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Indoleamine 2,3‐dioxygenase 1 (IDO1), the key enzyme in the catabolism of the essential amino acid tryptophan (Trp) through kynurenine pathway, induces immune tolerance and is considered as a critical immune checkpoint, but its impacts as a metabolism enzyme on glucose and lipid metabolism are overlooked. We aim to clarify the potential role of IDO1 in aerobic glycolysis in pancreatic cancer (PC). Analysis of database revealed the positive correlation in PC between the expressions of IDO1 and genes encoding important glycolytic enzyme hexokinase 2 (HK2), pyruvate kinase (PK), lactate dehydrogenase A (LDHA) and glucose transporter 1 (GLUT1). It was found that IDO1 could modulate glycolysis and glucose uptake in PC cells, Trp deficiency caused by IDO1 overexpression enhanced glucose uptake by stimulating GLUT1 translocation to the plasma membrane of PC cells. Besides, Trp deficiency caused by IDO1 overexpression suppressed the apoptosis of PC cells via promoting glycolysis, which reveals the presence of IDO1–glycolysis–apoptosis axis in PC. IDO1 inhibitors could inhibit glycolysis, promote apoptosis, and exhibit robust therapeutic efficacy when combined with GLUT1 inhibitor in PC mice. Our study reveals the function of IDO1 in the glucose metabolism of PC and provides new insights into the therapeutic strategy for PC. [ABSTRACT FROM AUTHOR]

Abstract Indoleamine 2,3‐dioxygenase 1 (IDO1), the key enzyme in the catabolism of the essential amino acid tryptophan (Trp) through kynurenine pathway, induces immune tolerance and is considered as a critical immune checkpoint, but its impacts as a metabolism enzyme on glucose and lipid metabolism are overlooked. We aim to clarify the potential role of IDO1 in aerobic glycolysis in pancreatic cancer (PC). Analysis of database revealed the positive correlation in PC between the expressions of IDO1 and genes encoding important glycolytic enzyme hexokinase 2 (HK2), pyruvate kinase (PK), lactate dehydrogenase A (LDHA) and glucose transporter 1 (GLUT1). It was found that IDO1 could modulate glycolysis and glucose uptake in PC cells, Trp deficiency caused by IDO1 overexpression enhanced glucose uptake by stimulating GLUT1 translocation to the plasma membrane of PC cells. Besides, Trp deficiency caused by IDO1 overexpression suppressed the apoptosis of PC cells via promoting glycolysis, which reveals the presence of IDO1–glycolysis–apoptosis axis in PC. IDO1 inhibitors could inhibit glycolysis, promote apoptosis, and exhibit robust therapeutic efficacy when combined with GLUT1 inhibitor in PC mice. Our study reveals the function of IDO1 in the glucose metabolism of PC and provides new insights into the therapeutic strategy for PC.

Background and aims: It is particularly important to identify the progression of non-alcoholic fatty liver disease (NAFLD) for prognosis evaluation and treatment guidance. The aim of this study was to explore the clinic use of exosomal protein-based detection as a valuable non-invasive diagnostic method for NAFLD. Methods: Exosomes were extracted from plasma of patients with NAFLD using Optima XPN-100 ultrafast centrifuge. The patients were recruited from outpatients and inpatients of Beijing Youan Hospital Affiliated to Capital Medical University. The exosomes were stained with fluorescent-labeled antibody and determined by ImageStream® X MKII imaging flow cytometry. Generalized linear logistic regression model was used to evaluate the diagnostic value of hepatogenic exosomes in NAFLD and liver fibrosis. Results: The percentage of hepatogenic exosomes glucose transporter 1 (GLUT1) in patients with non-alcoholic steatohepatitis (NASH) was significantly higher than that in patients with non-alcoholic fatty liver (NAFL). According to liver biopsy, we found that the percentage of hepatogenic exosomes GLUT1 in patients with advanced NASH (F2-4) was significantly higher than that in patients with early NASH (F0-1), and the same trend was observed in exosomes with CD63 and ALB. Compared with other clinical fibrosis scoring criteria (FIB-4, NFS, etc.), the diagnostic performance of hepatogenic exosomes GLUT1 was the highest and the area under the receiver-operating curves (AUROC) was 0.85 (95% CI 0.77–0.93). Furthermore, the AUROC of hepatogenic exosomes GLUT1 combined with fibrosis scoring was as high as 0.86–0.91. Conclusion: Hepatogenic exosome GLUT1 can be a molecular biomarker for early warning of NAFLD to distinguish the NAFL and NASH, and it also can be used as a novel non-invasive diagnostic biomarker for the staging liver fibrosis in NAFLD. [ABSTRACT FROM AUTHOR]

