Curcumin Inhibits the PPARδ-p-Akt-GLUT1 Pathway and Ameliorates the Antiproliferative Effects of Doxorubicin in MDA-MB-231 Cells.

OBJECTIVE: The aim of the present study was to examine whether GLUT1 was involved in the antiproliferative activity of curcumin and doxorubicin by understanding mechanistically how curcumin regulated GLUT1. METHODS: Expression level of GLUT1 in MCF-7 and MDA-MB-231 cells were quantitated using quantitative real-time PCR and western blot. GLUT1 activity was inhibited in MDA-MB-231 cells with the pharmacological inhibitor WZB117 to assess the anti-proliferative effects of doxorubicin using MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide).  To examine cell proliferation, trypan blue assay was used in cells transfected with GLUT1 siRNA or plasmid overexpressing GLUT1 with doxorubicin and/or commercially available curcumin. The role of PPARδ and Akt on the regulation of GLUT1 by curcumin was examined by overexpressing these proteins and western blot was employed to examine their protein expression. RESULTS: The data revealed that there was a 1.5 fold increase in GLUT1 mRNA and protein levels in MDA-MB-231 compared to MCF-7.  By inhibiting GLUT1 in triple negative breast cancer cell line, MDA-MB-231 with either the pharmacological inhibitor WZB117 or with GLUT1 siRNA, we observed the enhanced antiproliferative effects of doxorubicin. Additional observations indicated these effects can be reversed by the overexpression of GLUT1. Treatment of MDA-MB-231 with curcumin also revealed downregulation of GLUT1, with further growth suppressive effects when combined with doxorubicin.  Overexpression of GLUT1 blocked the growth suppressive role of curcumin and doxorubicin (p< 0.05). Mechanistically, we also observed that the regulation of GLUT1 by curcumin was mediated by the Peroxisome proliferator-activated receptor (PPAR) δ/Akt pathway. CONCLUSION: Our study demonstrates that regulation of GLUT1 by curcumin via the PPARδ/Akt signaling improves the efficacy of doxorubicin by promoting its growth inhibitory effects in MDA-MB-231 cells.

Identification of SLC2A1 as a predictive biomarker for survival and response to immunotherapy in lung squamous cell carcinoma.

BACKGROUND: As one of the common subtypes of non-small lung cancer, lung squamous cell carcinoma (LUSC) patients with advanced stage have few choices of treatment strategies. Therefore, it is urgent to discover genes that are associated with the survival and efficacy of immunotherapies. METHOD: Differential gene expression analyses were conducted using TCGA LUSC bulk-sequencing and single-cell RNA-sequencing data. Prognostic genes were identified from the TCGA LUSC cohort. Protein expression validation and survival analyses were performed. Experiments were conducted to explore the underlying mechanisms. In addition, the correlation between gene expression and pathological response to adjuvant immunochemotherapy was also investigated. RESULTS: After a series of bioinformatic analyses, solute carrier family 2 member 1(SLC2A1), encoding glucose transporter-1 (GLUT1), was found to be differentially expressed between tumor and normal tissues. GLUT1 was subsequently identified as an independent prognostic factor for LUSC. GSEA analysis revealed the glycolysis metabolism pathway of KEGG enriched in SLC2A1high tumor tissues. LASSO analyses revealed that tumor tissues with high expression of SLC2A1 were associated with high levels of protein lactylation. We found that SLC2A1 was preferentially expressed by SPP1+ macrophages in the tumor microenvironment, and the expression of SLC2A1 was associated with the abundance of SPP1+ macrophages. Immunofluorescence demonstrated GLUT1 and HIF1α colocalization in tumor-infiltrating macrophages. In vitro experiments showed HIF-1α-induced macrophage polarization under hypoxia, and GLUT1 inhibition blocked this polarization. In addition, SLC2A1 was negatively associated with the common immune checkpoint molecules, such as programmed cell death 1(PD-1), T cell immunoreceptor with Ig and ITIM domains (TIGIT), cytotoxic T-lymphocyte associated protein 4 (CTLA4) and lymphocyte activating 3 (LAG3), while showed a positive association with CD44. Finally, we observed that there was a significant correlation between pre-adjuvant-treatment GLUT1 expression and the pathological response. CONCLUSION: SLC2A1 expression was differentially upregulated in tumor tissues, and elevated GLUT1 expression was associated with worse survival and poor pathological response to adjuvant immunochemotherapy. Upregulation of GLUT1 promoted macrophage polarization into the M2 phenotype. The findings will contribute to guiding the treatment selection for LUSC patients and providing personalized immunotherapy strategies.

