Academic literature on the topic 'Hexosamine biosynthesis pathway'
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Journal articles on the topic "Hexosamine biosynthesis pathway"
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Lockhart, Deborah E. A., Mathew Stanley, Olawale G. Raimi, David A. Robinson, Dominika Boldovjakova, Daniel R. Squair, Andrew T. Ferenbach, Wenxia Fang, and Daan M. F. van Aalten. "Targeting a critical step in fungal hexosamine biosynthesis." Journal of Biological Chemistry 295, no. 26 (April 27, 2020): 8678–91. http://dx.doi.org/10.1074/jbc.ra120.012985.
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Aspergillus fumigatus is a human opportunistic fungal pathogen whose cell wall protects it from the extracellular environment including host defenses. Chitin, an essential component of the fungal cell wall, is synthesized from UDP-GlcNAc produced in the hexosamine biosynthetic pathway. As this pathway is critical for fungal cell wall integrity, the hexosamine biosynthesis enzymes represent potential targets of antifungal drugs. Here, we provide genetic and chemical evidence that glucosamine 6-phosphate N-acetyltransferase (Gna1), a key enzyme in this pathway, is an exploitable antifungal drug target. GNA1 deletion resulted in loss of fungal viability and disruption of the cell wall, phenotypes that could be rescued by exogenous GlcNAc, the product of the Gna1 enzyme. In a murine model of aspergillosis, the Δgna1 mutant strain exhibited attenuated virulence. Using a fragment-based approach, we discovered a small heterocyclic scaffold that binds proximal to the Gna1 active site and can be optimized to a selective submicromolar binder. Taken together, we have provided genetic, structural, and chemical evidence that Gna1 is an antifungal target in A. fumigatus.
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Frank, L. A., M. L. Sutton-McDowall, D. L. Russell, X. Wang, D. K. Feil, R. B. Gilchrist, and J. G. Thompson. "Effect of varying glucose and glucosamine concentration in vitro on mouse oocyte maturation and developmental competence." Reproduction, Fertility and Development 25, no. 8 (2013): 1095. http://dx.doi.org/10.1071/rd12275.
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The effects of hyper- and hypo-glycaemic conditions during the in vitro maturation of mouse cumulus–oocyte complexes on developmental competence were examined, with an emphasis on the role of the hexosamine biosynthesis pathway. A low (1 mM) glucose concentration achieved optimal oocyte competence (3-fold higher blastocyst development rate compared with high (30 mM) glucose, P < 0.05). In addition, glucose supplementation during only the first hour after release from the follicle was necessary and sufficient to support oocyte maturation and embryo development to the blastocyst stage. Glucosamine (a known hyperglycaemic mimetic and specific activator of the hexosamine pathway) was able to substitute for glucose during this first hour, indicating that flux through the hexosamine pathway is essential for oocyte competence. In the absence of glucose throughout the maturation period, glucosamine was not able to increase developmental competence, and at higher concentrations (2.5 and 5 mM) had a detrimental effect on MII and blastocyst development rates, compared with controls (P < 0.05). These experiments underscore the importance of glucose metabolic pathways during in vitro maturation and support the concept that excess flux through the hexosamine pathway has detrimental consequences.
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FILIPPIS, Anthony, Stella CLARK, and Joseph PROIETTO. "Increased flux through the hexosamine biosynthesis pathway inhibits glucose transport acutely by activation of protein kinase C." Biochemical Journal 324, no. 3 (June 15, 1997): 981–85. http://dx.doi.org/10.1042/bj3240981.
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The hexosamine biosynthesis pathway and protein kinase C (PKC) activation mediate hyperglycaemia-induced impaired glucose transport, but the relative role of each pathway is unknown. Following a 2 h preincubation of rat adipocytes in the presence of either high glucose (30 mM) plus insulin (0.7 nM) or glucosamine (3 mM), both high glucose and glucosamine inhibited subsequent basal and insulin-stimulated glucose transport, measured at 5.0 mM glucose. Azaserine, an inhibitor of the enzyme glutamine:fructose-6-phosphate aminotransferase, abolished the effect of high glucose, but not that of glucosamine. Ro-31-8220, an inhibitor of PKC, reversed the effects of both high glucose and glucosamine, suggesting that flux through the hexosamine biosynthesis pathway impaired glucose transport acutely by activating PKC. Both high glucose and glucosamine caused a 3-fold increase in PKC activity; this effect of high glucose, but not that of glucosamine, was partially decreased by azaserine. Neither high glucose nor glucosamine altered basal or insulin-stimulated plasma membrane GLUT1 levels, whereas both treatments decreased basal, but not insulin-stimulated, GLUT4 levels. Azaserine abolished the effect of high glucose, but not that of glucosamine, on basal plasma membrane GLUT4 levels. Ro-31-8220, which returned glucose transport to control values, caused a further decrease in plasma membrane GLUT4 levels. It is concluded that, in rat adipocytes, an acute increase in flux through the hexosamine biosynthesis pathway inhibits glucose transport by activation of PKC.
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Filippis, Christine, Anthony Filippis, Stella Clark, and Joseph Proietto. "Activation of PI 3-kinase by the hexosamine biosynthesis pathway." Molecular and Cellular Endocrinology 194, no. 1-2 (August 2002): 29–37. http://dx.doi.org/10.1016/s0303-7207(02)00213-7.
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Willems, Anke P., Baziel G. M. van Engelen, and Dirk J. Lefeber. "Genetic defects in the hexosamine and sialic acid biosynthesis pathway." Biochimica et Biophysica Acta (BBA) - General Subjects 1860, no. 8 (August 2016): 1640–54. http://dx.doi.org/10.1016/j.bbagen.2015.12.017.
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Przybyl, Joanna, Angela Tolwani, Sushama Varma, and Matt van de Rijn. "Abstract B015: Targeting hexosamine biosynthesis pathway for the treatment of desmoid tumors." Clinical Cancer Research 28, no. 18_Supplement (September 15, 2022): B015. http://dx.doi.org/10.1158/1557-3265.sarcomas22-b015.
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Abstract Cancer cells rewire metabolic pathways and energy production to support the enhanced proliferation, invasion and resistance to treatment. The three main glucose metabolism pathways that support growth of cancer cells are: a) the glycolysis pathway for energy production; b) the pentose phosphate pathway for biomass production; and c) the hexosamine biosynthesis pathway (HBP) for protein glycosylation. It is known that the activation of HBP leads to altered glycosylation of oncogenes, transcription factors and kinases in many types of cancer. These aberrations may lead to increased proliferation and survival of tumor cells, and may be associated with resistance to therapy. A better understanding of the role of HBP in malignancies has the potential for clinical implications. Several studies demonstrated that pharmacological inhibition of GFPT2 (glutamine-fructose-6-phosphate transaminase 2, the first and rate-limiting enzyme in HBP) and the enzymes that act downstream of HBP may exhibit anti-tumorigenic effect both in vitro and in vivo, and may modulate sensitivity to chemo-, radio- and immunotherapy. Most of these studies focused on carcinomas and the role of HPB in sarcoma has not been studied. We recently reported a remarkable enrichment of genes involved in HBP in a subset of leiomyosarcoma (LMS) and demonstrated that expression of GFPT2 in LMS is associated with poor clinical outcome. We identified the c-Myc oncoprotein as a potential target of HPB that may be stabilized by aberrant glycosylation in LMS. Here we show the results of a large-scale screening of 260 primary specimens of 33 types of soft tissue lesions. In addition to expression in a subset of LMS, we observed near universal expression of GFPT2 in 34 of 35 desmoid type fibromatosis (DTF), independent of the mutation type of the CTNNB1 gene. Gene Set Enrichment Analysis of a previously published 3SEQ transcriptomic dataset composed of DTF and 9 other types of fibrotic lesions identified significant enrichment of other genes implicated in HBP and multiple glycosylation-associated pathways in DTF compared to the other types of fibrotic lesions. Our analysis identified ATF6 (activating transcription factor 6) as a possible target regulated by aberrant glycosylation as a consequence of HBP activation in DTF. ATF6 is a glycoprotein that has been demonstrated to underlie the resistance to chemotherapy in osteosarcoma, to have a pro-oncogenic role in primary liver cancers and has been proposed as a therapeutic target in cystic fibrosis. Others have shown that targeting HBP can provide therapeutic benefit in a number of preclinical models of carcinoma. Our studies offer new insights into the mechanisms of DTF tumorigenesis and, when confirmed by in vitro studies, will provide a rationale to explore the potential of therapeutic targeting of HBP in DTF. Citation Format: Joanna Przybyl, Angela Tolwani, Sushama Varma, Matt van de Rijn. Targeting hexosamine biosynthesis pathway for the treatment of desmoid tumors [abstract]. In: Proceedings of the AACR Special Conference: Sarcomas; 2022 May 9-12; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(18_Suppl):Abstract nr B015.