Background: Dihydropyrimidinase-like 3 (DPYSL3) is a cytosolic phosphoprotein expressed in the nervous system and is crucial for neurogenesis. A previous study showed that increased DPYSL3 expression promotes tumour aggressiveness in pancreatic ductal adenocarcinoma, gastric cancer, and colon cancer. However, the role of DPYSL3 in affecting the biological behaviour of urothelial carcinoma (UC) is not yet understood. Methods: A UC transcriptomic dataset from the Gene Expression Omnibus and the Urothelial Bladder Cancer (BLCA) dataset from The Cancer Genome Atlas were used for the in silico study. We collected 340 upper urinary tract urothelial carcinoma (UTUC) and 295 urinary bladder urothelial carcinoma (UBUC) samples for the immunohistochemical study. Fresh tumour tissue from 50 patients was used to examine the DPYSL3 mRNA level. In addition, urothelial cell lines with and without DPYSL3 knockdown were used for the functional study. Results: The in silico study revealed that DPYSL3 correlated with advanced tumour stage and metastasis development while functioning primarily in the nucleobase-containing compound metabolic process (GO:0006139). DPYSL3 mRNA expression is significantly upregulated in advanced UC. Furthermore, overexpression of the DPYSL3 protein is significantly associated with the aggressive behaviour of UTUC and UBUC. DPYSL3 expression independently predicts disease-specific survival (DSS) and metastatic-free survival (MFS) in patients with UC. In non-muscle-invasive UBUC, DPYSL3 expression predicts local recurrence-free survival. UC cell lines with DPYSL3 knockdown exhibited decreased proliferation, migration, invasion, and human umbilical vein endothelial cells (HUVECs) tube formation but increased apoptosis and G1 arrest. Gene ontology enrichment analysis revealed that the enriched processes related to DPYSL3 overexpression in UC were tissue morphogenesis, cell mesenchyme migration, smooth muscle regulation, metabolic processes, and RNA processing. In vivo study revealed DPYSL3 knockdown in UC tumours significantly suppressed the growth of tumours and decreased MYC and GLUT1 protein expression. Conclusions: DPYSL3 promotes the aggressiveness of UC cells by changing their biological behaviours and is likely associated with cytoskeletal and metabolic process modifications. Furthermore, DPYSL3 protein overexpression in UC was associated with aggressive clinicopathological characteristics and independently predicted poor clinical outcomes. Therefore, DPYSL3 can be used as a novel therapeutic target for UC. [ABSTRACT FROM AUTHOR]

Background: The pumping function of corneal endothelial cells (CECs) plays a pivotal role in the maintenance of corneal water homeostasis. Corneal endothelial dysfunction (CED) leads to corneal edema and opacity, but with the exception of keratoplasty, no optimal therapeutic strategies have been established for CED. In this study, we aimed to investigate the ameliorative effect of ascorbic acid (AA) on CED and the underlying mechanism of action in the corneal endothelium. Methods: Rabbit corneal endothelial damage was induced by anterior chamber injection of benzalkonium chloride (BAK). AA was topically administered to the corneal surface, and the transparency and thickness of the cornea were assessed by external eye photography, slit-lamp photography, and ultrasonic pachymetry. To further analyze the mechanism, rabbit CECs and immortalized human CECs (B4G12 cells) were cultured. A ferric reducing/antioxidant and AA (FRASC) assay was performed to measure the AA concentration. Cell proliferation was evaluated by cell counting and bromodeoxyuridine (BrdU) labeling assays, and protein expression was examined by liquid chromatography–mass spectrometry (LC/MS) and immunoblotting. The involvement of glucose transporter 1 (GLUT1) and phospho-ERK was evaluated via GLUT1-siRNA and phospho-ERK inhibitor (PD98059) treatment. Interpretation: We observed that topical AA ameliorates BAK-induced rabbit corneal endothelial damage. Furthermore, we demonstrated that AA is transported into B4G12 cells via GLUT1, and afterward, AA increases ERK phosphorylation and promotes cell proliferation. Our findings indicate that CEC proliferation stimulated via the noncanonical AA-GLUT1-ERK axis contributes to AA-enhanced healing of CED.

Recent epidemiological and preclinical studies have revealed that aspirin possesses antitumor properties; one of the mechanisms results from inhibition of angiogenesis. However, the underlying mechanisms of such action remain to be elucidated, in particular, the effect of aspirin on glucose metabolism of vascular endothelial cells (ECs) has not yet been reported. Herein, we demonstrate that glucose transporter 1 (GLUT1), a main glucose transporter in ECs, can be down-regulated by aspirin. Exposure to 4-mM aspirin significantly decreased GLUT1 at the mRNA and protein level, resulting in impaired glucose uptake capacity in vascular ECs. In addition, we also showed that exposure to 4-mM aspirin led to an inhibition of intracellular ATP and lactate synthesis in vascular ECs, and a down-regulation of the phosphorylation level of NF-κB p65 was observed. Taken together, these findings indicate 4-mM aspirin inhibits glucose uptake and glucose metabolism of vascular ECs through down-regulating GLUT1 expression and suggest that GLUT1 has potential to be a target for aspirin in vascular ECs.

An association study was conducted to investigate the relation between 14 variants of glucose transporter 1 gene (SLC2A1) and the risk of type 2 diabetes (T2DM) leading to nephropathy. We also performed a meta-analysis of 11 studies investigating association between diabetic nephropathy (DN) and SLC2A1 variants. The cohort included 197 cases (T2DM with nephropathy), 155 diseased controls (T2DM without nephropathy) and 246 healthy controls. The association of variants with disease progression was tested using generalized odds ratio (ORG). The risk of type 2 diabetes leading to nephropathy was estimated by the OR of additive and co-dominant models. The mode of inheritance was assessed using the degree of dominance index (h-index). We synthesized results of 11 studies examining association between 5 SLC2A1 variants and DN. ORG was used to assess the association between variants and DN using random effects models. Significant results were derived for co-dominant model of rs12407920 [OR = 2.01 (1.17–3.45)], rs841847 [OR = 1.73 (1.17–2.56)] and rs841853 [OR = 1.74 (1.18–2.55)] and for additive model of rs3729548 [OR = 0.52 (0.29–0.90)]. The mode of inheritance for rs12407920, rs841847 and rs841853 was ‘dominance of each minor allele’ and for rs3729548 ‘non-dominance’. Frequency of one haplotype (C-G-G-A-T-C-C-T-G-T-C-C-A-G) differed significantly between cases and healthy controls [p = .014]. Regarding meta-analysis, rs841853 contributed to an increased risk of DN [(ORG = 1.43 (1.09–1.88); ORG = 1.58 (1.01–2.48)] between diseased controls versus cases and healthy controls versus cases, respectively. Further studies confirm the association of rs12407920, rs841847, rs841853, as well as rs3729548 and the risk of T2DM leading to nephropathy.