PIGT promotes cell growth, glycolysis, and metastasis in bladder cancer by modulating GLUT1 glycosylation and membrane trafficking.

BACKGROUND: Bladder cancer is very common worldwide. PIGT is a subunit of the glycosylphosphatidylinositol transamidase which involves in tumorigenesis and invasiveness. m6A modification of mRNA has been linked to cell proliferation, tumor progression and other biological events. However, how PIGT is regulated and what is the function of PIGT in bladder cancer remains to be elucidated. METHODS: PIGT was silenced or overexpressed to study its role in regulating bladder cancer. Cell proliferation and invasion were examined with the Cell Counting Kit-8, colony formation and Transwell assay, respectively. Cellular oxygen consumption rates or extracellular acidification rates were detected by a XF24 Analyzer. Quantitative RT-PCR and immunoblots were performed to detect mRNA and protein levels. RESULTS: PIGT was overexpressed in bladder cancer. Silencing PIGT inhibited cell proliferation, oxidative phosphorylation, and glycolysis. Overexpressing PIGT promoted cell proliferation, oxidative phosphorylation, glycolysis in vitro and tumor metastasis in vivo by activating glucose transporter 1 (GLUT1). PIGT also promoted GLUT1 glycosylation and membrane trafficking. Wilms' tumor 1-associated protein (WTAP) mediated PIGT m6A modification, and m6A reader, insulin-like growth factor 2 mRNA-binding protein (IGF2BP2), binds to the methylated PIGT to promote the stability of PIGT, leading to up-regulation of PIGT. CONCLUSION: WTAP mediates PIGT m6A modification to increase the stability of PIGT via the IGF2BP2, which enhances cell proliferation, glycolysis, and metastasis in bladder cancer by modulating GLUT1 glycosylation and membrane trafficking.

Revisiting the roles of glucose transporters in skeletal muscle physiology: is GLUT10 a novel player?

Skeletal muscle is the largest metabolic tissue responsible for systemic glucose handling. Glucose uptake into skeletal tissue is highly dynamic and delicately regulated, in part through the controlled expression and subcellular trafficking of multiple types of glucose transporters. Although the roles of GLUT4 in skeletal muscle metabolism are well established, the physiological significance of other, seemingly redundant, glucose transporters remain incompletely understood. Nonetheless, recent studies have shed light on the roles of several glucose transporters, such as GLUT1 and GLUT10, in skeletal muscle. Mice experiments suggest that GLUT10 could be a novel player in skeletal muscle metabolism in the context of mechanical overload, which is in line with the meta-analytical results of gene expression changes after resistance exercise in humans. Herein we discuss the knowns, unknowns, and implications of these recent findings.

Valproic acid elevates HIF-1α-mediated CGB expression and suppresses glucose uptake in BeWo cells.

Placental dysfunction can disrupt pregnancy. However, few studies have assessed the effects of chemical-induced toxicity on placental function. Here, we examined the effects of valproic acid (VPA) as a model chemical on production of hormones and on glucose uptake in human choriocarcinoma cell line BeWo. Cells were treated with forskolin to differentiate into syncytiotrophoblasts, which were then treated with VPA for 72 hr. Real-time RT-PCR analysis showed that VPA significantly increased the mRNA expression of chorionic gonadotropin ß (CGB), a hormone that is produced by the placenta in the first trimester of pregnancy, relative to that in the forskolin-only group. It also suppressed the increase in intracellular glucose uptake and GLUT1 level observed in the forskolin-only group. RNA-seq analysis and pathway database analysis revealed that VPA consistently decreased the level of HIF-1α protein and expression of its downstream target genes HK2 and ADM in the hypoxia pathway. Cobalt chloride, a HIF-1α inducer, inhibited CGB upregulation in VPA-treated cells and rescued VPA-induced suppression of glucose uptake and GLUT1 level. Thus, HIF-1α-mediated elevation of CGB expression and suppression of glucose uptake by VPA is a novel mechanism of placental dysfunction.