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Filhoulaud, Gaëlle, Ghislaine Guillemain, and Raphaël Scharfmann. "The Hexosamine Biosynthesis Pathway Is Essential for Pancreatic Beta Cell Development." Journal of Biological Chemistry 284, no. 36 (July 7, 2009): 24583–94. http://dx.doi.org/10.1074/jbc.m109.025288.
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Adeyemo, Adelola O., Dulce H. Gomez, Maitha A. Aldokhayyil, and Michael D. Brown. "Is The Hexosamine Biosynthesis Pathway Affected By Inflammation In Endothelial Cells?" Medicine & Science in Sports & Exercise 52, no. 7S (July 2020): 567. http://dx.doi.org/10.1249/01.mss.0000680392.11336.c3.
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Beriault, D., Y. Shi, and G. Werstuck. "Investigating the Role of the Hexosamine Biosynthesis Pathway in Diabetic Atherosclerosis." Canadian Journal of Cardiology 29, no. 10 (October 2013): S254. http://dx.doi.org/10.1016/j.cjca.2013.07.417.
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Penque, Brent A., April M. Hoggatt, B. Paul Herring, and Jeffrey S. Elmendorf. "Hexosamine Biosynthesis Impairs Insulin Action via a Cholesterolgenic Response." Molecular Endocrinology 27, no. 3 (March 1, 2013): 536–47. http://dx.doi.org/10.1210/me.2012-1213.
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Abstract Plasma membrane cholesterol accumulation has been implicated in cellular insulin resistance. Given the role of the hexosamine biosynthesis pathway (HBP) as a sensor of nutrient excess, coupled to its involvement in the development of insulin resistance, we delineated whether excess glucose flux through this pathway provokes a cholesterolgenic response induced by hyperinsulinemia. Exposing 3T3-L1 adipocytes to physiologically relevant doses of hyperinsulinemia (250pM–5000pM) induced a dose-dependent gain in the mRNA/protein levels of 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGR). These elevations were associated with elevated plasma membrane cholesterol. Mechanistically, hyperinsulinemia increased glucose flux through the HBP and O-linked β-N-acetylglucosamine (O-GlcNAc) modification of specificity protein 1 (Sp1), known to activate cholesterolgenic gene products such as the sterol response element-binding protein (SREBP1) and HMGR. Chromatin immunoprecipitation demonstrated that increased O-GlcNAc modification of Sp1 resulted in a higher binding affinity of Sp1 to the promoter regions of SREBP1 and HMGR. Luciferase assays confirmed that HMGR promoter activity was elevated under these conditions and that inhibition of the HBP with 6-diazo-5-oxo-l-norleucine (DON) prevented hyperinsulinemia-induced activation of the HMGR promoter. In addition, both DON and the Sp1 DNA-binding inhibitor mithramycin prevented the hyperinsulinemia-induced increases in HMGR mRNA/protein and plasma membrane cholesterol. In these mithramycin-treated cells, both cortical filamentous actin structure and insulin-stimulated glucose transport were restored. Together, these data suggest a novel mechanism whereby increased HBP activity increases Sp1 transcriptional activation of a cholesterolgenic program, thereby elevating plasma membrane cholesterol and compromising cytoskeletal structure essential for insulin action.
Dissertations / Theses on the topic "Hexosamine biosynthesis pathway"
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Al-Oanzi, Ziad Hail. "The role of the hexosamine biosynthesis pathway in control of hepatic glucose metabolism." Thesis, University of Newcastle upon Tyne, 2013. http://hdl.handle.net/10443/2186.
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Type 2 diabetes is associated with increased hepatic lipogenesis and glucose production. Enzymes of lipogenesis are co-ordinately induced by insulin and glucose. However, the enzyme glucose 6-phosphatase (G6Pc), which catalyses the final reaction in hepatic glucose production is repressed by insulin but induced by glucose and is markedly elevated in type 2 diabetes. Another gene that is repressed by insulin and induced by glucose in muscle is thioredoxin interacting protein (TXNIP), which is abnormally elevated in muscle in type 2 diabetes. TXNIP gene regulation in liver has not been reported. The induction of hepatic lipogenic enzymes by glucose is attributed to the transcription factor ChREBP-Mlx, whereas the glucose-induction of G6Pc is attributed to covalent modification of FOXO transcription factors by O- GlcNAc formed by the hexosamine biosynthesis pathway (HBP). The aim of this thesis was to investigate the role of the HBP in regulation by glucose of G6Pc and TXNIP gene expression in hepatocytes. This thesis investigated three commonly used methods to modulate HBP flux and covalent modification by O-GlcNAc. (1) An inhibitor of glutamine:fructose 6- phosphate amidotransferase (6-diazo-5-oxo-l-norleucine, DON), the rate limiting enzyme of the HBP, was established to be a valid tool to study glucose-regulated gene expression. (2) Substrates that enter the HBP after GFAT, such as glucosamine which is widely used to demonstrate links between HBP or O-GlcNAc modification and insulin resistance were shown to be invalid tools. (3) Inhibitors of O-GlcNAc modification or expression of O-GlcNAc transferase were of limited use to alter protein modification by O-GlcNAc. Glucose caused a larger induction of G6Pc and TXNIP mRNA in the absence of insulin than in its presence, and this induction could be largely accounted for by Mlx- dependent mechanisms (supporting involvement of ChREBP or MondoA) and by FOXO transcription factors. G6Pc and TXNIP expression were confirmed to be regulated by distinct mechanisms based on the induction of TXNIP but not G6Pc by the glucose analogue, 2-deoxyglucose, through an Mlx-independent mechanism. Insulin caused rapid translocation of both FOXO1 and FOXO3A from the nucleus to the cytoplasm. Both glucose and 2-deoxyglucose opposed the translocation of FOXO1 and FOXO3A by insulin, and they stimulated translocation of FOXO3A to the nucleus in the absence of insulin. Inhibition of HBP flux with the GFAT inhibitor had the following effects: (i) counteraction of the glucose-induction of both G6Pc and TXNIP mRNA; (ii) counteraction of glucose-induced translocation of ChREBP to the nucleus without affecting the signalling metabolite, fructose 2,6-bisphosphate; (iii) counteraction of glucose-induced translocation of FOXO1 and FOXO3A to the nucleus. A role for O- GlcNAc modification of both ChREBP and FOXO3A was supported by wheat-germ agglutinin precipitation. The results of this thesis support involvement of both HBP flux and O-GlcNAc modification of ChREBP and FOXO3A in glucose-regulated expression of G6Pc and FOXO3A but they do not support a role for glucosamine as an experimental tool to study glucose-induced insulin resistance.