Recent epidemiological and preclinical studies have revealed that aspirin possesses antitumor properties; one of the mechanisms results from inhibition of angiogenesis. However, the underlying mechanisms of such action remain to be elucidated, in particular, the effect of aspirin on glucose metabolism of vascular endothelial cells (ECs) has not yet been reported. Herein, we demonstrate that glucose transporter 1 (GLUT1), a main glucose transporter in ECs, can be down-regulated by aspirin. Exposure to 4-mM aspirin significantly decreased GLUT1 at the mRNA and protein level, resulting in impaired glucose uptake capacity in vascular ECs. In addition, we also showed that exposure to 4-mM aspirin led to an inhibition of intracellular ATP and lactate synthesis in vascular ECs, and a down-regulation of the phosphorylation level of NF-κB p65 was observed. Taken together, these findings indicate 4-mM aspirin inhibits glucose uptake and glucose metabolism of vascular ECs through down-regulating GLUT1 expression and suggest that GLUT1 has potential to be a target for aspirin in vascular ECs. [ABSTRACT FROM AUTHOR]

Fucoidans are complex fucose-containing sulfated polysaccharides with pronounced anticancer effects. Their structure-anticancer activity relationships are difficult to determine due to fucoidans' complex, often irregularities-including structures. Fucoidan-active enzymes can be used for this propose. We have investigated two new recombinant endo -fucanases FWf3 and FWf4 from the marine bacterium Wenyingzhuangia fucanilytica CZ1127T that belong to the 107 family of glycoside hydrolases (GH). Both enzymes cleaved α-(1→4)-glycosidic bonds but in fucoidan fragments with different sulfation patterns. FWf3 is the first characterized endo -fucanase that cleaves glycosidic bonds between 2O- and 2,4diO-sulfated L-fucose residues. The obtained endo -fucanases were used to produce low- and high-molecular weight fucoidan derivatives with different sulfate group locations. Low- and high-molecular weight fucoidan derivatives rich with 2,4diO-sulfation were shown to inhibit MDA-MB-231 cell colony formation more efficiently than the native fucoidan and the derivatives sulfated otherwise. Such derivatives effectively suppressed the mitochondrial membrane potential of MDA-MB-231 cells and reduced the expression of the glucose transporter 1 (GLUT1). Co-treatment of MDA-MB-231 cells with the fucoidan derivatives and oligomycin (an OXPHOS inhibitor) resulted in a synergistic anticancer effect. The data obtained demonstrate, that fucoidan and its 2,4diO-sulfated derivatives can be an effective adjunct in TNBC therapy targeting cell metabolism. [Display omitted] [ABSTRACT FROM AUTHOR]

An association study was conducted to investigate the relation between 14 variants of glucose transporter 1 gene (SLC2A1) and the risk of type 2 diabetes (T2DM) leading to nephropathy. We also performed a meta-analysis of 11 studies investigating association between diabetic nephropathy (DN) and SLC2A1 variants. The cohort included 197 cases (T2DM with nephropathy), 155 diseased controls (T2DM without nephropathy) and 246 healthy controls. The association of variants with disease progression was tested using generalized odds ratio (ORG). The risk of type 2 diabetes leading to nephropathy was estimated by the OR of additive and co-dominant models. The mode of inheritance was assessed using the degree of dominance index (h-index). We synthesized results of 11 studies examining association between 5 SLC2A1 variants and DN. ORG was used to assess the association between variants and DN using random effects models. Significant results were derived for co-dominant model of rs12407920 [OR = 2.01 (1.17-3.45)], rs841847 [OR = 1.73 (1.17-2.56)] and rs841853 [OR = 1.74 (1.18-2.55)] and for additive model of rs3729548 [OR = 0.52 (0.29-0.90)]. The mode of inheritance for rs12407920, rs841847 and rs841853 was 'dominance of each minor allele' and for rs3729548 'non-dominance'. Frequency of one haplotype (C-G-G-A-T-C-C-T-G-T-C-C-A-G) differed significantly between cases and healthy controls [p = .014]. Regarding meta-analysis, rs841853 contributed to an increased risk of DN [(ORG = 1.43 (1.09-1.88); ORG = 1.58 (1.01-2.48)] between diseased controls versus cases and healthy controls versus cases, respectively. Further studies confirm the association of rs12407920, rs841847, rs841853, as well as rs3729548 and the risk of T2DM leading to nephropathy. [ABSTRACT FROM AUTHOR]

Diabetes mellitus is known to exacerbate cerebral ischemic injury. In the present study, we investigated antiapoptotic and anti-inflammatory effects of oral supplementation of ascorbic acid (AA) on cerebral injury caused by middle cerebral artery occlusion and reperfusion (MCAO/Re) in rats with streptozotocin-induced diabetes. We also evaluated the effects of AA on expression of sodium-dependent vitamin C transporter 2 (SVCT2) and glucose transporter 1 (GLUT1) after MCAO/Re in the brain. The diabetic state markedly aggravated MCAO/Re-induced cerebral damage, as assessed by infarct volume and edema. Pretreatment with AA (100 mg/kg, p.o.) for two weeks significantly suppressed the exacerbation of damage in the brain of diabetic rats. AA also suppressed the production of superoxide radical, activation of caspase-3, and expression of proinflammatory cytokines (tumor necrosis factor-α and interleukin-1β) in the ischemic penumbra. Immunohistochemical staining revealed that expression of SVCT2 was upregulated primarily in neurons and capillary endothelial cells after MCAO/Re in the nondiabetic cortex, accompanied by an increase in total AA (AA + dehydroascorbic acid) in the tissue, and that these responses were suppressed in the diabetic rats. AA supplementation to the diabetic rats restored these responses to the levels of the nondiabetic rats. Furthermore, AA markedly upregulated the basal expression of GLUT1 in endothelial cells of nondiabetic and diabetic cortex, which did not affect total AA levels in the cortex. These results suggest that daily intake of AA attenuates the exacerbation of cerebral ischemic injury in a diabetic state, which may be attributed to anti-apoptotic and anti-inflammatory effects via the improvement of augmented oxidative stress in the brain. AA supplementation may protect endothelial function against the exacerbated ischemic oxidative injury in the diabetic state and improve AA transport through SVCT2 in the cortex.