Photochemically Controlled Release of the Glucose Transporter 1 Inhibitor for Glucose Deprivation Responses and Cancer Suppression Research.

Cancer cells need a greater supply of glucose mainly due to their aerobic glycolysis, known as the Warburg effect. Glucose transport by glucose transporter 1 (GLUT1) is the rate-limiting step for glucose uptake, making it a potential cancer therapeutic target. However, GLUT1 is widely expressed and performs crucial functions in a variety of cells, and its indiscriminate inhibition will cause serious side effects. In this study, we designed and synthesized a photocaged GLUT1 inhibitor WZB117-PPG to suppress the growth of cancer cells in a spatiotemporally controllable manner. WZB117-PPG exhibited remarkable photolysis efficiency and substantial cytotoxicity toward cancer cells under visible light illumination with minimal side effects, ensuring its safety as a potential cancer therapy. Furthermore, our quantitative proteomics data delineated a comprehensive portrait of responses in cancer cells under glucose deprivation, underlining the mechanism of cell death via necrosis rather than apoptosis. We reason that our study provides a potentially reliable cancer treatment strategy and can be used as a spatiotemporally controllable trigger for studying nutrient deprivation-related stress responses.

Generation of antibodies to an extracellular region of the transporters Glut1/Glut4 by immunization with a designed antigen.

Monoclonal antibodies are one of the fastest growing class of drugs. Nevertheless, relatively few biologics target multispanning membrane proteins because of technical challenges. To target relatively small extracellular regions of multiple membrane-spanning proteins, synthetic peptides, which are composed of amino acids corresponding to an extracellular region of a membrane protein, are often utilized in antibody discovery. However, antibodies to these peptides often do not recognize parental membrane proteins. In this study, we designed fusion proteins in which an extracellular helix of the membrane protein glucose transporter 1 (Glut1) was grafted onto the scaffold protein Adhiron. In the initial design, the grafted fragment did not form a helical conformation. Molecular dynamics simulations of full-length Glut1 suggested the importance of intramolecular interactions formed by surrounding residues in the formation of the helical conformation. A fusion protein designed to maintain such intramolecular interactions did form the desired helical conformation in the grafted region. We then immunized an alpaca with the designed fusion protein and obtained VHH (variable region of heavy-chain antibodies) using the phage display method. The binding of these VHH antibodies to the recombinant Glut1 protein was evaluated by surface plasmon resonance, and their binding to Glut1 on the cell membrane was further validated by flow cytometry. Furthermore, we also succeeded in the generation of a VHH against another integral membrane protein, glucose transporter 4 (Glut4) with the same strategy. These illustrates that our combined biochemical and computational approach can be applied to designing other novel fusion proteins for generating site-specific antibodies.

GLUT1 mediates the release of HMGB1 from airway epithelial cells in mixed granulocytic asthma.

Asthma is quite heterogenous and can be categorized as eosinophilic, mixed granulocytic (presence of both eosinophils and neutrophils in the airways) and neutrophilic. Clinically, mixed granulocytic asthma (MGA) often tends to be severe and requires large doses of corticosteroids. High mobility group box 1 (HMGB1) is one of the epithelium-derived alarmins that contributes to type 2 inflammation and asthma. This study was aimed to investigate the role of glucose transporter 1 (GLUT1) in modulation of airway epithelial HMGB1 production in MGA. Induced sputum and bronchial biopsy specimens were obtained from healthy subjects and asthma patients. BALB/c mice, the airway epithelial cell line BEAS-2B, or primary human bronchial epithelial cells (HBECs) were immunized with allergens. Intracellular and extracellular HMGB1 were both detected. The role of GLUT1 was assessed by using a pharmacological antagonist BAY876. MGA patients have a significant higher sputum HMGB1 level than the health and subjects with other inflammatory phenotypes. Nuclear-to-cytoplasmic translocation of HMGB1 was also observed in the bronchial epithelia. Allergen exposure markedly induced GLUT1 expression in murine lungs and cultured epithelial cells. Pharmacological antagonism of GLUT1 with BAY876 dramatically decreased airway hyperresponsiveness, neutrophil and eosinophil accumulation, as well as type 2 inflammation in murine models of MGA. Besides, the allergen-induced up-regulation of HMGB1 was also partly recovered by BAY876, accompanied by inhibited secretion into the airway lumen. In vitro, treatment with BAY876 relieved the allergen-induced over-expression and secretion of HMGB1 in airway epithelia. Taken together, our data indicated that GLUT1 mediates bronchial epithelial HMGB1 release in MGA.