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Rajamani, Uthra. "Hyperglycemia-induced activation of the hexosamine biosynthetic pathway causes myocardial cell death." Thesis, Stellenbosch : University of Stellenbosch, 2009. http://hdl.handle.net/10019.1/1142.
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Thesis (PhD (Physiological Sciences))--University of Stellenbosch, 2009.ENGLISH ABSTRACT: OBJECTIVE – Oxidative stress increases flux through the hexosamine biosynthetic pathway (HBP) resulting in greater O-GlcNAcylation of target proteins. Since increased oxidative stress and HBP flux are associated with insulin resistance, we hypothesized that its activation leads to greater O-GlcNAcylation of BAD (pro-apoptotic) and increased myocardial apoptosis. RESEARCH DESIGN AND METHODS – To investigate our hypothesis, we employed two experimental models: 1) H9c2 cardiomyoblasts exposed to high glucose (33 mM glucose) ± HBP modulators ± antioxidant treatment vs. matched controls (5.5 mM glucose); and 2) a rat model of high fat diet-induced insulin resistance and hyperglycemia. We evaluated apoptosis in vitro by Hoechst nuclear staining, Annexin-V staining, caspase activity measurements and immunoblotting while in vivo apoptosis was assessed by immunoblotting. In vitro reactive oxygen species (ROS) levels were quantified by H2DCFDA staining (fluorescence microscopy, flow cytometry). We determined overall and BAD O-GlcNAcylation, both by immunoblotting and immunofluorescence microscopy. As BAD-Bcl-2 dimer formation enhances apoptosis, we performed immunoprecipitation analysis and immunofluorescence microscopy (co-localization) to determine BAD-cl-2 dimerization. In vivo overall O-GlcNAcylation, BAD O-GlcNAcylation and BAD-Bcl-2 dimerization was determined by immunoprecipitation and immunoblotting. 4 RESULTS – High glucose treatment of cells significantly increased the degree of apoptosis as revealed by Hoechst nuclear staining (54 ± 9%, p<0.01 vs. 5.5 mM), Annexin-V staining (43 ± 5%), caspase activity assay (26 ± 2%) and immunoblotting. In parallel, overall OGlcNAcylation (p<0.001 vs. 5.5 mM), BAD O-GlcNAcylation (p<0.05 vs. 5.5 mM) and ROS levels were increased (fluorescence microscopy – p<0.05 vs. 5.5 mM; flow cytometry – p<0.001 vs. 5.5 mM). HBP inhibition using DON and antioxidant treatment (α-OHCA) attenuated these effects while HBP activation by PUGNAc exacerbated it. Likewise, insulin resistant rat hearts exhibited significantly higher caspase-3 (p<0.05 vs. controls), overall O-GlcNAcylation (p<0.05 vs. controls) and BAD O-GlcNAcylation levels (p<0.05 vs. 5.5 mM). BAD-Bcl-2 dimer formation was increased in cells exposed to hyperglycemia [immunoprecipitation analysis and co-localization] and in insulin resistant hearts. CONCLUSIONS - Our study identified a novel pathway whereby hyperglycemia results in greater oxidative stress, resulting in increased HBP activation and increased BAD OGlcNAcylation. We also found greater BAD-Bcl-2 dimerization increasing myocardial apoptosis, suggesting that this pathway may play a crucial role in the onset of the diabetic cardiomyopathy.
AFRIKAANSE OPSOMMING: DOELWIT – Oksidatiewe stres verhoog fluks deur die heksosamien biosintetiese weg (HBW) wat in „n groter O-GlcNAsetilering van teiken proteïene resulteer. Weens die feit dat verhoogde oksidatiewe stres en HBW fluks verband hou met insulienweerstandigheid, hipotetiseer ons dat die aktivering hiervan tot groter O-GlcNAsetilering van BAD (pro-aptoptoties) en verhoogde miokardiale apoptose lei. NAVORSINGS ONTWERP EN METODES – Om die hipotese te ondersoek het ons twee modelle ontplooi: 1) H9c2 kardiomioblaste is blootgestel aan hoë glukose konsentrasie (33mM glucose) ± HBW moduleerders ± antioksidant behandeling vs. gepaarde kontrole (5.5mM glucose); en 2) „n hoë vet dieetgeïnduseerde insulienweerstandige rotmodel en hiperglukemie. Ons het apoptose in vitro deur middel van Hoescht nukleuskleuring geëvalueer, kasapase aktiwiteit bepalings en immunoblotting terwyl apoptose in vivo getoets is deur immunoblotting. Reaktiewe suurstofspesie (RSS) vlakke is deur middel van H2DCFDA verkleuring (fluoresensie mikroskopie, vloeisitometrie) bepaal. Algehele en BAD O-GlcNAsetilering is beide deur immunoblotting en immunofluoresensie mikroskopie bepaal. BAD-Bcl-2 dimeervorming bevorder apoptose, om BAD-cl-2 dimerisasie te bepaal is daar van immunopresipitering analise en immunofluoresensie mikroskopie (ko-lokalisasie) gebruik gemaak. In vivo is algehele OGlcNAsetiliering, BAD O-GlcNAsetiliering en BAD-Bcl-2 dimerisasie deur immunopresipitasie en immunoblotting bepaal. 6 RESULTE – Hoë glukose behandeling van selle het die graad van apotpose betekenisvol verhoog soos blootgelê deur Hoechst nukleuskleuring (54 ± 9%, p<0.01 vs. 5.5 mM), Annexin-V kleuring (43 ± 5%), kaspase aktiviteit assay (26 ± 2%) en immunoblotting. In parallel, algehele OGlcNAsetilering (p<0.001 vs. 5.5 mM), BAD O-GlcNAsetilering (p<0.05 vs. 5.5 mM) en RSS vlakke is verhoog (fluoresensie mikroskopie– p<0.05 vs. 5.5 mM; vloeisitometrie– p<0.001 vs. 5.5 mM). HBW inhibering deur van DON en van antioksidant behandeling gebruik te maak (α- OHCA) het hierdie effekte verlaag terwyl HBW aktivering deur PUGNAc dit verhoog het. Netso, het insulienweerstandige rotharte betekenisvolle hoë kaspase -3 (p<0.05 vs. kontrole), algeheel O-GlcNAsetilering (p<0.05 vs. kontrole) en BAD O-GlcNAsetiliering vlakke (p<0.05 vs. 5.5 mM) getoon. BAD-Bcl-2 dimeervorming is verhoog in hiperglukemies blootgestelde selle [immunopresipitering analise en ko-lokalisering] en in insulienweerstandige harte. GEVOLGTREKKINGS – Ons studie het „n nuwe weg geïdenifiseer waar hiperglukemie in groter oksidatiewe stres resulteer wat weer HBW aktivering verhoog en BAD O-GlcNAsetilering verhoog het. Ons het verder bevind dat groter BAD-Bcl-2 dimerisasie miokardiale apoptose verhoog wat voorstel dat hierdie weg „n belangrike rol in diabetiese kardiomiopatie speel.
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Williams, Gordon. "Increased hexosamine biosynthetic pathway flux impairs myocardial GLUT4 translocation." Thesis, Stellenbosch : University of Stellenbosch, 2009. http://hdl.handle.net/10019.1/2893.