Glucose transporter 1 (GLUT1) is a facilitative glucose transporter overexpressed in various types of tumors; thus, it has been considered as an important target for cancer therapy. GLUT1 works through conformational switching from an outward-open (OOP) to an inward-open (IOP) conformation passing through an occluded conformation. It is critical to determine which conformation is preferred by bound ligands because the success of structure-based drug design depends on the appropriate starting conformation of the target protein. To find out the most favorable GLUT 1 conformation for ligand binding, we ran systemic molecular docking studies for different conformations of GLUT1 using known GLUT1 inhibitors. Our data revealed that the IOP is the preferred conformation and that residues Phe291, Phe379, Glu380, Trp388, and Trp412 may play critical roles in ligand binding to GLUT1. Our data suggests that conformational differences in these five amino acids in the different conformers of GLUT1 may be used to design ligands that inhibit GLUT1.

The blood-brain barrier (BBB) plays a vital role in the central nervous system (CNS). A comprehensive understanding of BBB development has been hampered by difficulties in observing the differentiation of brain endothelial cells (BECs) in real-time. Here, we generated two transgenic zebrafish line, Tg(glut1b:mCherry) and Tg(plvap:EGFP) , to serve as in vivo reporters of BBB development. We showed that barriergenesis ( i.e. the induction of BEC differentiation) occurs immediately as endothelial tips cells migrate into the brain parenchyma. Using the Tg(glut1b:mCherry) transgenic line, we performed a genetic screen and identified a zebrafish mutant with a nonsense mutation in gpr124 , a gene known to play a role in CNS angiogenesis and BBB development. We also showed that our transgenic plvap:EGFP line, a reporter of immature brain endothelium, is initially expressed in newly formed brain endothelial cells, but subsides during BBB maturation. Our results demonstrate the ability to visualize the in vivo differentiation of brain endothelial cells into the BBB phenotype and establish that CNS angiogenesis and barriergenesis occur simultaneously. [ABSTRACT FROM AUTHOR]

Despite the recent development of anti-myeloma drugs, the prognosis of high-risk multiple myeloma remains poor. Therefore, new effective treatment strategies for this disease are needed. It has been reported that high intensity of 18-fluorodeoxyglucose positron emission tomography is high-risk factor in myeloma, suggesting that glucose uptake can be therapeutic target in high-risk myeloma. In this study, we addressed the utility of glucose transporter 1 (GLUT1) as a therapeutic target for myeloma with increased glucose uptake. We found myeloma cell lines with elevated glucose uptake activity via GLUT1 up-regulation. STF-31, a selective GLUT1 inhibitor, completely suppressed the glucose uptake activity and induced apoptosis in GLUT1 expressing myeloma cells. On the other hand, this agent little shows the cytotoxicity in normal peripheral blood mononuclear cells. Moreover, STF-31 synergistically enhanced the cell death induced by melphalan, doxorubicin, and bortezomib. GLUT1 may be promising therapeutic target in myeloma with elevated glucose uptake. [ABSTRACT FROM AUTHOR]

We previously used the expression of various combinations and configurations of MyoD and E12, two basic helix-loop-helix transcription factors (TF), to produce populations of myotubes assuming distinct morphology, myofibrillar development and Ca 2+ dynamics, from mammalian C2C12 myoblasts in non-differentiation growth conditions. Here, we assessed the synthetically generated myotubes in terms of energetics, otherwise necessary to sustain their mechanical output as bio-actuators. We found that the myotubes exhibit changed expression of key regulators for the uptake and utilization of two major cellular fuels, glucose and lactate. Furthermore, while lactate transport was uniformly slowed in all the populations of myotubes, glucose uptake and utilization were modified by particular TF configuration. Our approach allows the production of a class of biomaterials with predetermined energetics that could be applied in biorobotics, where fuel of choice could be used, and also in reparative medicine where, for example, particular population of myotubes could be additionally employed as glucose sinks to mitigate effects of secondary metabolic syndrome. [ABSTRACT FROM AUTHOR]

Abstract: Hypoxia-inducible transcription factor-1 (HIF-1) plays a decisive role in cell survival and adaptation to hypoxic stress by controlling the expression of genes involved in oxygen homeostasis. HIF-1 activity is fine-tuned through specific post-translational modifications of its essential HIF-1α subunit. Among these modifications, phosphorylation is important for HIF-1 transcriptional activity. Studies have shown that the mitogen-activated protein kinases, p42/p44 MAPKs, directly phosphorylate HIF-1α and increase HIF-1-mediated transcription. Pin1, a peptidyl-prolyl cis/trans isomerase, targets a number of proteins containing a phosphorylated Ser/Thr-Pro motif. Pin1 isomerization causes a change in target protein conformation which can modify their activity. Here, we identify Pin1 as an important HIF-1α partner. Immunoprecipitation and pull-down studies show that Pin1 interacts with HIF-1α. We demonstrate that the interaction between Pin1 and HIF-1α is regulated through p42/p44 MAPK pathway activation. By performing proteolysis studies, our results indicate that Pin1 catalytic activity generates a conformational change in HIF-1α. Finally, our work shows that Pin1 is required for gene-specific HIF-1 transcriptional activity. Our results indicate that the prolyl isomerase Pin1 regulates HIF-1 transcriptional activity by interacting with HIF-1α and promoting conformational changes in a p42/p44 MAPK phosphorylation-dependent manner. [Copyright &y& Elsevier]