The genetic variant SLC2A1 -rs1105297 is associated with the differential analgesic response to a glucose-based treatment in newborns.

ABSTRACT: Neonatal pain is a critical issue in clinical practice. The oral administration of glucose-based solutions is currently one of the most common and effective nonpharmacologic strategies for neonatal pain relief in daily minor procedures. However, a varying degree of analgesic efficacy has been reported for this treatment. Environmental, maternal, and genetic factors may explain this variability and potentially allow for a personalized analgesic approach, maximizing therapeutic efficacy and preventing side effects. We investigated the exposome (ie, the set of clinical and anthropometric variables potentially affecting the response to the therapy) and the genetic variability of the noradrenaline transporter gene (solute carrier family 6 member 2 [ SLC6A2 ]) and 2 glucose transporter genes (solute carrier family 2 member 1 [ SLC2A1 ] and 2 [ SLC2A2 ]) in relation to the neonatal analgesic efficacy of a 33% glucose solution. The study population consisted in a homogeneous sample of more than 1400 healthy term newborns. No association for the exposome was observed, whereas a statistically significant association between the G allele of SLC2A1 -rs1105297 and a fourfold decreased probability of responding to the therapy was identified after multiple-testing correction (odds ratio of 3.98, 95% confidence interval 1.95-9.17; P = 4.05 × 10 -4 ). This allele decreases the expression of SLC2A1-AS1 , causing the upregulation of SLC2A1 in the dorsal striatum, which has been suggested to be involved in reward-related processes through the binding of opioids to the striatal mu-opioid receptors. Altogether, these results suggest the involvement of SLC2A1 in the analgesic process and highlight the importance of host genetics for defining personalized analgesic treatments.

Ferroptosis-related gene glutathione peroxidase 4 promotes reprogramming of glucose metabolism via Akt-mTOR axis in intrahepatic cholangiocarcinoma.

The role of the ferroptosis-related gene glutathione peroxidase 4 (GPX4) in oncology has been extensively investigated. However, the clinical implications of GPX4 in patients with intrahepatic cholangiocarcinoma (ICC) remain unknown. This study aimed to evaluate the prognostic impact of GPX4 and its underlying molecular mechanisms in patients with ICC. Fifty-seven patients who underwent surgical resection for ICC between 2010 and 2017 were retrospectively analyzed. Based on the immunohistochemistry, patients were divided into GPX4 high (n = 15) and low (n = 42) groups, and clinical outcomes were assessed. Furthermore, the roles of GPX4 in cell proliferation, migration and gene expression were analyzed in ICC cell lines in vitro and in vivo. The results from clinical study showed that GPX4 high group showed significant associations with high SUVmax on 18F-fluorodeoxyglucose-positron emission tomography (≥8.0, P = 0.017), multiple tumors (P = 0.004), and showed glucose transporter 1 (GLUT1) high expression with a trend toward significance (P = 0.053). Overall and recurrence-free survival in the GPX4 high expression group were significantly worse than those in the GPX4 low expression group (P = 0.038 and P < 0.001, respectively). In the experimental study, inhibition of GPX4 attenuated cell proliferation and migration in ICC cell lines. Inhibition of GPX4 also decreased the expression of glucose metabolism-related genes, such as GLUT1 or HIF1α. Mechanistically, these molecular changes are regulated in Akt-mechanistic targets of rapamycin axis. In conclusion, this study suggested the pivotal value of GPX4 serving as a prognostic marker for patients with ICC. Furthermore, GPX4 can mediate glucose metabolism of ICC.

Lauric acid epigenetically regulates lncRNA HOTAIR by remodeling chromatin H3K4 tri-methylation and modulates glucose transport in SH-SY5Y human neuroblastoma cells: Lipid switch in macrophage activation.