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Thesis (MSc (Physiological Sciences))--University of Stellenbosch, 2009.Aims and Background: According to the World Health Organization type 2 diabetes will constitute a major global burden of disease within the next few decades. In agreement, reports show that rapid urbanization and lifestyle changes in South Africa are major factors responsible for these projections. Therefore, any perturbations that alter the regulatory steps that control myocardial glucose uptake by the cardiac-enrich glucose transporter, GLUT4, will lead in the development of diabetic cardiomyopathy and cardiac hypertrophy. Although considerable efforts are been put into unraveling molecular mechanisms underlying this process, less is known regarding the spatio-temporal regulation of GLUT4. In light of this, our specific aim was to establish in vitro fluorescence microscopy- and flow cytometry-based models for visualization and assessment of myocardial GLUT4 translocation using H9c2 cardiac-derived myoblasts. After successful establishment of our in vitro-based model for myocardial GLUT4 translocation, our second aim was to determine the role of the hexosamine biosynthetic pathway (HBP) in this process. Here, we employed HBP modulators to alter flux and subsequently evaluate its effect on myocardial GLUT4 translocation. To further strengthen our hypothesis, we also investigated the role of the HBP in hearts of an in vivo type 2 diabetes mouse model. Hypothesis: We hypothesize that increased flux through the HBP impairs myocardial GLUT4 translocation by greater O-linked glycosylation of the insulin signaling pathway, ultimately leading to myocardial insulin resistance. Methods: Rat cardiac-derived H9c2 myoblasts were cultured until ~ 80-90 % confluent for 3 days and thereafter subcultured in Lab-Tek chamber slides (~ 15, 000 cells per well) for 24 hours. Cells were then serum starved for 3 hours by insulin administration of 100 nM for 0, 5 and 30 minutes, respectively. We employed a method to quantify the relative proportion of GLUT4 at the sarcolemma using immunofluorescence microscopy- and flow cytometry-based models for visualization and assessment of myocardial GLUT4 translocation. Using these methods we investigated the role HBP have during GLUT4 translocation. The HBP were then activated through the following: a) high glucose and glutamine concentrations; b) low glucose and glucosamine stimulation; and c) over-expression of the HBP rate- limiting enzyme, i.e. GFAT. Subsequently, cardiac-derived myoblasts were fixed and probed for ~ 24 hours with antibodies specific for intracellular- and membrane-bound GLUT4, anti-myc GLUT4 (9E10) and O-GlcNAc. To assess GLUT4 translocation and O-GlcNAcylation we employed the following secondary antibodies: FITC Green for intracellular-bound GLUT4; and b) Texas Red for membrane-bound GLUT4 (immunofluorescence microscopy) and Phycoerythrin for flow cytometry-based model. Cells were thereafter viewed by multi-dimension imaging using an inverted system microscope (Olympus IX81) and a BD FACS Aria cell sorter for flow cytometric analysis. We also assessed HBP in an in vivo context by probing heart tissue - from insulin resistant db/db mice - with a GFAT monoclonal antibody. Results: The db/db mouse represents an ideal model to confirm our hypothesis in an in vivo context. In agreement, our preliminary results show increased GFAT expression versus heterozygous db/+ controls. Our in vitro model show myocardial GLUT4 translocation at 5 minute peak response when H9c2 cardiac-derived myoblasts were stimulated with 100 nM insulin, and GLUT4 vesicles return to normal after longer insulin stimulatory times (10, 15 and 30 minutes. Myocardial Glut4 v translocation was impaired when cells were stimulated with 100 nM wortmannin. Our transfection based model (immunofluorescence microscopy- and flow cytometry-based models) confirms 5 minute peak response under real time conditions. High glucose concentration (25 mM glucose), glucosamine concentrations (2.5 mM, 5 mM, and 10 mM) and over-expression of GFAT led to an impairment of myocardial GLUT4 translocation. Employment of an HBP activator (50 μM PUGNAc) also caused impairment of myocardial GLUT4 translocation. Myocardial GLUT4 translocation was restored when cells were treated with an HBP inhibitor (40 μM DON). High glucose concentrations (25 mM glucose), glucosamine concentrations (2.5 mM, 5 mM, and 10 mM) and over-expression of GFAT resulted in an increase in O-GlcNAcylation. HBP activation (50 μM PUGNAc) showed an increase in O-GlcNAcylation, while administration of 40 μM DON reversed this effect. Discussion and conclusion: We successfully established an in vitro experimental system to assay myocardial GLUT4 translocation. Our data show that dysregulated flux through the HBP impairs myocardial GLUT4 translocation. It is likely that the HBP becomes dysregulated during the pre-diabetic/early diabetic state and that O-GlcNAcylation of members of the insulin signaling pathway occurs during this stage. This will lead to myocardial insulin resistance, and in the long term, will contribute to the onset of the diabetic cardiomyopathy. Investigations to find unique inhibitors of this maladaptive pathway should therefore result in the development of novel therapeutic agents that will lead to a reduction in the growing global burden of disease for type 2 diabetes and associated cardiovascular diseases.
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Sylvain-Drolet, Guillaume. "Transcriptional regulation of mortalin and the hexosamine biosynthetic pathway by the orphan nuclear receptor ERRalpha." Thesis, McGill University, 2011. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=97252.
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The orphan nuclear receptor estrogen-related receptor α (ERRα, NR3B1) plays a major role in transcription regulation of metabolic genes. Its role in glycolysis regulation is well known. However, we ignore yet if it could be implicated in a signaling pathway connected to glycolysis, the Hexosamine Biosynthetic Pathway (HBP). The HBP is an important energy and nutrient-sensing pathway, which modulates O-linked N-acetylglucosamine (O-GlcNAc) post-translational modification. We demonstrate here that ERRα is involved in transcriptional regulation of HBP genes. ERRα can be localized to promoters of several HBP genes, such as Ogt and Oga. Both encode two enzymes that directly modulate the O-GlcNAc cycling. Moreover, ERRα activates HBP genes transcription in collaboration with its coactivator PGC-1α. In ERRα-null mice, the HBP genes expression is downregulated. However, we can observe more O-GlcNAcylated proteins in absence of ERRα. This was demonstrated more specifically on the mitochondrial chaperone Mortalin. Mortalin is encoded by Hspa9, and we show that this gene is an ERRα target. Moreover, Mortalin is much more O-GlcNAcylated in absence of the ERRα. This suggests that in addition to Mortalin transcriptional regulation, ERRα is involved in Mortalin post-translational modification through regulation of the HBP.Le récepteur nucléaire orphelin relié à l'estrogène (ERRα, NR3Β1) joue un rôle majeur dans la régulation de gènes métaboliques. Son rôle dans la régulation de la glycolyse est très bien connu. Cependant, nous ignorons encore s'il pourrait être impliqué dans la régulation d'une voie de signalisation directement liée à la glycolyse, la voie de signalisation des hexosamines (HBP). La HBP est une importante voie de signalisation servant de détecteur d'énergie et de nutriments et régulant la modification post-traductionelle O-lié N-acétylglucosamine (O-GlcNAc). Nous démontrons qu'ERRα est impliqué dans la régulation des gènes de HBP. ERRα se lie aux promoteurs de plusieurs gènes de HBP, comme Ogt et Oga, codant pour deux enzymes qui régulent directement le cycle des O-GlcNAc. De plus, ERRα active la transcription des gènes de HBP en collaboration avec son coactivateur PGC-1α. Dans les souris déficientes pour le gène ERRα, l'expression des gènes de HBP se trouve réduite. Cependant, nous pouvons observer plus de protéines O-GlcNAcylées en absence d'ERRα. Ceci a été démontré plus spécifiquement sur la chaperone mitochondriale Mortalin. Mortalin est codée par le gène Hspa9 et nous démontrons que ce gène est une cible d'ERRα. De plus, Mortalin est beaucoup plus O-GlcNAcylée en absence d'ERRα. Ceci suggère qu'en plus de réguler la transcription de Mortalin, ERRα est impliqué dans la régulation des modifications post-traductionelles de Mortalin, en régulant la voie de signalisation des hexosamines.
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Imbriolo, Jamie. "The hexosamine biosynthetic pathway induces gene promoter activity of the cardiac-enriched isoform of acetyl-CoA carboxylase." Thesis, Stellenbosch : Stellenbosch University, 2013. http://hdl.handle.net/10019.1/79957.