Abstract: Chronic kidney disease is a risk factor for cardiovascular mortality. This study uncovers pieces of hematological and erythrocyte protein variability observed in end stage renal disease (ESRD) in relation to disease progression/duration and mortality. Using a variety of experimental approaches, erythropoietin/dialysis-treated patients were compared to healthy individuals and had been followed for 36months. During that period, half of the patients died from cardiovascular diseases. The high levels of uremic toxins in those patients were associated with damaged erythrocytes, bad tolerance and poor response to hemodialysis therapy. The postmortem study revealed significant variation in alkaline phosphatase, duration of dialysis, erythrocyte transformation and intracellular hemoglobin concentration compared to the survived patients. The erythrocyte proteins showed substantial remodeling characteristic of pathologic regulation of cell hydration and susceptibility to the dialysis-induced oxidation defects. According to the follow-up study, duration of hemodialysis was associated with a trend towards increased intracellular hemoglobin concentration, membrane expression of glucose transporter-1 and stomatin as well as lower levels of circulating stomatocytes. The uremic index variation in long survived patients is accurately reflected in plasma and erythrocyte oxidative stress modifications. The ESRD patients exhibit impressive compensatory responses to the chronic challenges of the uremic milieu. Biological significance: This study demonstrates novel blood modifications probably associated with the duration of erythropoietin/hemodialysis treatment, disease progression and cardiovascular mortality in end stage renal disease. The observed variability adds new pieces to the erythrocyte pathophysiology puzzle in end stage renal disease and suggests novel hematologic and proteomic factors for consideration in future large scale studies on cardiovascular morbidity and mortality candidate biomarkers in uremic patients. [Copyright &y& Elsevier]

Abstract: In heart failure mitochondrial dysfunction is thought to be responsible for energy depletion and contractile dysfunction. The difficulties in procuring fresh left ventricular (LV) myocardium from humans for assessment of mitochondrial function have resulted in the reliance on surrogate markers of mitochondrial function and limited our understanding of cardiac energetics. We isolated mitochondria from fresh LV wall tissue of patients with heart failure and reduced systolic function undergoing heart transplant or left ventricular assist device placement, and compared their function to mitochondria isolated from the non-failing LV (NFLV) wall tissue with normal systolic function from patients with pulmonary hypertension undergoing heart–lung transplant. We performed detailed mitochondrial functional analyses using 4 substrates: glutamate–malate (GM), pyruvate–malate (PM) palmitoyl carnitine–malate (PC) and succinate. NFLV mitochondria showed preserved respiratory control ratios and electron chain integrity with only few differences for the 4 substrates. In contrast, HF mitochondria had greater respiration with GM, PM and PC substrates and higher electron chain capacity for PM than for PC. Surprisingly, HF mitochondria had greater respiratory control ratios and lower ADP-independent state 4 rates than NFLV mitochondria for GM, PM and PC substrates demonstrating that HF mitochondria are capable of coupled respiration ex vivo. Gene expression studies revealed decreased expression of key genes in pathways for oxidation of both fatty acids and glucose. Our results suggest that mitochondria from the failing LV myocardium are capable of tightly coupled respiration when isolated and supplied with ample substrates. Thus energy starvation in the failing heart may be the result of dysregulation of metabolic pathways, impaired substrate supply or reduced mitochondrial number but not the result of reduced mitochondrial electron transport capacity. [Copyright &y& Elsevier]

Abstract: Pre-diabetes, a risk factor for type 2 diabetes development, leads to metabolic changes at testicular level. Peroxisome proliferator-activated receptor γ coactivator 1 α (PGC-1α) and Sirtuin 3 (Sirt3) are pivotal in mitochondrial function. We hypothesized that pre-diabetes disrupts testicular PGC-1α/Sirt3 axis, compromising testicular mitochondrial function. Using a high-energy-diet induced pre-diabetic rat model, we evaluated testicular levels of PGC-1α and its downstream targets, nuclear respiratory factors 1 (NRF-1) and 2 (NRF-2), mitochondrial transcription factor A (TFAM) and Sirt3. We also assessed mitochondrial DNA (mtDNA) content, mitochondrial function, energy levels and oxidative stress parameters. Protein levels were quantified by Western Blot, mtDNA content was determined by qPCR. Mitochondrial complex activity and oxidative stress parameters were spectrophotometrically evaluated. Adenine nucleotide levels, adenosine and its metabolites (inosine and hypoxanthine) were determined by reverse-phase HPLC. Pre-diabetic rats showed increased blood glucose levels and impaired glucose tolerance. Both testicular PGC-1α and Sirt3 levels were decreased. NRF-1, NRF-2 and TFAM were not altered. Testicular mtDNA content was decreased. Mitochondrial complex I activity was increased, whereas mitochondrial complex III activity was decreased. Adenylate energy charge was decreased in pre-diabetic rats, as were ATP and ADP levels. Conversely, AMP levels were increased, evidencing a decreased ATP/AMP ratio. Concerning to oxidative stress pre-diabetes decreased testicular antioxidant capacity and increased lipid and protein oxidation. In sum, pre-diabetes compromises testicular mitochondrial function by repressing PGC-1α/Sirt3 axis and mtDNA copy number, declining respiratory capacity and increasing oxidative stress. This study gives new insights into overall testicular bioenergetics at this prodromal stage of disease. [Copyright &y& Elsevier]