Lauric acid (LA) induces apoptosis in cancer and promotes the proliferation of normal cells by maintaining cellular redox homeostasis. Earlier, we postulated LA-mediated regulation of the NF-κB pathway by an epigenetic mechanism. However, the molecular mechanism and possible epigenetic events remained enigmatic. Herein, taking the lead from the alteration in cellular energetics in cancer cells upon LA exposure, we investigated whether LA exposure can epigenetically influence lncRNA HOTAIR, regulate glucose metabolism, and shift the cellular energetic state. Our results demonstrate LA induced modulation of lncRNA HOTAIR in a dose and time dependent manner. In addition, HOTAIR induces the expression of glucose transporter isoform 1 (GLUT1) and is regulated via NF-κB activation. Silencing HOTAIR by siRNA-mediated knockdown suppressed GLUT1 expression suggesting the key role of HOTAIR in LA-mediated metabolic reprogramming. Further, from our ChIP experiments, we observed that silencing HOTAIR subdues the recruitment of NF-κB on the GLUT1 (SLC2A1) promoter region. In addition, by performing western blot and immunocytochemistry studies, we found a dose dependent increase in Histone 3 Lysine 4 tri-methylation (H3K4me3) in the chromatin landscape. Taken together, our study demonstrates the epigenetic regulation in LA-treated SH-SY5Y cancer cells orchestrated by remodeling chromatin H3K4me3 and modulation of lncRNA HOTAIR that apparently governs the GLUT1 expression and regulates glucose uptake by exerting transcriptional control on NF-κB activation. Our work provides insights into the epigenetic regulation and metabolic reprogramming of LA through modulation of lncRNA HOTAIR, remodeling chromatin H3K4 tri-methylation, and shifting the energy metabolism in SH-SY5Y neuroblastoma cells.

SOX2 promotes vasculogenic mimicry by accelerating glycolysis via the lncRNA AC005392.2-GLUT1 axis in colorectal cancer.

Vasculogenic mimicry (VM), a new model of angiogenesis, fulfills the metabolic demands of solid tumors and contributes to tumor aggressiveness. Our previous study demonstrated the effect of SOX2 in promoting VM in colorectal cancer (CRC). However, the underlying mechanisms behind this effect remain elusive. Here, we show that SOX2 overexpression enhanced glycolysis and sustained VM formation via the transcriptional activation of lncRNA AC005392.2. Suppression of either glycolysis or AC005392.2 expression curbed SOX2-driven VM formation in vivo and in vitro. Mechanistically, SOX2 combined with the promoter of AC005392.2, which decreased H3K27me3 enrichment and thus increased its transcriptional activity. Overexpression of AC005392.2 increased the stability of GLUT1 protein by enhancing its SUMOylation, leading to a decrease in the ubiquitination and degradation of GLUT1. Accumulation of GLUT1 contributed to SOX2-mediated glycolysis and VM. Additionally, clinical analyses showed that increased levels of AC005392.2, GLUT1, and EPHA2 expression were positively correlated with SOX2 and were also associated with poor prognoses in patients with CRC. Our study conclusively demonstrates that the SOX2-lncRNA AC005392.2-GLUT1 signaling axis regulates VM formation in CRC, offering a foundation for the development of new antiangiogenic drugs or new drug combination regimens.

Pigment epithelium-derived factor (PEDF) represses the glucose transporter 1 (GLUT1) mRNA expression and may be a potential therapeutic agent in psoriasis: a case-control and experimental study.

We investigated the whole blood GLUT1 mRNA expression and serum pigment epithelium-derived factor (PEDF), interleukin-6 (IL-6), fetuin-A, and pentraxin-3 (PTX3) levels in psoriatic patients and tested their correlations with the severity of psoriasis using the psoriasis area and severity index (PASI) score. Also, we tested the GLUT1 mRNA expression after an in vitro treatment of human skin fibroblast (HSF) cell lines with PEDF. The case-control part of the study recruited 74 participants (44 psoriatic patients and 30 healthy volunteers). Whole blood GLUT1 mRNA fold changes were estimated by RT-PCR, and serum PEDF, IL-6, fetuin-A, and PTX3 levels were measured by ELISA kits. In the experimental part, the HSF cell lines were treated with different concentrations of PEDF for different times to test its effect on the GLUT1 mRNA expression. The whole blood GLUT 1 expression significantly increased in psoriatic patients and correlated positively with serum IL-6, fetuin-A, PTX3 levels and with the severity of psoriasis while negatively with serum PEDF levels. The PEDF-treated HSF cell lines showed a time- and dose-dependent decline in the GLUT 1 mRNA expression. The whole blood GLUT 1 mRNA is a non-invasive biomarker that is associated with the severity of psoriasis. PEDF represses GLUT 1 expression and may be a potential therapeutic agent in psoriasis.Trial registration: ClinicalTrials.gov Identifier: NCT04242082.