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Thesis (PhD)--Stellenbosch University, 2013.ENGLISH ABSTRACT: The cardiac isoform of acetyl-CoA carboxylase (ACCβ) produces malonyl-CoA, a potent inhibitor of mitochondrial fatty acid (FA) uptake; thus increased ACCβ activity decreases fatty acid utilization thereby potentially leading to intracellular myocardial lipid accumulation and insulin resistance (IR). Previous studies show that greater flux through the hexosamine biosynthetic pathway (HBP) contributes to the development of IR. In light of this, we hypothesize that increased HBP flux induces ACCβ gene expression thereby contributing to the onset of IR. Our initial work focused on ACCβ gene promoter regulation and suggest that the HBP modulates upstream stimulatory factor 2 (USF2) thereby inducing ACCβ gene expression. Here, we further investigated HBP-mediated regulation of ACCβ gene expression by transiently transfecting cardiac-derived H9c2 cells with an expression vector encoding the rate-limiting HBP enzyme (GFAT) ± the full length ACCβ and 4 truncated promoter-luciferase constructs, respectively. GFAT overexpression increased ACCβ gene promoter activity for the full length and 3 larger deletion constructs (p<0.001 vs. controls). However, GFAT-mediated and USF2-mediated ACCβ promoter induction was blunted when co-transfected with the -38/+65 deletion construct suggesting that USF2 binds to the proximal promoter region (near start codon). Further investigation proves that USF2 binds to ACCβ promoter and activates it, but that USF2 is not O-GlcNAc modified even though there is a strong correlation between increased O-GlcNac levels and USF2 activation of ACCβ. This would suggest that there is another O-GlcNac modified factor involved in this regulatory pathway. Our study demonstrates that increased HBP flux induces ACCβ gene promoter activity via HBP modulation of USF2. We propose that ACCβ induction reduces fatty acid oxidation, thereby leading to intracellular lipid accumulation (FA uptake>>FA oxidation) and the onset of cardiac IR.
AFRIKAANSE OPSOMMING: Die kardiale isoform van asetiel-CoA karboksilase (ACCβ) produseer maloniel-CoA, ‘n kragtige inhibeerder van mitochondriale vetsuur (VS) opname, en om hierdie rede sal verhoogde ACCβ aktiwiteit, vetsuur gebruik verlaag en potensieël aanleiding gee tot intrasellulêre miokardiale lipiedophoping en insulienweerstand (IW). Vorige studies toon dat groter fluks deur die heksosamienbiosintetiese weg (HBW) bydra tot die ontwikkeling van IW. In die lig hiervan hipotetiseer ons dat verhoogde HBW fluks, ACCβ geenuitdrukking induseer, en sodoende tot die onstaan van IW bydra. Ons aanvanglike werk het op ACCβ geenpromotorregulering gefokus, en voorgestel dat die HBW die opstroom stimuleringsfaktor 2 (USF2) moduleer en dus ACCβ geen uitdrukking induseer. Hier het ons verder die HBW-gemedieërde regulering van ACCβ-geenuitdrukking deur kortstondige tranfeksie van kardiaalverkrygde H9c2 selle met ‘n uitdrukkingsvektor wat kodeer vir die tempo-bepalende HBW ensiem (GFAT) ± die volle lengte ACCβ, en vier afgestompte promotor-lusiferase konstrukte onderskeidelik, te ondersoek. GFAT ooruidrukking het ACCβ geenpromotor aktiwiteit vir die volle lengte, en drie groter uitwissingskonstrukte verhoog (p<0.001 vs. kontrole). Hoewel GFAT- en USF2-gemedieërde ACCβ promotorinduksie tydens ko-transfeksie van die -38/+65 uitwissingskonstruk versag was, is dit voorgestel dat USF2 aan die proksimale promotor area (naby die beginkodon) bind. Verdere ondersoek bewys ook dat USF2 aan die ACCβ promotor bind en dit aktiveer, maar dat USF2 nie O-GlcNAc gemodifiseer word nie ten spyte van ‘n sterk korrelasie tussen verhoogde O-GlcNac vlakke en USF2 aktivering van ACCβ. Dit kan dus voogestel word dat daar ‘n alternatiewe O-GlcNac gemodifiseerde faktor betrokke is in hierdie reguleringsweg. Ons studie demonstreer dat verhoogde HBW fluks ACCβ geenpromotor aktiwiteit via HBW modulering van USF2 veroorsaak. Ons stel voor dat ACCβ induksie vetsuuroksidasie verlaag en so tot intrasellulêre lipiedophoping (VS opname >> VS oksidasie) en die onstaan van kardiale IW lei.
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Harris, E. R. (Eurinah Roberta). "Does the hexosamine biosynthetic pathway play a role in mediating the beneficial effects of oleic acid in the heart?" Thesis, Stellenbosch : Stellenbosch University, 2012. http://hdl.handle.net/10019.1/20124.
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Thesis (MSc)--Stellenbosch University, 2012.ENGLISH ABSTRACT: Background:Obesity is a growing global burden; current studies have projected the prevalence of obese / overweight individuals to increase to ~1.35 billion by 2030. A number of factors contribute to cardiovascular diseases, of which the focus of this study is what effect an increased level of free fatty acids has on the flux through the hexosamine biosynthetic pathway (HBP). It has been widely proven that an increased flux through the HBP causes an increase in protein O-GlcNAcylation, which leads to increased reactive oxygen species (ROS) production as well as an increase in cell death (apoptosis). Methods: For the purpose of this study a cell model was used. H9c2 cardiomyoblasts were cultured in 5ml Dulbecco‟s Modified Eagles Medium (DMEM) supplemented with 10% foetal bovine serum and 1% penicillin-streptomycin. The cells were then exposed to 0.25mM monounsaturated fatty acid (oleic acid) for 24, 48 and 72 hours respectively. The cultured cells were then evaluated to assess the degree ROS production, overall O-GlcNAcylation and cell death (apoptosis and necrosis), using flow cytometry and immunofluorescence microscopy. Results: We found that oleic acid causes a significant decrease in ROS production at the 48 hour time point when analysed on the flow cytometer, which indicates that oleic acid is metabolized by the cells in a independent manner. Oleic acid also caused a significant decrease in cell death at all the time intervals. With regard to the HBP, oleic acid activates this pathway but causes downstream cardioprotective effects that do not necessarily occur along this pathway. Conclusion: This study explored whether a monounsaturated fatty acid, oleic acid, is able to act as a novel cardioprotective agent. The in vitro data supports this concept and we showed that it is able to blunt oxidative stress and cell death. It was also found that although oleic acid activated the HBP, it did not mediate its protective effects via this pathway only.
