoblasts and specifically mediates syncytiotrophoblast layer formation. Syncytin deficiency has been observed in fetal growth-restricted placentas. Abnormal fetal growth, especially fetal growth restriction, is associated with the decreased expression of glucose transporters. Here, we aimed to determine the role of syncytin in fetal growth restriction in placental glucose transport capacity. MethodsTo better explore the function of syncytin in fetal growth-restricted placenta, we generated an inducible knockout mouse model of syncytin-a gene. The expression levels of glucose transporters in BeWo cells were measured before and after HERV-W knockdown. ResultsSyncytin-A disruption was associated with significant abnormalities in placental and fetal development in mice. Syncytin-A destruction causes extensive abnormalities in the maternal-fetal exchange structures in the labyrinth, including an extremely reduced number and dramatically irregular distribution of fetal vessels. Moreover, glucose transporter 1, glucose transporters 3, and connexin 26 expression levels decreased after E14.5. Consistently, low glucose transporter 1, glucose transporter 3, and connexin 26 levels were observed in HERV-W-silenced BeWo cells. DiscussionSyncytin-A is crucial for both syncytiotrophoblast layer development and morphogenesis, suggesting that syncytin-A disruption leads to fetal growth restriction associated with abnormalities in the maternal-fetal exchange barrier and decreased glucose transport.

Different Glucose Metabolic Features According to Cancer and Immune Cells in the Tumor Microenvironment.

BackgroundA close metabolic interaction between cancer and immune cells in the tumor microenvironment (TME) plays a pivotal role in cancer immunity. Herein, we have comprehensively investigated the glucose metabolic features of the TME at the single-cell level to discover feasible metabolic targets for the tumor immune status.MethodsWe examined expression levels of glucose transporters (GLUTs) in various cancer types using The Cancer Genome Atlas (TCGA) data and single-cell RNA-seq (scRNA-seq) datasets of human cancer tissues including melanoma, head and neck, and breast cancer. In addition, scRNA-seq data of immune cells in the TME acquired from human melanoma after immune checkpoint inhibitors were analyzed to investigate the dynamics of glucose metabolic profiles of specific immune cells.ResultsPan-cancer bulk RNA-seq showed that the GLUT3-to-GLUT1 ratio was positively associated with immune cell enrichment score. The scRNA-seq datasets of various human cancer tissues showed that GLUT1 was highly expressed in cancer cells, while GLUT3 was highly expressed in immune cells in TME. The scRNA-seq data obtained from human melanoma tissues pre- and post-immunotherapy showed that glucose metabolism features of myeloid cells, particularly including GLUTs expression, markedly differed according to treatment response.ConclusionsDifferently expressed GLUTs in TME suggest that GLUT could be a good candidate a surrogate of tumor immune metabolic profiles and a target for adjunctive treatments for immunotherapy.

Plant hvu-MIR168-3p enhances expression of glucose transporter 1 (SLC2A1) in human cells by silencing genes related to mitochondrial electron transport chain complex I

Diet is a crucial factor for preventing most diseases. Edible plant extracts are known to contain exosome-like nanoparticles, in which food-derived plant microRNAs are included and may serve as a novel functional component in human health. Here, we demonstrated that hvu-MIR168-3p included in the nanoparticles of rice aleurone cells down-regulated the expression of the genes related to mitochondrial electron transport chain complex I in human cells. Subsequently, hvu-MIR168-3p enhanced protein and RNA expression levels of glucose transporter I and caused a decrease in the blood glucose level, which findings were obtained by in vitro and in vivo experiments, respectively. These findings suggest that a cross-kingdom relationship between plants and humans with respect to hvu-MIR168-3p exists and may contribute to preventive medicine for GLUT1-related dysfunctions including glucose metabolism, aging, and tumor immunology.

Berberine exerts its antineoplastic effects by reversing the Warburg effect via downregulation of the Akt/mTOR/GLUT1 signaling pathway.