An arrayed CRISPR knockout screen identifies genetic regulators of GLUT1 expression.

Glucose, a primary fuel source under homeostatic conditions, is transported into cells by membrane transporters such as glucose transporter 1 (GLUT1). Due to its essential role in maintaining energy homeostasis, dysregulation of GLUT1 expression and function can adversely affect many physiological processes in the body. This has implications in a wide range of disorders such as Alzheimer's disease (AD) and several types of cancers. However, the regulatory pathways that govern GLUT1 expression, which may be altered in these diseases, are poorly characterized. To gain insight into GLUT1 regulation, we performed an arrayed CRISPR knockout screen using Caco-2 cells as a model cell line. Using an automated high content immunostaining approach to quantify GLUT1 expression, we identified more than 300 genes whose removal led to GLUT1 downregulation. Many of these genes were enriched along signaling pathways associated with G-protein coupled receptors, particularly the rhodopsin-like family. Secondary hit validation confirmed that removal of select genes, or modulation of the activity of a corresponding protein, yielded changes in GLUT1 expression. Overall, this work provides a resource and framework for understanding GLUT1 regulation in health and disease.

LncRNA FTO-IT1 promotes glycolysis and progression of hepatocellular carcinoma through modulating FTO-mediated N6-methyladenosine modification on GLUT1 and PKM2.

BACKGROUND: Long non-coding RNAs (LncRNAs) have been extensively studied to play essential roles in tumor progression. However, more in-depth studies are waiting to be solved on how lncRNAs regulate the progression of hepatocellular carcinoma (HCC). METHODS: Different expression levels of lncRNAs in HCC cells were compared by analysis of Gene Expression Omnibus and The Cancer Genome Atlas databases. The effects of lncRNA FTO Intronic Transcript 1 (FTO-IT1) on HCC cells were assessed by gain- and loss-of-function experiments. Colony formation assay, Edu assay, glucose uptake and lactic acid production assay were performed to evaluate the regulation of proliferation and glycolysis of HCC cells by FTO-IT1. The binding between protein interleukin enhancer binding factor 2/3 (ILF2/ILF3) and FTO-IT1 was determined by RNA pull-down, mass spectroscopy and RNA immunoprecipitation experiments. RNA stability assay, quantitative reverse transcription PCR and Western blot were employed to determine the regulatory mechanisms of FTO-IT1 on fat mass and obesity-associated (FTO). Methylated RNA immunoprecipitation assay was used to assessed the regulation of key enzymes of glycolysis by FTO. The role of FTO-IT1/FTO in vivo was confirmed via xenograft tumor model. RESULTS: LncRNA FTO-IT1, an intronic region transcript of FTO gene, was highly expressed in HCC and associated with poor prognosis of patients with HCC. FTO-IT1 was related to proliferation and glycolysis of HCC cells, and contributed to the malignant progression of HCC by promoting glycolysis. Mechanistically, FTO-IT1 induced stabilization of FTO mRNA by recruiting ILF2/ILF3 protein complex to 3'UTR of FTO mRNA. As a demethylase for N6-methyladenosine (m6A), FTO decreased m6A modification on mRNAs of glycolysis associated genes including GLUT1, PKM2, and c-Myc which alleviated the YTH N6-methyladenosine RNA binding protein 2 (YTHDF2)-mediated mRNA degradation. Therefore, the upregulated expression of FTO-IT1 leaded to overexpression of GLUT1, PKM2, and c-Myc by which enhanced glycolysis of HCC. Meanwhile, it was found that c-Myc transcriptional regulated expression of FTO-IT1 by binding to its promoter area under hypo-glucose condition, forming a reciprocal loop between c-Myc and FTO-IT1. CONCLUSIONS: This study identified an important role of the FTO-IT1/FTO axis mediated m6A modification of glycolytic genes contributed to glycolysis and tumorigenesis of HCC, and FTO-IT1 might be served as a new therapeutic target for HCC.

Intermedin alleviates diabetic vascular calcification by inhibiting GLUT1 through activation of the cAMP/PKA signaling pathway.