Highlights: [•] Adipose tissue hypoxia induce production of hypoxia markers. [•] Hypoxia mediates the inflammatory primarily via macrophages rather than adipocytes. [•] Resveratrol downregulates most of the hypoxia mediated effects. [Copyright &y& Elsevier]

Abstract: It is becoming increasingly evident that improving the cure rate of many cancers will require treatment regimens hit more than one validated tumor targets. Developing an anti-cancer agent that targets two oncoproteins simultaneously is a promising strategy for accomplishing this goal. It would be expected to promote drug efficacy, reduce therapy-resistant without introducing additional toxic side effects. HIF-1α is a key regulator of the cellular response to hypoxia and is involved in tumor angiogenesis and cancer cell survival, glucose metabolism, and invasion. Stat3 has several oncogenic functions, including suppression of anti-tumor immune responses and promotion of inflammation. Recently, we have identified the perylene derivative, TEL03, as a dual inhibitor that targets both HIF-1α and Stat3. TEL03 blocks the expression of both HIF-1α and Stat3, regulated oncogenes (e.g., Bcl-2, VEGF, Glut1, and others) in cancer cells, and induces cancer cell apoptosis. The results demonstrated that: (i) TEL03 blocks Stat3 phosphorylation, and inhibits Stat3 transcriptional activity; and (ii) interferes the binding of HIF-1α to p300/CBP inducing its degradation by proteasomes under hypoxic conditions. Our in vivo tests showed that as a dual inhibitor, TEL03 dramatically inhibited tumor growth, and provided the evidence that targeting both HIF-1α and Stat3 simultaneously could be a promising strategy for breast and pancreatic cancer therapies. [Copyright &y& Elsevier]

Highlights: [•] Caffeine reduced p53α expression and induced the expression of p53β. [•] Caffeine alters the expression of serine/arginine-rich splicing factors. [•] SRSF3 is the primary target of caffeine for the regulation of cellular functions. [Copyright &y& Elsevier]

Highlights: [•] Inhibition of GSK3β enhanced HIF1α protein expression and expression of its target genes in osteosarcoma cells. [•] GSK3β did not affect HIF1α protein stability in osteosarcoma cells. [•] Inhibition of GSK3β upregulated HIF1α mRNA expression in osteosarcoma cells. [•] Inhibition of GSK3β enhanced HIF1α mRNA stability in osteosarcoma cells. [•] Inhibition of GSK3β upregulated nucleolin mRNA expression in osteosarcoma cells. [Copyright &y& Elsevier]

Highlights: [•] 1,25(OH)2D reduces glucose addiction of ras-oncogene transformed MCF10A breast epithelial cells. [•] 1,25(OH)2D reduces glucose utilization in glycolysis, lactate production and the TCA cycle. [•] Regulation of cellular glucose metabolism is a novel mechanism for 1,25(OH)2D action in cancer prevention. [Copyright &y& Elsevier]

Highlights: [•] We used a rat model of hyperglycemia-enhanced hemorrhagic transformation. [•] Rosiglitazone reduced infarct volume after focal ischemia. [•] Rosiglitazone reduced hemorrhagic transformation. [•] Rosiglitazone increased expression of collagen IV by preservation of GLUT1. [ABSTRACT FROM AUTHOR]

Highlights: [•] P. falciparum secretes 17 FIKK kinases into host erythrocytes. [•] We report here the first heterologous expression and purification of a FIKK kinase. [•] We show that recombinant FIKK kinase PfFk4.1 is an active protein kinase. [•] We identify dematin as a substrate of recombinant Fk4.1. [ABSTRACT FROM AUTHOR]

Abstract: Targeting of intracerebral functional regions has been limited by the inability to transport drugs across the blood–brain barrier (BBB) and by poor accumulation in these regions. To overcome these hurdles, liposomes modified with P-aminophenyl-α-d-mannopyranoside (MAN) were used as a fluorescent dye carrier through the BBB and used the specific distribution of liposomes (LIP) modified with MAN (MAN–LIP) to target various functional regions of the brain. An in vitro BBB model was established to evaluate the transendothelial ability of MAN–LIP, and liposomes uptake by C6 glioma cells was analyzed by flow cytometry and live cell imaging. Liposome targeting was evaluated using in vivo and ex vivo imaging. After MAN–LIP administration, the transendothelial ability and the delivery of fluorescent dye to the brain significantly increased. MAN–LIP concentrated in the cortex at 4h, shifting distribution to the cerebellum and brainstem at 12h. The fluorescence intensity in the hippocampus and pontine nuclei remained high and stable over a period of 12h. The results demonstrate that MAN–LIP is able to enhance cellular uptake in vitro and also promotes penetration through the BBB and accumulation in the brain with a distinct spatio-temporal pattern. [Copyright &y& Elsevier]

Highlights: [•] This is the first study to show the structural requirements for the binding of a porphyrin compound to HSP90. [•] An in silico molecular docking model and experimental approaches revealed the anti-angiogenic activities of PPIX. [•] A tetrapyrrole macrocycle and propionate chains of porphyrins coordinately interact with the ATP-binding pocket of HSP90. [•] Our results may offer key structural information on the potential development of a new class of HSP90 inhibitors. [Copyright &y& Elsevier]

Highlights: [•] Albumin overload induced the expression of HIF-1α protein and mRNA in HK-2 cells. [•] HIF-1α protein was predominantly localized in nucleus in albumin-overloaded cells. [•] The HIF-1 target gene mRNAs (GLUT1 and GAPDH) were up-regulated by albumin. [ABSTRACT FROM AUTHOR]