Glucose transporter 1 (GLUT1) plays a primary role in the glucose metabolism of cancer cells. However, to the best of our knowledge, there are currently no anticancer drugs that inhibit GLUT1 function. The present study aimed to investigate the antineoplastic activity of berberine (BBR), the main active ingredient in numerous Traditional Chinese medicinal herbs, on HepG2 and MCF7 cells. The results of Cell Counting Kit-8 assay, colony formation assay and flow cytometry revealed that BBR effectively inhibited the proliferation of tumor cells, and induced G2/M cell cycle arrest and apoptosis. Notably, the results of luminescence ATP detection assay and glucose uptake assay showed that BBR also significantly inhibited ATP synthesis and markedly decreased the glucose uptake ability, which suggested that the antitumor effect of BBR may occur via reversal of the Warburg effect. In addition, the results of reverse transcription-quantitative PCR, western blotting and immunofluorescence staining indicated that BBR downregulated the protein expression levels of GLUT1, maintained the cytoplasmic internalization of GLUT1 and suppressed the Akt/mTOR signaling pathway in both HepG2 and MCF7 cell lines. Augmentation of Akt phosphorylation levels by the Akt activator, SC79, abolished the BBR-induced decrease in ATP synthesis, glucose uptake, GLUT1 expression and cell proliferation, and reversed the proapoptotic effect of BBR. These findings indicated that the antineoplastic effect of BBR may involve the reversal of the Warburg effect by downregulating the Akt/mTOR/GLUT1 signaling pathway. Furthermore, the results of the co-immunoprecipitation assay demonstrated that BBR increased the interaction between ubiquitin conjugating enzyme E2 I (Ubc9) and GLUT1, which suggested that Ubc9 may mediate the proteasomal degradation of GLUT1. On the other hand, BBR decreased the interaction between Gα-interacting protein-interacting protein at the C-terminus (GIPC) and GLUT1, which suggested that the retention of GLUT1 in the cytoplasm may be achieved by inhibiting the interaction between GLUT1 and GIPC, thereby suppressing the glucose transporter function of GLUT1. The results of the present study provided a theoretical basis for the application of the Traditional Chinese medicine component, BBR, for cancer treatment.

Calcitriol Suppresses Warburg Effect and Cell Growth in Human Colorectal Cancer Cells.

Increasing lines of evidence indicate that the biologically active form of vitamin D, calcitriol (1,25-dihydroxyvitamin D3), prevents cancer progression by reducing cell proliferation, increasing cell differentiation, and inhibiting angiogenesis, among other potential roles. Cancer cells in solid tumors preferably undergo the “Warburg effect” to support cell growth by upregulating glycolysis, and the glycolytic intermediates further serve as building blocks to generate biomass. The objective of the current study is to investigate whether calcitriol affects glucose metabolism and cell growth in human colorectal cancer cells. Calcitriol reduced the expression of cyclin D1 and c-Myc. In addition, calcitriol reduced the expression of glucose transporter 1 (GLUT1) and key glycolytic enzymes and decreased extracellular acidification rate but increased oxygen consumption rate in human colorectal cancer cells. In a subcutaneous HT29 xenograft NOD/SCID mouse model, the volume and weight of the tumors were smaller in the calcitriol groups as compared with the control group, and the expression levels of GLUT1 and glycolytic enzymes, hexokinase 2 and lactate dehydrogenase A, were also lower in the calcitriol groups in a dose-responsive manner. Our data indicate that calcitriol suppresses glycolysis and cell growth in human colorectal cancer cells, suggesting an inhibitory role of the biologically active form of vitamin D in colorectal cancer progression.

Ketamine ameliorates depressive-like behaviors in mice through increasing glucose uptake regulated by the ERK/GLUT3 signaling pathway

To investigate the effects of ketamine on glucose uptake and glucose transporter (GLUT) expression in depressive-like mice. After HA1800 cells were treated with ketamine, 2-[N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)Amino]-2-Deoxyglucose (2-NBDG) was added to the cells to test the effects of ketamine on glucose uptake, production of lactate, and expression levels of GLUT, ERK1/2, AKT, and AMPK. Adult female C57BL/6 mice were subjected to chronic unpredictable mild stress (CUMS), 27 CUMS mice were randomly divided into the depression, ketamine (i.p.10 mg/kg), and FR180204 (ERK1/2 inhibitor, i.p.100 mg/kg) + ketamine group. Three mice randomly selected from each group were injected with 18F-FDG at 6 h after treatment. The brain tissue was collected at 6 h after treatment for p-ERK1/2 and GLUTs. Treatment with ketamine significantly increased glucose uptake, extracellular lactic-acid content, expression levels of GLUT3 and p-ERK in astrocytes and glucose uptake in the prefrontal cortex (P < 0.05), and the immobility time was significantly shortened in depressive-like mice (P < 0.01). An ERK1/2 inhibitor significantly inhibited ketamine-induced increases in the glucose uptake in depressive-like mice (P < 0.05), as well as prolonged the immobility time (P < 0.01). The expression levels of p-ERK1/2 and GLUT3 in depressive-like mice were significantly lower than those in normal control mice (P < 0.01). Ketamine treatment in depressive-like mice significantly increased the expression levels of p-ERK1/2 and GLUT3 in the prefrontal cortex (P < 0.01), whereas an ERK1/2 inhibitor significantly inhibited ketamine-induced increases (P < 0.01).Our present findings demonstrate that ketamine mitigated depressive-like behaviors in female mice by activating the ERK/GLUT3 signal pathway, which further increased glucose uptake in the prefrontal cortex.

