Evaluation Methods for Facilitative Glucose Transport in Cells and Their Applications

Yamamoto, Norio; Ashida, Hitoshi

doi:10.3136/fstr.18.493

Cited by 11 publications

(17 citation statements)

References 73 publications

(69 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…While most of the assays described in the earlier literature employed radioactive 2DG, radiolabeled analogs are costly and require specialized training and equipment. Described below is a rapid, inexpensive, and reproducible nonradioactive microplate assay for 2DG uptake into cultured cells (Yamamoto et al, 2006;Yamamoto and Ashida, 2012). The principle of the assay (Fig.…”

Section: Measurement Of Nonradiolabeled 2-deoxyglucose Uptakementioning

confidence: 99%

“…These indicators were not used extensively because their fluorescence is difficult to detect. However, recent developments in photomultiplier technology have now made the NBDGs more practical for use (Yamamoto and Ashida, 2012), with 2-NBDG shown to be of value for monitoring glucose uptake into a variety of mammalian cells. The 2-NBDG technique can be used in a microplate format (Leira et al, 2002), with flow cytometry (Zou et al, 2005), and with direct visualization using digital imaging fluorescence microscopy (Lloyd et al, 1999).…”

Section: Basic Protocolmentioning

confidence: 99%

“…To avoid the use of radioisotopes, in the 1990s Manchester and colleagues as well as Sasson and colleagues developed enzymatic assays for evaluating 2DG uptake into tissue samples and cultured cells (Yamamoto and Asida, ). These assays are based on the analysis of DG6P levels, which is accomplished by quantifying the NADPH produced during the G6PDH‐induced oxidation of DG6P to 6‐phosphoglucuronic acid.…”

Section: Commentarymentioning

confidence: 99%

See 2 more Smart Citations

Measurement of Glucose Uptake in Cultured Cells

Yamamoto¹,

Ueda-Wakagi

Sato³

et al. 2015

CP Pharmacology

Self Cite

115

View full text Add to dashboard Cite

Facilitative glucose uptake transport systems are ubiquitous in animal cells and are responsible for transporting glucose across cell surface membranes. Evaluation of glucose uptake is crucial in the study of numerous diseases and metabolic disorders such as myocardial ischemia, diabetes mellitus, and cancer. Detailed in this unit are laboratory methods for assessing glucose uptake into mammalian cells. The unit is divided into five sections: (1) a brief overview of glucose uptake assays in cultured cells; (2) a method for measuring glucose uptake using radiolabeled 3-O-methylglucose; (3) a method for measuring glucose uptake using radiolabeled 2-deoxyglucose (2DG); (4) a microplate method for measuring 2DG-uptake using an enzymatic, fluorometric assay; and (5) a microplate-based method using a fluorescent analog of 2DG.

show abstract

Section: Measurement Of Nonradiolabeled 2-deoxyglucose Uptakementioning

confidence: 99%

Section: Basic Protocolmentioning

confidence: 99%

Section: Commentarymentioning

confidence: 99%

See 1 more Smart Citation

Measurement of Glucose Uptake in Cultured Cells

Yamamoto¹,

Ueda-Wakagi

Sato³

et al. 2015

CP Pharmacology

Self Cite

115

View full text Add to dashboard Cite

show abstract

“…However, aerobic glycolysis in tumor cells may be limited by glucose uptake . Glucose derivatives labeled with various radioactive isotopes and fluorescent dyes have been used to monitor increased glucose uptake associated with a high glycolytic rate in vivo . These include 2‐deoxy‐D‐[1,2‐ 3 H]‐glucose, 2‐deoxy‐D‐[1‐ 14 C]‐glucose, 2‐deoxy‐2‐( 18 F)‐fluoro‐D‐glucose ( 18 F‐FDG) and the fluorescent probe 2‐[N‐(7‐nitrobenz‐2‐oxa‐1,3‐diaxol‐4‐yl)amino]‐2‐deoxyglucose (2‐NBDG) .…”