This study investigated the expression of the plasma membrane markers aquaporin 1 (AQP1), glucose transporter 1 (GLUT1) and the a1 subunit of Na, K-ATPase in normal canine mammary glands and in benign and malignant mammary tumours, using immunohistochemistry and semi-quantitative histomorphometry. AQP1 immunoreactivity was absent from the majority of specimens studied. GLUT1 immunoreactivity was observed in normal mammary tissue and particularly in the epithelial and mesenchymal cells of benign, and in the epithelial cells of malignant tumours, respectively. Na, K-ATPase immunoreactivity was present in normal and neoplastic mammary epithelium and was significantly increased in the epithelium of both benign and malignant tumours. These results suggest that GLUT1 is more highly expressed in neoplastic epithelium and mesenchyme and that Na, K-ATPase is more highly expressed in neoplastic mammary epithelium. In consequence, these membrane proteins may have potential as diagnostic and prognostic biomarkers of canine mammary neoplasia. [ABSTRACT FROM AUTHOR]

The pumping function of corneal endothelial cells (CECs) plays a pivotal role in the maintenance of corneal water homeostasis. Corneal endothelial dysfunction (CED) leads to corneal edema and opacity, but with the exception of keratoplasty, no optimal therapeutic strategies have been established for CED. In this study, we aimed to investigate the ameliorative effect of ascorbic acid (AA) on CED and the underlying mechanism of action in the corneal endothelium. Rabbit corneal endothelial damage was induced by anterior chamber injection of benzalkonium chloride (BAK). AA was topically administered to the corneal surface, and the transparency and thickness of the cornea were assessed by external eye photography, slit-lamp photography, and ultrasonic pachymetry. To further analyze the mechanism, rabbit CECs and immortalized human CECs (B4G12 cells) were cultured. A ferric reducing/antioxidant and AA (FRASC) assay was performed to measure the AA concentration. Cell proliferation was evaluated by cell counting and bromodeoxyuridine (BrdU) labeling assays, and protein expression was examined by liquid chromatography–mass spectrometry (LC/MS) and immunoblotting. The involvement of glucose transporter 1 (GLUT1) and phospho-ERK was evaluated via GLUT1-siRNA and phospho-ERK inhibitor (PD98059) treatment. We observed that topical AA ameliorates BAK-induced rabbit corneal endothelial damage. Furthermore, we demonstrated that AA is transported into B4G12 cells via GLUT1, and afterward, AA increases ERK phosphorylation and promotes cell proliferation. Our findings indicate that CEC proliferation stimulated via the noncanonical AA-GLUT1-ERK axis contributes to AA-enhanced healing of CED. [ABSTRACT FROM AUTHOR]

Drug resistance is often a limiting factor in successful chemotherapy. Our laboratory has been interested in studying mechanisms of resistance to drugs that are targeted to the thymidylate biosynthesis pathway especially those that target thymidylate synthase (TS) and dihydrofolate reductase (DHFR). We have used leukemia as a model system to study resistance to methotrexate (MTX) and colorectal cancer as the model system to study 5-fluorouracil (5-FU) resistance. In leukemias, we and others have shown that transport, efflux, polyglutamylation and hydrolase activities are major determinants of MTX resistance. We have further reported that some leukemic cells have an increase in DHFR gene copy number possibly contributing to the resistant phenotype. Recently, we have begun to study in detail the molecular mechanisms that govern translational regulation of DHFR in response to MTX as an additional resistance mechanism. Studies thus far involving colorectal tumors obtained from patients have focused predominantly on the predictive value of levels of TS expression and p53 mutations in determining response to 5-FU. Although the predictive value of these two measures appears to be significant, given the variety of resistance to 5-FU observed in cell lines, it is not likely that these are the only measures predictive of response or responsible for acquired resistance to this drug. The enzyme uridine–cytidine monophosphate kinase (UMPK) is an essential and rate-limiting enzyme in 5-FU activation while dihydropyrimidine dehydrogenase (DPD) is a catabolic enzyme that inactivates 5-FU. Alterations in UMPK and DPD may therefore explain failure of 5-FU response in the absence of alterations in TS or p53. Transcription factors that regulate TS may also influence drug sensitivity. We have found that mRNA levels of the E2F family of transcription factors correlates with TS message levels and are higher in lung metastases than in liver metastases of colorectal cancers. Moreover, gene copy number of the E2F-1 gene appears to be increased in a significant number of samples obtained from metastases of colorectal cancer. We have also generated mutants of both DHFR and TS that confer resistance to MTX as well as 5-FU by random as well as site-directed mutagenesis. These mutants used alone or as fusion cDNAs of the mutants have proven to be useful in transplant studies where transfer of these mutant cDNAs to bone marrow cells have been shown to confer drug resistance to recipients. The fusion cDNAs of DHFR such as the DHFR–herpes simplex virus type 1 thymidine kinase (HSVTK) are also useful for regulation of gene expression in vivo using MTX as the small molecule regulator that can be monitored by positron emission tomography (PET) scanning or by optical imaging using a fusion construct such as DHFR–EGFP. [Copyright &y& Elsevier]

Salidroside is reported to have a wide range of pharmacological properties and has been proven to play a key anti-cancer effect. This study investigated the effects of purified salidroside, an ingredient of Rhodiola rosea, on the proliferation of two human gastric cancer cell lines and further investigating its possible molecular mechanisms. We verified that salidroside exerts a dose-dependent inhibitory effect on the proliferation of SGC-7901 and MKN-45 human gastric cancer cells. Moreover, salidroside can induce cell apoptosis, which was accompanied by an increase in nuclear fragmentation. In addition, salidroside inhibited glycolysis, as evidenced by the reduced expression levels of the glycolysis-related enzymes pyruvate kinase isoenzyme M2 (PKM2), enolase 1 (ENO1) and glucose transporter 1 (GLUT1), which could play important roles in the metabolism of gastric cancer cells. Further investigation showed that salidroside exerted potent anti-proliferative effects by inhibiting glycolysis in human gastric cancer cells in vitro. In vivo, xenograft tumors treated with salidroside were significantly smaller than those in the control animals. Therefore, salidroside could be a promising therapeutic prospect in the treatment of gastric cancer.