AFRIKAANSE OPSOMMING: Agtergrond: Vetsug is 'n groeiende wêreldlas; huidige studies voorspel dat die voorkoms van vetsugtige / oorgewig individue toe sal neem tot ~1.35 biljoen teen 2030. Alhoewel verskeie faktore tot kardiovaskulêre siektes bydra is die fokus van hierdie studie om die effek van verhoogde vryvetsuurvlakke op die fluks deur die heksosamienbiosintestiese weg (HBW) te ondersoek. Dit is reeds bewys dat verhoogde fluks deur die HBW 'n verhoging in proteïen O-GlcNAsilering lei, wat verder tot verhoogde reaktiewe suusrtofspesies (ROS) vorming aanleiding gee en ook seldood (apoptose) verhoog. Metodes:'n Selmodel is vir die doel van hierdie studie gebruik. H9c2 kardiomioblaste is in 5ml Dulbecco's Modified Eagles Medium (DMEM) gekweek en gesupplementeer met 10% fetale beesserum en 1% penisillien-streptomysien. Die selle is blootgestel aan 'n 0.25mM mono onversadigde vetsuur (oleïensuur ) vir 24, 48 en 72 uur onderskeidelik. Die gekweekte selle is gevolglik ondersoek vir die graad van ROS ontwikkeling, algehele O-GlcNAsilering en seldood (apoptosis en nekrose), deur van vloeisitometrie en immunofluoresensie mikroskopie gebruik te maak. Resultate: Ons het bevind dat oleïensuur 'n betekenisvolle verlaging in ROS ontwikkeling teen 48 uur soos bepaal deur die vloeisitometer, veroorsaak. Dit wys daarop dat oleïensuur deur die selle op 'n onafhanklike wyse gemetaboliseer is. Oleïensuur het ook 'n betekenisvolle verlaging in seldood by alle tydsintervalle veroorsaak. Met betrekking tot die HBW het oleïensuur hierdie weg geaktiveer maar afstroom kardiobeskermings effekte versoorsaak wat nie noodwendig langs hierdie weg onstaan nie. Gevolgtrekking:Hierdie studie het die moontlikheid van 'n mono-onversadige vetsuur, oleïensuur, om op te tree as 'n nuwe kardiobeskermingsmiddel ondersoek. Die in vitro data ondersteun hierdie konsep en hier is aangetoon dat dit wel oksidatiewe stres en seldood onderdruk. Daar is verder bevind dat alhoewel oleïensuur die HBW aktiveer dit nie die beskermings effekte alleenlik via hierdie weg medieer nie.
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Imbriolo, Jamie. "Increased flux through the hexosamine biosynthetic pathway leads to the induction of acetol-CoA caboxylase gene expression in the heart." Thesis, Stellenbosch : Stellenbosch University, 2008. http://hdl.handle.net/10019.1/21459.
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Thesis (MSc)--Stellenbosch University, 2008.ENGLISH ABSTRACT: Gene expression of the cardiac isoform of acetyl-CoA carboxylase (ACCb) is induced in a glucose-dependent manner. ACCb produces malonyl-CoA, a potent inhibitor of mitochondrial fatty acid uptake. Previous studies show that increased flux through the hexosamine biosynthetic pathway (HBP) under hyperglycaemic conditions may contribute to the development of insulin resistance. In light of this, we hypothesised that increased HBP flux induces cardiac ACCb gene expression thereby contributing to the onset of insulin resistance. We tested our hypothesis by transiently transfecting cardiac-derived rat H9c2 myoblasts with a 1,317 bp human ACCb promoter-luciferase construct (pPIIb-1317) and an expression construct encoding the rate-limiting step of the HBP i.e. glutamine: fructose 6-phosphate amidotransferase (GFAT). Overexpression of GFAT increased ACCb gene promoter activity by 75 ± 23% versus controls (n=6, p<0.001). When cotransfection experiments were repeated in the presence of varying concentrations of L-glutamine (0 mM, 4 mM, 8 mM), a substrate for the HBP, ACCb promoter activity was dose-dependently increased. To further corroborate these findings, we employed two inhibitors of GFAT, i.e. 40 μM azaserine and 40 μM 6-diazo-5-oxo-Lnorleucine were administered to transfected cells for a period of 24 hours. Here both azaserine and 6-diazo-5-oxonorleucine attenuated ACCb gene promoter activity. In agreement, co-transfections with two dominant negative GFAT constructs also diminished ACCb gene promoter activity. We next inhibited two enzymes of the HBP acting downstream of GFAT, i.e. O-GlcNAc transferase and O-GlcNAcase using alloxan (0.1 mM, 1 mM and 2 mM) and streptozotocin (5 mM and 10 mM), respectively, for a period of 24 hours. Addition of alloxan attenuated ACCb gene promoter activity by 35.6 ± 1.9% (n=16, p<0.001) and streptozotocin increased activity by 32 ± 12% (n=12, p<0.001). We also investigated USF1 and USF2 as transcriptional regulatory candidates for HBP-induced ACCβ promoter regulation. Our data implicates USF2 as an important transcriptional regulator of HBP-induced ACCβ promoter regulation. In summary, this study demonstrates that increased flux through the hexosamine biosynthetic pathway induces ACCb gene promoter activity. We further propose that such an induction would reduce cardiac fatty acid oxidation, thereby leading to intracellular lipid accumulation due to a mismatch between sarcolemmal FA uptake and mitochondrial FA oxidation in the insulin resistant setting (i.e. hyperlipidaemia).
AFRIKAANSE OPSOMMING: Geen uitdrukking van die kardiale isoform asetiel-KoA karboksilase (ACCb) word in ‘n glukose afhanklike wyse geïnduseer. ACCb produseer maloniel-KoA, ‘n kragtige inhibeerder van mitochondriale vetsuuropname. Vorige studies toon aan dat verhoogde fluks deur die heksosamien biosintestiese weg (HBW) onder hiperglukemiese toestande bydra tot die ontwikkeling van insulienweerstand. In die lig hiervan, word daar gehipotetiseer dat verhoogde HBP fluks kardiale ACCb geenuitdrukking induseer en so bydra tot die ontstaan van insulienweerstand. Ons hipotese is getoets deur die kardiale afkomstige rot H9c2 mioblaste met ‘n 1.317 bp mens ACCb-lusiferase promotor konstruk (pPII-1317) te transfekteer en ‘n uitdrukking te konstrueer wat die tempo bepalende stap van HBP i.e. glutamien: fruktose-6-fosfaat amidotransferase (GFAT) kodeer. Ooruitdrukking van GFAT verhoog ACCb geenpromotor aktiviteit deur 75 ± 23% teenoor kontrole (n=6, p<0.001). Die herhaling van ko-transfeksie eksperimente is herhaal in die teenwoordigheid van variëerbare L-glutamienkonsentrasies (0 mM, 4 mM, 8 mM), ’n substraat vir die HBP, ACCb promotor aktiwiteit is dosisafhanglik verhoog. Om die bevindinge verder te staaf, is twee inhibeerders van GFAT, i.e. 40 μM azaserien en 40 μM 6-diazo-5-oxo-L-norleusien aan transfeksie selle toegedien vir ’n tydperk van 24 uur. Beide azaserien en 6-diazo-5-oxo-L-norleusien verlaag ACCb geenpromotor aktiwiteit. In ooreenstemming met die bogenoemde het ko-transfeksies met twee dominante negatiewe GFAT konstrukte ook ACCb geenpromoter aktiwiteit verminder. Die volgende stap is om twee ensieme van die HBP wat stroomaf van GFAT aktief is, vir ‘n periode van 24 uur te inhibeer i.e. O-GlcNAc transferase en O-GlcNAcase deur alloxan (0.1 mM, 1 mM en 2 mM) and streptozotosien (5 mM en 10 mM) onderskeidelik vir ‘n 24 uur periode te gebruik. Toevoeging van alloxan het die ACCb geenpromotor aktiwiteit by 35.6 ± 1.9% (n=16, p<0.001) verlaag en streptozotosien aktiwiteit verhoog by 32 ± 12% (n=12, p<0.001). Ons het ook die USF1 en USF2 as transkripsie regulerings kandidate vir HBP-geïnduseerde ACCβ promotor regulering ondersoek. Ons data impliseer dat USF2 as ‘n belangrike transkripsie reguleerder van HBP-geïndiseerde ACCβ promotor regulering is. Samevattend het hierdie studie demonstreer dat verhoogde fluks deur die hexosamien biosintetiese weg ACCb geenpromotor aktiwiteit induseer. Ons stel verder voor dat hierdie induksie die kardiale vetsuuroksidasie verlaag wat daartoe lei dat intrasellulêre lipied akkumulasie as gevolg van onparing tussen sarkolemma vetsuuropname en mitochondriale vetsuuroksidasie in ’n insulien weerstandige situasie (i.e. hiperlipidaemia).