BACKGROUND AND AIMS: Vascular calcification (VC) is regarded as an independent risk factor for cardiovascular events in type 2 diabetic patients. Glucose transporter 1 (GLUT1) involves VC. Intermedin/Adrenomedullin-2 (IMD/ADM2) is a cardiovascular protective peptide that can inhibit multiple disease-associated VC. However, the role and mechanism of IMD in diabetic VC remain unclear. Here, we investigated whether IMD inhibits diabetic VC by inhibiting GLUT1. METHODS AND RESULTS: It was found that plasma IMD concentration was significantly decreased in type 2 diabetic patients and in fructose-induced diabetic rats compared with that in controls. Plasma IMD content was inversely correlated with fasting blood glucose level and VC severity. IMD alleviated VC in fructose-induced diabetic rats. Deficiency of Adm2 aggravated and Adm2 overexpression attenuated VC in high-fat diet-induced diabetic mice. In vitro, IMD mitigated high glucose-induced calcification of vascular smooth muscle cells (VSMCs). Mechanistically, IMD reduced advanced glycation end products (AGEs) content and the level of receptor for AGEs (RAGE). IMD decreased glucose transporter 1 (GLUT1) levels. The inhibitory effect of IMD on RAGE protein level was blocked by GLUT1 knockdown. GLUT1 knockdown abolished the effect of IMD on alleviating VSMC calcification. IMD receptor antagonist IMD17-47 and cyclic adenosine monophosphate/protein kinase A (cAMP/PKA) inhibitor H89 abolished the inhibitory effects of IMD on GLUT1 and VSMC calcification. CONCLUSIONS: These findings revealed that IMD exerted its anti-calcification effect by inhibiting GLUT1, providing a novel therapeutic target for diabetic VC.

Downregulation of the glucose transporter GLUT 1 in the cerebral microvasculature contributes to postoperative neurocognitive disorders in aged mice.

INTRODUCTION: Glucose transporter 1 (GLUT1) is essential for glucose transport into the brain and is predominantly expressed in the cerebral microvasculature. Downregulation of GLUT1 precedes the development of cognitive impairment in neurodegenerative conditions. Surgical trauma induces blood-brain barrier (BBB) disruption, neuroinflammation, neuronal mitochondria dysfunction, and acute cognitive impairment. We hypothesized that surgery reduces the expression of GLUT1 in the BBB that in turn disrupts its integrity and contributes to metabolic dysregulation in the brain that culminates in postoperative cognitive impairment. METHODOLOGY: Using an abdominal surgery model in aged WT mice, we assessed the perioperative changes in cognitive performance, tight junction proteins expression, GLUT1 expression, and the associated metabolic effects in the hippocampus. Thereafter, we evaluated the effects of these parameters in aged mice with conditional overexpression of GLUT1, and then again in aged mice with conditional overexpression of GLUT1 with or without prior exposure to the GLUT1 inhibitor ST-31. RESULTS: We showed a significant decline in cognitive performance, along with GLUT1 reduction and diminished glucose metabolism, especially in the ATP level in the postoperative mice compared with controls. Overexpression of GLUT1 expression alleviated postoperative cognitive decline and improved metabolic profiles, especially in adenosine, but did not directly restore ATP generation to control levels. GLUT1 inhibition ameliorated the postoperative beneficial effects of GLUT1 overexpression. CONCLUSIONS: Surgery-induced GLUT1 reduction significantly contributes to postoperative cognitive deficits in aged mice by affecting glucose metabolism in the brain. It indicates the potential of targeting GLUT1 to ameliorate perioperative neurocognitive disorders.