Study on effect of gypenosides on insulin sensitivity of rats with diabetes mellitus via regulating NF-κB signaling pathway

This study focused on the ameliorative effects of gypenosides(GPS) on insulin sensitivity and inflammatory factors in rats with type 2 diabetes mellitus(T2 DM) and explored their possible molecular mechanisms. After the successful establishment of T2 DM model, diabetic rats were randomly divided into four groups, including model group, GPS groups(200, 100 mg·kg~(-1)) and metformin group(100 mg·kg~(-1)), with healthy rats serving as the control. After 6-week intragastric administration, fasting blood glucose(FBG) and oral glucose tolerance were examined. The levels of insulin, C-peptide, tumor necrosis factor-α(TNF-α), interleukin-1β(IL-1β), interleukin-6(IL-6) and C-reactive protein(CRP) in serum were examined. Then the homeostasis model assessment of insulin resistance(HOMA-IR) and insulin sensitivity index(ISI) were calculated. The protein expression levels of phosphorylated insulin receptor substrate-1(p-IRS-1) and phosphorylated protein kinase B(p-Akt) in skeletal muscle were measured by Western blot, as well as those of phosphorylated inhibitor of nuclear factor-κB(NF-κB) kinase β(p-IKKβ), phosphorylated alpha inhibitor of NF-κB(p-IκBα) and phosphorylated p65 subunit of NF-κB(p-p65) in adipose tissue. The relative expression levels of glucose transporter 4(GLUT4) mRNA in skeletal muscle and NF-κB mRNA in adipose tissue were measured by qRT-PCR, and the morphological changes of pancreatic tissue were observed. Compared with the model group, the GPS groups witnessed significant decrease in FBG, marked amelioration of impaired oral glucose tolerance and significant increase in ISI. Further, the high-dose GPS group saw significantly reduced HOMA-IR, TNF-α, IL-1β and CRP, significantly increased expression levels of p-IRS-1(Tyr), p-Akt and GLUT4, and markedly inhibited p-IRS-1(Ser), p-IKKβ, p-IκBα, p-p65 and NF-κB. The concentration of CRP and the expression levels of p-IRS-1(Ser), p-IKKβ, p-IκBα and NF-κB were remarkably reduced in the low-dose GPS group. However, GPS was found less effective in the regulation of serum insulin, C-peptide and IL-6 levels and the alleviation of pancreatic islet injury. The results indicated that GPS can reduce FBG and improve insulin sensitivity in diabetic rats possibly by regulating the NF-κB signaling pathway, inhibiting inflammation, and thereby regulating the expression of key proteins in the insulin signaling pathway.

Hypouricemic Effects of Chrysanthemum indicum L. and Cornus officinalis on Hyperuricemia-Induced HepG2 Cells, Renal Cells, and Mice.

Hyperuricemia, abnormally excess accumulation of uric acid, is caused by an imbalance between the production and excretion of uric acid and is a major cause of gout. We compared the effects of extracts from Chrysanthemum indicum L. (Ci) and Cornus officinalis Siebold and Zucc. (Co) on hyperuricemia, both individually and in combination (FSU-CC), using hypoxanthine-treated human liver cancer (HepG2) cells, primary mouse renal proximal tubule cells, and potassium oxonate induced hyperuricemic mice. The Ci contained 7.62 mg/g luteolin and 0 mg/g loganin, Co contained 0 mg/g luteolin and 4.90 mg/g loganin, and FSH-CC contained 3.95 mg/g luteolin and 2.48 mg/g loganin. We found that treatment with Ci, Co, and FSU-CC suppressed the activity of xanthine oxidase and mRNA expression of xanthine dehydrogenase while inducing an increase in the expression levels of the organic anion transporter 1 (OAT1) and organic anion transporter 3 (OAT3) proteins and a decrease in the expression levels of glucose transporter 9 (GLUT9) and urate transporter 1 (URAT1) proteins. Particularly, treatment and supplementation with FSU-CC showed stronger effects than those of supplementation with either Ci or Co alone. We observed that the excretion of creatinine and uric acid in the combination of Ci and Co was higher than that observed in their individual supplementations and was similar to that of the normal group. Therefore, our data suggest that a combination of Ci and Co may potentially be used for the development of effective natural anti-hyperuricemic functional foods.