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Otto, Delita. "Exploring underlying mechanisms driving the onset of stress-induced insulin resistance." Thesis, Stellenbosch : Stellenbosch University, 2012. http://hdl.handle.net/10019.1/20125.
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Thesis (MSc)--Stellenbosch University, 2012.ENGLISH ABSTRACT: Physical and psychological stressors trigger activation of the hypothalamo-pituitary-adrenocortical (HPA) axis that leads to enhanced secretion of glucocorticoids e.g. cortisol. Moreover, chronic activation of this pathway may elevate oxidative stress that is linked to the onset of insulin resistance and cardiovascular diseases (CVD). Our laboratory previously found that oxidative stress increases flux through metabolic circuits such as the hexosamine biosynthetic pathway (HBP), in effect increasing its modification of target proteins post-transcriptionally with O-GlcNAc moeities. This in turn may alter protein function and contribute to the onset of myocardial insulin resistance and impaired contractile function. Since the underlying mechanisms linking chronic stress to cardiometabolic pathophysiology are poorly understood, we hypothesised that cortisol elicits myocardial oxidative stress, HBP activation, and decreased glucose uptake (due to attenuated glucose transport functionality) with detrimental outcomes, i.e. insulin resistance and apoptosis. To investigate this hypothesis we established an in vitro model using HL-1 cardiomyocytes, with which we evaluated the degree of O-GlcNAcylation and oxidative stress in response to a range of time-dose treatments with dexamethasone (synthetic glucocorticoid). Glucose transporter 4 (GLUT4) translocation to the sarcolemma was also assessed. In agreement with the literature, results suggest that GLUT4 translocation is significantly decreased subsequent to dexamethasone treatment. Although no significant differences were observed with regards to oxidative stress or O-GlcNAcylation, the data show that dexamethasone increased the latter with a maximal effect after two hours exposure to the 10-6 M dose. Although our results were not conclusive, the data suggest a potential novel link between dexamethasone exposure, HBP activation and decreased GLUT4 translocation. Based on our findings we propose that detrimental effects of chronic stress on the heart may be mediated by increased HBP flux. Given that glucocorticoid excess and GLUT4 dysregulation have been associated with insulin resistance (and related metabolic derangements and diseases), these results provide new targets for potential therapeutic agents.
AFRIKAANSE OPSOMMING: Fisiese sowel as psigologiese stressors veroorsaak die aktivering van die hipotalamiese-hipo seale-bynier (HHB) pad wat lei tot die verhoogde sekresie van glukokortikoïede soos kortisol. Kroniese aktivering van hierdie pad kan ook oksidatiewe stres verhoog wat weer tot insulienweerstandigheid en kardiovaskulêre siektes (KVS) kan lei. Navorsing uit ons laboratorium het voorheen bewys dat oksidatiewe stres 'n toename in vloei deur metaboliese paaie soos die heksoamine biosintetiese pad (HBP) kan veroorsaak deur die modi sering van teikenproteïene met O-GlcNAc motiewe. Dit kan weer proteïen funksie verander en bydra tot die ontstaan van miokardiale insulienweerstandigheid en verswakte kontraktiele funksie. Die onderliggende meganismes wat kroniese stres aan kardiometaboliese pato siologie verbind word nog nie goed verstaan nie, daarom is ons hipotese dat kortisol miokardiale oksidatiewe stres veroorsaak, die HBP pad aktiveer, en glukose opname verminder (deur die funksionele onderdrukking van glukose transport), wat nadelige uitkomste soos insulienweerstandigheid en apoptose tot gevolg kan hê. Om hierdie hipotese te ondersoek, is 'n in vitro model van HL-1 kardiomiosiete gebruik waarmee die graad van O-GlcNAsilering en oksidatiewe stres in reaksie op 'n reeks tyd-konsentrasie behandelings met deksametasoon (sintetiese glukokortikoïed), bepaal is. Glukose transporter 4 (GLUT4) translokasie na die sarkolemma is ook geasseseer. In ooreenstemming met die literatuur, is GLUT4 translokasie insiggewend onderdruk tydens deksometasoon behandeling. Alhoewel geen insiggewende verskille rakende oksidatiewe stres en O-GlcNAsilering gevind is nie, het ons data aangedui dat laasgenoemde deur deksametasoon vermeerder het na twee ure van blootstelling aan die 10-6 M konsentrasie. Alhoewel ons resultate geen afdoende bewys lewer nie, stel dit wel voor dat daar 'n potensiële verbintenis tussen deksametasoon behandeling en 'n afname in GLUT4 translokasie is. Gebasseer op ons bevindings, stel ons voor dat die nadelige e ekte van kroniese stres op die hart bemiddel kan word deur 'n toename in vloei deur die HBP. Gegewe dat 'n oormaat glukokortikoïede en GLUT4 wanregulering geassosieer is met insulien weerstandigheid (en verbandhoudende metaboliese veranderinge en siektes), verskaf hierdie resultate nuwe teikens vir potensiële terapeutiese ingrepe.
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Frank, Laura Alice. "The role of the hexosamine biosynthesis pathway and β-O-linked glycosylation in determining oocyte developmental competence." Thesis, 2012. http://hdl.handle.net/2440/96463.
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Maternal diabetes and conditions such as obesity in which blood glucose levels are elevated are associated with reduced fertility and poor pregnancy outcomes. Many studies have examined the effects of hyperglycaemia on the early embryo and fetus; however, it is becoming increasingly evident that the peri-conceptual environment surrounding the oocyte has a significant impact on developmental competence and the long-term health of offspring. In this thesis, I aimed to investigate the role of the hexosamine biosynthesis pathway (HBP) in oocyte developmental competence. The HBP is a glucose-metabolising pathway which can also be upregulated by glucosamine, a potent hyperglycaemic mimetic which enters the HBP downstream of the rate-limiting enzyme. The HBP produces uridine diphosphate-N acetylglucosamine, which can be used for the β-O-linked glycosylation (O-GlcNAcylation) of proteins, regulating their function in a similar manner to phosphorylation. Firstly I established the effect of hyper- and hypo-glycaemic conditions during in vitro maturation (IVM) of mouse cumulus-oocyte complexes (COCs) on a range of measures associated with oocyte developmental competence, including cumulus expansion, meiotic maturation, cleavage and blastocyst development rates. A low (1 mM) glucose concentration achieved optimal oocyte competence, and glucose supplementation during only the first hour of IVM was necessary and sufficient to support oocyte maturation and embryo development to the blastocyst stage. Glucosamine was able to substitute for glucose during this first hour. In the absence of glucose throughout IVM, glucosamine was not able to increase developmental competence, and at higher concentrations (2.5 and 5 mM) had a detrimental effect on these outcomes. These experiments underscored the importance of the other glucose metabolic pathways, during COC maturation, and supported the concept that excess flux through the HBP has detrimental consequences. Using Western blots and immunohistochemistry, it was shown that both glucosamine and high glucose levels induced an increase in total O-GlcNAcylation in COCs, which was reduced in the presence of an inhibitor of the β-O-linked glycosyltransferase enzyme. Several specific proteins were identified using mass spectrometry as potential targets of O-GlcNAcylation in COCs, including heat-shock protein 90 (HSP90, both α and β isoforms). While glucosamine treatment of COCs significantly decreased blastocyst development rate, inhibiting HSP90 with 17-allylamino-17-demethoxygeldanamycin during IVM in the presence of glucosamine recovered blastocyst rates to control levels. This effect was not due to an increase in overall HSP90 levels, since inhibiting HSP90 in control COCs did not affect blastocyst rate. These results suggest O-GlcNacylated HSP90 has an aberrant function in the COC. This study is the first to examine in detail O-GlcNAcylation levels in the COC, and their correlation to oocyte developmental competence. HSP90 was identified as a potential target of O-GlcNAcylation in the COC, and subsequently shown to mediate oocyte developmental competence. This research is significant because of the increasing numbers of women wishing to become pregnant who have high blood glucose levels due to diabetes, obesity or poor diet. I have generated critically needed knowledge towards understanding how these lifestyle factors affect fertility and identifying possible avenues for new therapies.Thesis (Ph.D.) -- University of Adelaide, School of Paediatrics and Reproductive Health, 2012
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Lin, Tzu, and 林芷. "The role of Hexosamine Biosynthetic Pathway in Adipocyte Differentiation." Thesis, 2008. http://ndltd.ncl.edu.tw/handle/97175327786173568976.