Section: Conventional Assessment Of Glycolysismentioning

confidence: 99%

Probing carbohydrate metabolism using hyperpolarized ¹³C‐labeled molecules

et al. 2018

View full text Add to dashboard Cite

Glycolysis is a fundamental metabolic process in all organisms. Anomalies in glucose metabolism are linked to various pathological conditions. In particular, elevated aerobic glycolysis is a characteristic feature of rapidly growing cells. Glycolysis and the closely related pentose phosphate pathway can be monitored in real time by hyperpolarized 13 C-labeled metabolic substrates such as 13 C-enriched, deuterated D-glucose derivatives, [2-13 C]-D-fructose, [2-13 C] dihydroxyacetone, [1-13 C]-Dglycerate, [1-13 C]-D-glucono-δ-lactone and [1-13 C] pyruvate in healthy and diseased tissues. Elevated glycolysis in tumors (the Warburg effect) was also successfully imaged using hyperpolarized [U-13 C 6 , U-2 H 7 ]-D-glucose, while the size of the preexisting lactate pool can be measured by 13 C MRS and/or MRI with hyperpolarized [1-13 C]pyruvate. This review summarizes the application of various hyperpolarized 13 C-labeled metabolites to the real-time monitoring of glycolysis and related metabolic processes in normal and diseased tissues. KEYWORDS dynamic nuclear polarization, glycolysis, hyperpolarized 13 C NMR, metabolic probes 1 | INTRODUCTION Glucose is a fundamental source of energy in living cells. It is present in all living organisms and utilized by both aerobic and anaerobic organisms. 1In eukaryotes, oxidation of glucose through glycolysis and the citric acid cycle (or tricarboxylic acid cycle, TCA cycle) yields energy in the form of ATP along with the release of carbon dioxide (CO 2 ) and water. Glycolysis, the breakdown of glucose, includes several reversible and three irreversible (committed) enzymatic reactions leading to the end-product pyruvate (Figure 1). 2,3 The enzymes of the three committed steps in glycolysis are allosterically controlled both positively and negatively. 4-8 Pyruvate is a key metabolic intermediate with many potential fates, including the production of carbohydrates through gluconeogenesis, fatty acids, amino acids or energy (ATP) via acetyl-CoA. Anomalies in glucose metabolism are linked to various pathological conditions, 9-11 so any method that could reliably monitor glucose metabolism in vivo not only would be valuable in fundamental studies of those diseases but could also be a valuable diagnostic biomarker. It has been widely documented that tumors and other rapidly proliferating cells have a dramatic increased rate of glucose uptake and lactate production even in the presence of adequate oxygen supply (the Warburg effect). 9,12,13 In addition, the expression levels of the mono-carboxylate transporters MCT1 and MCT4 are higher in cancer, and this facilitates the export of lactate from cancer cells. Although there is still an ongoing debate about why aerobic glycolysis is advantageous for tumor growth, in general cancer cells modulate glucose uptake as well as several glycolytic enzymes to match their energy demands by rapidly, albeit /journal/nbm 1 of 23 inefficiently, producing ATP in aerobic glycolysis, and to fulfill their increased demand for anabolic intermediates. [14][15]...

show abstract

“…3A). The uptake of 6-NBDG, a fluorescent glucose analog that is not further metabolized to enter the glycolytic pathway (21,43), was significantly reduced by S100B (Fig. 3B).…”

Section: Effects Of Recombinant S100b Protein On Insulin Activation Of Glycolysis In L6 Cellsmentioning

confidence: 99%

S100B impairs glycolysis via enhanced poly(ADP-ribosyl)ation of glyceraldehyde-3-phosphate dehydrogenase in rodent muscle cells

Hosokawa

Hamada

Fujiya

et al. 2017

American Journal of Physiology-Endocrinology and Metabolism

View full text Add to dashboard Cite

S100 calcium-binding protein B (S100B), a multifunctional macromolecule mainly expressed in nerve tissues and adipocytes, has been suggested to contribute to the pathogenesis of obesity. To clarify the role of S100B in insulin action and glucose metabolism in peripheral tissues, we investigated the effect of S100B on glycolysis in myoblast and myotube cells. Rat myoblast L6 cells were treated with recombinant mouse S100B to examine glucose consumption, lactate production, glycogen accumulation, glycolytic metabolites and enzyme activity, insulin signaling, and poly(ADP-ribosyl)ation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Glycolytic metabolites were investigated by enzyme assays or metabolome analysis, and insulin signaling was assessed by Western blot analysis. Enzyme activity and poly(ADP-ribosyl)ation of GAPDH was evaluated by an enzyme assay and immunoprecipitation followed by dot blot with an anti-poly(ADP-ribose) antibody, respectively. S100B significantly decreased glucose consumption, glucose analog uptake, and lactate production in L6 cells, in either the presence or absence of insulin. In contrast, S100B had no effect on glycogen accumulation and insulin signaling. Metabolome analysis revealed that S100B increased the concentration of glycolytic intermediates upstream of GAPDH. S100B impaired GAPDH activity and increased poly(ADP-ribosyl)ated GAPDH proteins. The effects of S100B on glucose metabolism were mostly canceled by a poly(ADP-ribose) polymerase inhibitor. Similar results were obtained in CC myotube cells. We conclude that S100B as a humoral factor may impair glycolysis in muscle cells independent of insulin action, and the effect may be attributed to the inhibition of GAPDH activity from enhanced poly(ADP-ribosyl)ation of the enzyme.

show abstract

Evaluation Methods for Facilitative Glucose Transport in Cells and Their Applications

Cited by 11 publications

References 73 publications

Measurement of Glucose Uptake in Cultured Cells

Measurement of Glucose Uptake in Cultured Cells

Probing carbohydrate metabolism using hyperpolarized ¹³C‐labeled molecules

S100B impairs glycolysis via enhanced poly(ADP-ribosyl)ation of glyceraldehyde-3-phosphate dehydrogenase in rodent muscle cells

Contact Info

Product

Resources

About

Evaluation Methods for Facilitative Glucose Transport in Cells and Their Applications

Cited by 11 publications

References 73 publications

Measurement of Glucose Uptake in Cultured Cells

Measurement of Glucose Uptake in Cultured Cells

Probing carbohydrate metabolism using hyperpolarized 13C‐labeled molecules

S100B impairs glycolysis via enhanced poly(ADP-ribosyl)ation of glyceraldehyde-3-phosphate dehydrogenase in rodent muscle cells

Contact Info

Product

Resources

About

Probing carbohydrate metabolism using hyperpolarized ¹³C‐labeled molecules