GLUT1 Mediates the Metabolic Reprogramming and Inflammation of CCR2$

BACKGROUND: Macrophages expressing CC chemokine receptor 2 (CCR2) possess characteristics and performance akin to M1 polarized macrophages, which promote inflammation. Advanced heart failure (HF) patients with higher abundance of CCR2+ macrophages are more likely to experience adverse remodeling. The precise mechanism of CCR2+ macrophages in how they affect the progression of dilated cardiomyopathy remains unknown. METHODS: Cardiac biopsy samples from dilated cardiomyopathy patients (DCM) were used for immunohistochemistry and immunofluorescence staining. PCR is employed to identify the IL-1ß, IL-6, TNF-α, TGF-ß, MMP2, MMP9, PKM1, PKM1, GLUT1, GLUT2, GLUT3, GLUT4, PDK1, PFKFB3, PFK1 and HK2 mRNA expression of CCR2+ monocytes/macrophages from the peripheral blood of DCM patients. Seahorse was used to evaluate the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) of CCR2+ monocytes/macrophages. 2-DG was used to simulate a lack of glucose. Lentivirus containing GLUT1 inhibitory sequence was used to knockdown GLUT1 gene expression of CCR2+ monocytes/macrophages. Western Blot and immunofluorescence staining was used to evaluate the expression of NLRP3. RESULTS: Immunostaining results of cardiac biopsy tissue from dilated cardiomyopathy (DCM) patients demonstrated that the progression to HF was associated with an increase in the number of CCR2+ macrophages. PCR results demonstrated that CCR2 monocytes and macrophages derived from the blood of DCM patients expressed elevated levels of inflammatory factors and up regulation of glycolysis related genes. In addition, OCR and glucose uptake experiments confirmed that increased glucose uptake of these cells was associated with greater inflammation and correlated with a worsening of cardiac function. limiting the glucose supply to CCR2+ monocytes and macrophages, or suppressing the activity of glucose transporter 1 (GLUT1) could reduce inflammation levels. CONCLUSIONS: These results suggest that CCR2+ monocytes and macrophages rely on metabolic reprogramming to trigger inflammatory response and contribute to myocardial injury and the progression of DCM.

Iron promotes glycolysis to drive colon tumorigenesis.

Colorectal cancer (CRC) is the third most common cancer and is also the third leading cause of cancer-related death in the USA. Understanding the mechanisms of growth and progression of CRC is essential to improve treatment. Macronutrients such as glucose are energy source for a cell. Many tumor cells exhibit increased aerobic glycolysis. Increased tissue micronutrient iron levels in both mice and humans are also associated with increased colon tumorigenesis. However, if iron drives colon carcinogenesis via affecting glucose metabolism is still not clear. Here we found the intracellular glucose levels in tumor colonoids were significantly increased after iron treatment. 13C-labeled glucose flux analysis indicated that the levels of several labeled glycolytic products were significantly increased, whereas several tricarboxylic acid cycle intermediates were significantly decreased in colonoids after iron treatment. Mechanistic studies showed that iron upregulated the expression of glucose transporter 1 (GLUT1) and mediated an inhibition of the pyruvate dehydrogenase (PDH) complex function via directly binding with tankyrase and/or pyruvate dehydrogenase kinase (PDHK) 3. Pharmacological inhibition of GLUT1 or PDHK reactivated PDH complex function and reduced high iron diet-enhanced tumor formation. In conclusion, excess iron promotes glycolysis and colon tumor growth at least partly through the inhibition of the PDH complex function.

Metabolic classification suggests the GLUT1/ALDOB/G6PD axis as a therapeutic target in chemotherapy-resistant pancreatic cancer.

Metabolic reprogramming is known as an emerging mechanism of chemotherapy resistance, but the metabolic signatures of pancreatic ductal adenocarcinomas (PDACs) remain unclear. Here, we characterize the metabolomic profile of PDAC organoids and classify them into glucomet-PDAC (high glucose metabolism levels) and lipomet-PDAC (high lipid metabolism levels). Glucomet-PDACs are more resistant to chemotherapy than lipomet-PDACs, and patients with glucomet-PDAC have a worse prognosis. Integrated analyses reveal that the GLUT1/aldolase B (ALDOB)/glucose-6-phosphate dehydrogenase (G6PD) axis induces chemotherapy resistance by remodeling glucose metabolism in glucomet-PDAC. Increased glycolytic flux, G6PD activity, and pyrimidine biosynthesis are identified in glucomet-PDAC with high GLUT1 and low ALDOB expression, and these phenotypes could be reversed by inhibiting GLUT1 expression or by increasing ALDOB expression. Pharmacological inhibition of GLUT1 or G6PD enhances the chemotherapy response of glucomet-PDAC. Our findings uncover potential metabolic heterogeneity related to differences in chemotherapy sensitivity in PDAC and develop a promising pharmacological strategy for patients with chemotherapy-resistant glucomet-PDAC through the combination of chemotherapy and GLUT1/ALDOB/G6PD axis inhibitors.