Human endometrial stromal cells (ESCs) differentiate into decidual cells by the action of progesterone, which is essential for implantation and maintenance of pregnancy. We previously reported that glucose uptake by human ESCs increases during decidualization and that glucose is indispensable for decidualization. Although glucose transporter 1 (GLUT1) is upregulated during decidualization, it remains unclear whether it is involved in glucose uptake. Here, we attempted to determine the role of GLUT1 during decidualization as well as the factors underlying its upregulation. ESCs were incubated with cAMP to induce decidualization. Knockdown of GLUT1 suppressed cAMP-increased glucose uptake and the expressions of specific markers of decidualization, IGF-binding protein-1 (IGFBP-1), and prolactin (PRL). To investigate the regulation of GLUT1 expression, we focused on CCAAT enhancer-binding protein β (C/EBPβ) and Wilms' tumor 1 (WT1) as the upstream transcription factors regulating GLUT1 expression. Knockdown of either C/EBPβ or WT1 suppressed cAMP-increased GLUT1 expression and glucose uptake. cAMP treatment also increased the recruitment of C/EBPβ and WT1 to the GLUT1 promoter region. Interestingly, cAMP increased the H3K27 acetylation (H3K27ac) and p300 recruitment in the GLUT1 promoter region. Knockdown of C/EBPβ or WT1 inhibited these events, indicating that both C/EBPβ and WT1 contribute to the increase of H3K27ac by recruiting p300 to the GLUT1 promoter region during decidualization. These findings indicate that GLUT1 is involved in glucose uptake in ESCs during decidualization, thus facilitating the establishment of pregnancy., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Glucose transporter 1 (GLUT1) is a facilitative glucose transporter overexpressed in various types of tumors; thus, it has been considered as an important target for cancer therapy. GLUT1 works through conformational switching from an outward-open (OOP) to an inward-open (IOP) conformation passing through an occluded conformation. It is critical to determine which conformation is preferred by bound ligands because the success of structure-based drug design depends on the appropriate starting conformation of the target protein. To find out the most favorable GLUT 1 conformation for ligand binding, we ran systemic molecular docking studies for different conformations of GLUT1 using known GLUT1 inhibitors. Our data revealed that the IOP is the preferred conformation and that residues Phe291, Phe379, Glu380, Trp388, and Trp412 may play critical roles in ligand binding to GLUT1. Our data suggests that conformational differences in these five amino acids in the different conformers of GLUT1 may be used to design ligands that inhibit GLUT1. [ABSTRACT FROM AUTHOR]

Chronic infection of HPV16 E6/E7 is frequently associated with lung cancers, especially in non-smokers and in Asians. In our previous studies, we found that HPV16 E6/E7 up-regulated HIF-1α at protein level and further up-regulated GLUT1 at both protein and mRNA levels in well-established lung cancer cell lines. In one of our further mechanism study, the results demonstrated that HPV16 E6/E7 up-regulated the expression of GLUT1 through HPV-LKB1-HIF-1α-GLUT1 axis. However, there are multiple pathways involved in HPV16 E6/E7 regulation of HIF-1α expression. In current study, using double directional genetic manipulation in well-established lung cancer cell lines, we showed that both E6 and E7 down-regulated the expression of RRAD at both protein and mRNA levels. Like LKB1, RRAD is one of the cancer suppressor genes. The loss of RRAD further activated NF-κB by promoted cytoplasmic p65 translocated to nucleus, and up-regulated the expression level of the p-p65 in nucleus. Furthermore, p-p65 up regulated HIF-1α and GLUT1 at both protein and mRNA levels. Thus, we proposed HPV16 E6/E7 up-regulated the expression of GLUT1 through HPV-RRAD-p65- HIF-1α- GLUT1 axis. In conclusion, we demonstrated for the first time that E6 and E7 promoted the expression of HIF-1α and GLUT1 by relieving the inhibitory effect of RRAD which resulted in the activation of NF-κB by promoting cytoplasmic p65 translocated to nucleus, and up-regulated the expression of the p-p65 in nucleus in lung cancer cells. Our findings provided new evidence to support the critical role of RRAD in the pathogenesis of HPV-related lung cancer, and suggested novel therapeutic targets., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Activation of the immune system occurs in response to the recognition of foreign antigens and receipt of optimal stimulatory signals by immune cells, a process that requires energy. Energy is also needed to support cellular growth, differentiation, proliferation, and effector functions of immune cells. In HIV-infected individuals, persistent viral replication, together with inflammatory stimuli contributes to chronic immune activation and oxidative stress. These conditions remain even in subjects with sustained virologic suppression on antiretroviral therapy. Here we highlight recent studies demonstrating the importance of metabolic pathways, particularly those involving glucose metabolism, in differentiation and maintenance of the activation states of T cells and monocytes. We also discuss how changes in the metabolic status of these cells may contribute to ongoing immune activation and inflammation in HIV- infected persons and how this may contribute to disease progression, establishment and persistence of the HIV reservoir, and the development of co-morbidities. We provide evidence that other viruses such as Epstein-Barr and Flu virus also disrupt the metabolic machinery of their host cells. Finally, we discuss how redox signaling mediated by oxidative stress may regulate metabolic responses in T cells and monocytes during HIV infection., (Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.)