HIF-1α may promote glycolysis in psoriasis vulgaris via upregulation of CD147 and GLUT1.

To analyze the expressions and distributions of hypoxia-inducible factor-1α (HIF-1α), CD147, and glucose transporter 1 (GLUT1) in epidermis from psoriasis vulgaris and normal people, and to explore the associations among these proteins and their roles in hypoxic HaCaT cell line. The expression levels of HIF-1α, CD147, and GLUT1 were determined by immunohistochemistry staining in skin biopsies from 48 psoriasis vularis patients and 33 healthy subjects. Cobalt chloride (CoCl2) was added into the culture media of HaCaT cells to mimic hypoxia while RNA interference and transfection technologies were used to explore the association among these proteins by quantitative real-time polymerase chain reaction and Western blotting. Glycolytic capacity was detected by ATP and lactate measurements. HIF-1α, CD147, and GLUT1 were highly expressed and the glycolytic capacity was increased in lesions of psoriasis vulgaris; HIF-1α upregulated the expression of CD147 and GLUT1, increased the lactate production and decreased the ATP level in CoCl2-treated HaCaT cells, while CD147 and GLUT1 directly or indirectly bound to each other. Glycolytic capacity increases in the injured keratinocytes of psoriasis vulgaris, suggesting that HIF-1α, CD147, and GLUT1 are associated with glycolysis, which can be considered as the promising targets for psoriasis therapy.

Prognostic value of glycolysis markers in head and neck squamous cell carcinoma: a meta-analysis.

Glycolysis markers including glucose transporter 1 (GLUT1), monocarboxylate transporter 4 (MCT4), hexokinase 2 (HK2), pyruvate kinase M2 (PKM2) and glucose transporter 4 (GLUT4) play vital roles in head and neck squamous cell carcinoma (HNSCC). However, their prognostic value in HNSCC is still controversial. In this meta-analysis, we searched the PubMed, Web of Science and Cochrane Library databases and included thirty-seven studies (3272 patients) that met the inclusion criteria. Higher expression levels of the glycolysis markers in tumor tissues correlated with poorer overall survival (OS; P < 0.001), disease-free survival (DFS; P = 0.03) and recurrence-free survival (RFS; P < 0.001) of HNSCC patients. Subgroup and sensitivity analyses demonstrated that higher expression levels of GLUT1 (P < 0.001), MCT4 (P = 0.002), HK2 (P = 0.002) and PKM2 (P < 0.001) correlated with poorer OS among HNSCC patients. Higher expression of MCT4 (P < 0.001) and PKM2 (P = 0.008) predicted poorer DFS among HNSCC patients. However, GLUT4 expression levels did not associate with clinical outcomes in HNSCC patients. These results demonstrate that glycolysis markers, such as GLUT1, MCT4, HK2 and PKM2, are potential prognostic predictors and therapeutic targets in HNSCC.

Effects of epigallocatechin-3-gallate combined with ascorbic acid and glycerol on the stability and uric acid-lowering activity of epigallocatechin-3-gallate

Context Epigallocatechin-3-gallate (EGCG) is unstable and easily oxidized, which limits its applications. Ascorbic acid (Vc) is a natural antioxidant. Objective The effects of EGCG combined with Vc and glycerol on stability and uric acid-lowering activity of EGCG were examined. Materials and methods EGCG (aqueous solution), EGCG + Vc (aqueous solution), EGCG (glycerol solution) and EGCG + Vc (glycerol solution) were prepared and incubated under different conditions in vitro. The recovery rate of EGCG was calculated by HPLC. Kunming mice were randomly divided into normal control group, model group, allopurinol (5 mg/kg), EGCG (10 mg/kg), EGCG + Vc (both 10 mg/kg), EGCG (10 mg/kg) + glycerol (60%), and EGCG (10 mg/kg) + Vc (10 mg/kg) + glycerol (60%) (n = 6). Allopurinol was injected intraperitoneally to mice, others were administered intragastrically to (2 cases) mice. All mice were continuously administrated for 7 days, once a day. Results EGCG recovery rates of EGCG group and EGCG + Vc + glycerol group respectively reached to 32.34 ± 1.86% and 98.90 ± 0.64% when they were incubated for 4 h at 80 °C. EGCG recovery rates reached to 91.82 ± 5.13% (incubated for 6 h at pH 8) and 88.85 ± 2.63% (incubated for 4 h in simulated intestinal fluid) when EGCG incubated with Vc and glycerol. Compared with the model group, UA values of EGCG + Vc + glycerol group reduced by 43.49% while EGCG group reduced by 25.63%. The activities of xanthine oxidase (XOD, 31.41 U/gprot) and adenosine deaminase (ADA, 10.05 U/mgprot), and the mRNA expression levels of glucose transporter 9 (GLUT9, 1.03) and urate transporter 1 (URAT1, 0.44) in EGCG + Vc + glycerol group were notably lower than those of EGCG group (38.12 U/gprot, 13.16 U/mgprot, 1.54, and 0.55). The mRNA expression levels of ATP-binding cassette superfamily G member 2 (ABCG2, 1.39) and organic anion transport 1/2 (OAT1/2, 2.34, 2.53) in EGCG + Vc + glycerol group were notably higher than those of EGCG group (0.57, 1.13, and 1.16). Discussion and conclusions Our findings suggest that when EGCG used in combination with Vc and glycerol could effectively increase its biology activities and can be generalized to the broader pharmacological studies. This sheds light on the development and application of EGCG in the fields of food and medicine.