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碩士高雄醫學大學
醫學遺傳學研究所
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Obesity is a rapidly growing global threat against public health. The World Health Organisation (WHO) has defined obesity as a condition with excessive fat accumulation in the body. Increasing adipose tissue relates in adipokines secretion and involves in development of metabolic complications. However, regulation of adipocyte differentiation and adipokines secretion still need further clarified. O-linked N acetyl-glucosamine (O-GlcNAc) modification has been implicated in regulation of signaling pathway, cell function, cell cycle and gene expression in several cell types. Hexosamine biosynthetic pathway (HBP) generates the sugar nucleotide UDP-GlcNAc, which is the donor for O-GlcNAc modification of nucleocytoplasmic proteins. When the GFAT-1 (rate limiting enzyme of HBP) was overexpressed in mice, the mice developed fat cell hypertrophy and excess weight. The aim of this study is to investigate the pathophysiological role of HBP and protein O-GlcNAc modification on adipocyte by observing the expression of genes involving in early stage of differentiation and adipokines affecting late stage complications. Our results demonstrate that the O-GlcNAc modified proteins are increased in the process of 3T3-L1 preadipocyte differentiation since very early stage after induction. Genes involve in early stage of differentiation such as SREBP-1c, PPAR-γ, C/EBP-α, C/EBP-β, and C/EBP-δ are upregulated. Adiponectin, visfatin, apelin, retinol-binding protein-4 and angiotensinogen mRNAs are increased in mature adipocytes. However, blocking HBP by AZA or DON can decrease the end product of HBP—O-GlcNAc modification and prevents the differentiation of 3T3-L1 cells. Decreasing HBP activity also diminishes gene expressions which are necessary for adipocyte differentiation in early stage and prevents mRNA overexpression of above adipokines. The results suggest that HBP activation and change of protein O-GlcNAc modification maybe a novel pathway mediating differentiation and adipokine expression in adipocytes.
Book chapters on the topic "Hexosamine biosynthesis pathway"
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Le Minh, Giang, and Mauricio J. Reginato. "Role of Hexosamine Biosynthetic Pathway on Cancer Stem Cells: Connecting Nutrient Sensing to Cancer Cell Plasticity." In Reference Module in Biomedical Sciences. Elsevier, 2021. http://dx.doi.org/10.1016/b978-0-12-820472-6.00038-4.
Full textConference papers on the topic "Hexosamine biosynthesis pathway"
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Zhang, Weiruo, Gina Bouchard, Alice Yu, Majid Shafiq, Mehran Jamali, Joseph Shrager, Kelsey Ayers, et al. "Abstract 4403: FDG uptake in human lung adenocarcinoma associated with invasion through the hexosamine biosynthesis pathway." In Proceedings: AACR Annual Meeting 2017; April 1-5, 2017; Washington, DC. American Association for Cancer Research, 2017. http://dx.doi.org/10.1158/1538-7445.am2017-4403.
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Sharma, Nikita S., Vineet K. Gupta, Roey Hadad, Bhuwan Giri, Anthony Ferrantella, Vikas Dudeja, Ashok K. Saluja, and Sulagna Banerjee. "Abstract A59: Targeting the glutamine dependent hexosamine biosynthesis pathway sensitizes pancreatic tumors to anti-PD1 therapy." In Abstracts: AACR Special Conference on Tumor Immunology and Immunotherapy; November 27-30, 2018; Miami Beach, FL. American Association for Cancer Research, 2020. http://dx.doi.org/10.1158/2326-6074.tumimm18-a59.
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Efimova, Elena V., Oliver K. Appelbe, Steve Seung-Young Lee, Natalia Ricco, Aishwarya Ramamarthy, Nicolas Rymut, Tamica Collins, Donald J. Wolfgeher, Sara Warrington, and Stephen J. Kron. "Abstract 3505: Tumor radiosensitization by suppression of hexosamine biosynthetic pathway." In Proceedings: AACR Annual Meeting 2018; April 14-18, 2018; Chicago, IL. American Association for Cancer Research, 2018. http://dx.doi.org/10.1158/1538-7445.am2018-3505.
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Wu, Chang-Chih, Thomas Lynch, Joseph Moloughney, Aixa Navia, Olufunmilola Ibironke, Po-Chien Chou, Nicole M. Vega-Cotto, et al. "Abstract 2441: mTOR complex 2 modulates glycosylation of CD147 via the hexosamine biosynthetic pathway." In Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA. American Association for Cancer Research, 2014. http://dx.doi.org/10.1158/1538-7445.am2014-2441.
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Jacinto, Estela, Joseph Moloughney, Thomas Lynch, Chang-Chih Wu, Olufunmilola Ibironke, Aixa Navia, Po-Chien Chou, Sisi Zhang, Joshua Rabinowitz, and Guy Werlen. "Abstract 1150: mTORC2 enhances flux through the hexosamine biosynthetic pathway by regulation of GFAT1 expression." In Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA. American Association for Cancer Research, 2015. http://dx.doi.org/10.1158/1538-7445.am2015-1150.
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Saeui, Christopher T., and Kevin Yarema. "Abstract LB-054: Small molecule inhibitors of the hexosamine biosynthetic pathway and the cancer O-GlcNAcome." In Proceedings: AACR 107th Annual Meeting 2016; April 16-20, 2016; New Orleans, LA. American Association for Cancer Research, 2016. http://dx.doi.org/10.1158/1538-7445.am2016-lb-054.
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Chiaradonna, F., F. Ricciardiello, G. Votta, R. Palorini, I. Raccagni, L. Brunelli, L. De Gioia, R. Pastorelli, RM Moresco, and B. La Ferla. "PO-259 Inhibition of the hexosamine biosynthetic pathway by targeting PGM3 causes breast cancer growth arrest and apoptosis." In Abstracts of the 25th Biennial Congress of the European Association for Cancer Research, Amsterdam, The Netherlands, 30 June – 3 July 2018. BMJ Publishing Group Ltd, 2018. http://dx.doi.org/10.1136/esmoopen-2018-eacr25.291.
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Taparra, Kekoa, Hailun Wang, Katriana Nugent, Reem Malek, Jen Groves, Gokben Yildirir, Brian Simons, Dean Felsher, Natasha Zachara, and Phuoc Tran. "Abstract 1054: SNAI1 regulates the hexosamine biosynthetic pathway to promote tumorigenesis and oncogene-induced senescence escape in lung cancer." In Proceedings: AACR 107th Annual Meeting 2016; April 16-20, 2016; New Orleans, LA. American Association for Cancer Research, 2016. http://dx.doi.org/10.1158/1538-7445.am2016-1054.
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Bryant, Jerry, Archito T. Tamayo, Richard Mendez, John Lee, Changping Li, David J. Yang, David Rollo, Richard J. Ford, and Lan V. Pham. "Abstract 218: The hexosamine biosyntheic pathway and O-linked glycosylation for targeted therapy in diffuse large B-cell lymphoma." In Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL. American Association for Cancer Research, 2012. http://dx.doi.org/10.1158/1538-7445.am2012-218.
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