Paclitaxel inhibits the migration of CD133+ U251 malignant glioma cells by reducing the expression of glycolytic enzymes.

Energy metabolic reprogramming (EMR) allows for the rearrangement of a series of metabolic genes and proteins when tumor cells adapt to their microenvironment. EMR is characterized by changes in the metabolic pattern and metabolic intermediates to meet the needs of tumor cells for their malignant proliferation and infiltrative growth. The present study investigated the role of low-dose paclitaxel (PTX) in changing the expression levels of key genes and proteins during glycolysis in CD133+ U251 glioma cells and explored the relevant regulatory mechanisms of action at the molecular level. CD133 immunomagnetic beads were applied to malignant CD133+ U251 glioma cells, which were then divided into a negative control and an experimental group treated with 1, 2, 4 or 8 µM PTX for 72 h. Cell Counting Kit-8 (CCK-8) was used to measure U251 cell proliferation. RNA and protein were extracted from the malignant glioma cells in all groups to observe changes in the expression levels of key glycolytic enzymes, such as glucose transporter 1 (GLUT1), pyruvate kinase M (PKM) and lactate dehydrogenase A (LDHA), using reverse transcription-quantitative PCR and western blot assays. Transwell migration assays were performed to quantify the effects of PTX solution on U251 cells. CD133+ U251 glioma cells were isolated successfully. CD1133+ cells had a higher rate of proliferation compared with CD1133- cells. In CD1133+ cells treated with PTX, a dose-dependent reduction in the expression levels of the key glycolytic enzymes GLUT1, PKM and LDHA was observed at both the mRNA and protein levels. PTX solution also inhibited cell migration. Differences between the control and experimental groups were statistically significant (P<0.05). Since glycolysis plays an indispensable role in the proliferation and migration of stem cell-like glioma cells, PTX may inhibit tumor cell growth by downregulating the gene and protein expression levels of glycolytic enzymes in CD133+ glioma cells.

Metabolic Alterations Predispose to Seizure Development in High-Fat Diet-Treated Mice: the Role of Metformin.

The link between epilepsy and type 2 diabetes (T2DM) and/or metabolic syndrome (MetS) has been poorly investigated. Therefore, we tested whether a high-fat diet (HFD), inducing insulin-resistant diabetes and obesity in mice, would increase susceptibility to develop generalized seizures induced by pentylentetrazole (PTZ) kindling. Furthermore, molecular mechanisms linked to glucose brain transport and the effects of the T2DM antidiabetic drug metformin were also studied along with neuropsychiatric comorbidities. To this aim, two sets of experiments were performed in CD1 mice, in which we firstly evaluated the HFD effects on some metabolic and behavioral parameters in order to have a baseline reference for kindling experiments assessed in the second section of our protocol. We detected that HFD predisposes towards seizure development in the PTZ-kindling model and this was linked to a reduction in glucose transporter-1 (GLUT-1) expression as observed in GLUT-1 deficiency syndrome in humans but accompanied by a compensatory increase in expression of GLUT-3. While we confirmed that HFD induced neuropsychiatric alterations in the treated mice, it did not change the development of kindling comorbidities. Furthermore, we propose that the beneficial effects of metformin we observed towards seizure development are related to a normalization of both GLUT-1 and GLUT-3 expression levels. Overall, our results support the hypothesis that an altered glycometabolic profile could play a pro-epileptic role in human patients. We therefore recommend that MetS or T2DM should be constantly monitored and possibly avoided in patients with epilepsy, since they could further aggravate this latter condition.