Measurement of Glucose Uptake in Cultured Cells

Yamamoto, Norio; Ueda-Wakagi, Manabu; Sato, Takuya; Kawasaki, Kengo; Sawada, Keisuke; Kawabata, Kyuichi; Akagawa, Mitsugu; Ashida, Hitoshi

doi:10.1002/0471141755.ph1214s71

Cited by 115 publications

(77 citation statements)

References 50 publications

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“…3 H(G) (2-NBDG) (Molecular Probes, Life Technologies, Eugene, OR) in RPMI 1640 (Gibco, Life Technologies, Frederick, MD), as previously described (30). After 30 min of incubation (378C under 95% humidity, 5% CO 2 , and atmospheric O 2 levels), the reaction was stopped by adding cold 13 PBS, and fluorescence was read (l ex = 485 nm; l em = 538 nm) (Fluoroskan Ascent; Thermo Fisher Scientific, Madrid, Spain).…”

Section: Cellular Glucose Uptakementioning

confidence: 99%

Beneficial Effect of Pistachio Consumption on Glucose Metabolism, Insulin Resistance, Inflammation, and Related Metabolic Risk Markers: A Randomized Clinical Trial

et al. 2014

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OBJECTIVETo examine whether a pistachio-rich diet reduces the prediabetes stage and improves its metabolic risk profile. RESEARCH DESIGN AND METHODSPrediabetic subjects were recruited to participate in this Spanish randomized clinical trial between 20 September 2011 and 4 February 2013. In a crossover manner, 54 subjects consumed two diets, each for 4 months: a pistachio-supplemented diet (PD) and a control diet (CD). A 2-week washout period separated study periods. Diets were isocaloric and matched for protein, fiber, and saturated fatty acids. A total of 55% of the CD calories came from carbohydrates and 30% from fat, whereas for the PD, these percentages were 50 and 35%, respectively (including 57 g/day of pistachios). RESULTSFasting glucose, insulin, and HOMA of insulin resistance decreased significantly after the PD compared with the CD. Other cardiometabolic risk markers such as fibrinogen, oxidized LDL, and platelet factor 4 significantly decreased under the PD compared with the CD (P < 0.05), whereas glucagon-like peptide-1 increased. Interleukin-6 mRNA and resistin gene expression decreased by 9 and 6%, respectively, in lymphocytes after the pistachio intervention (P < 0.05, for PD vs. CD). SLC2A4 expression increased by 69% in CD (P = 0.03, for PD vs. CD). Cellular glucose uptake by lymphocytes decreased by 78.78% during the PD (P = 0.01, PD vs. CD). CONCLUSIONSChronic pistachio consumption is emerging as a useful nutritional strategy for the prediabetic state. Data suggest that pistachios have a glucose-and insulin-lowering effect, promote a healthier metabolic profile, and reverse certain metabolic deleterious consequences of prediabetes.

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Section: Cellular Glucose Uptakementioning

confidence: 99%

Beneficial Effect of Pistachio Consumption on Glucose Metabolism, Insulin Resistance, Inflammation, and Related Metabolic Risk Markers: A Randomized Clinical Trial

et al. 2014

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“…The application of 2‐NBDG as a fluorescent tracer for assessing GLUT‐mediated sugar uptake is a well‐established method . To evaluate the cellular uptake mechanism of the title compounds, we performed a competitive 2‐NBDG cell uptake assay using the aminodeoxypyranoside‐conjugated platinum(II) complexes compared with oxaliplatin in HT29 cells using glucose‐free culture medium.…”

Section: Resultsmentioning

confidence: 99%

Methyl 6‐Amino‐6‐deoxy‐d‐pyranoside‐Conjugated Platinum(II) Complexes for Glucose Transporter (GLUT)‐Mediated Tumor Targeting: Synthesis, Cytotoxicity, and Cellular Uptake Mechanism

Gao

Yang³

et al. 2016

ChemMedChem

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Methyl 6-aminodeoxy-d-pyranoside-derived platinum(II) glycoconjugates were designed and synthesized based on the clinical drug oxaliplatin for glucose transporter (GLUT)-mediated tumor targeting. In addition to a substantial improvement in water solubility, the conjugates exhibited cytotoxicity similar to or higher than that of oxaliplatin in six different human cancer cell lines. GLUT-mediated transport of the complexes was investigated with a cell-based fluorescence competition assay and GLUT-inhibitor-mediated cytotoxicity analysis in a GLUT-overexpressing human colorectal adenocarcinoma (HT29) cell line. The antitumor effect of the aminodeoxypyranoside-conjugated platinum(II) complexes was found to depend significantly on the GLUT inhibitor, and the cellular uptake of the molecules was regulated by GLUT-mediated transport. The results from this study demonstrate the potential advantages of aminodeoxypyranosides as sugar motifs for glycoconjugation for Warburg-effect-targeted drug design. These fundamental results also support the potential of aminodeoxypyranoside-conjugated platinum(II) complexes as lead compounds for further preclinical evaluation.

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“…Cells were incubated with 1 mmol/l 2-deoxy-D-glucose for 5 min at 37°C/5% CO 2 before washing (PBS), lysing with 0.1 mol/l NaOH, and snap freezing. Glucose uptake was measured by the fluorescence method of Yamamoto et al [23].…”

Section: Methodsmentioning

confidence: 99%

Evaluation of anti-insulin receptor antibodies as potential novel therapies for human insulin receptoropathy using cell culture models

2018

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Aims/hypothesis Bi-allelic loss-of-function mutations in the INSR gene (encoding the insulin receptor [INSR]) commonly cause extreme insulin resistance and early mortality. Therapeutic options are limited, but anti-INSR antibodies have been shown to activate two mutant receptors, S323L and F382V. This study evaluates four well-characterised murine anti-INSR monoclonal antibodies recognising distinct epitopes (83-7, 83-14, 18-44, 18-146) as surrogate agonists for potential targeted treatment of severe insulin resistance arising from insulin receptoropathies. Methods Ten naturally occurring mutant human INSRs with defects affecting different aspects of receptor function were modelled and assessed for response to insulin and anti-INSR antibodies. A novel 3T3-L1 adipocyte model of insulin receptoropathy was generated, permitting conditional knockdown of endogenous mouse Insr by lentiviral expression of species-specific short hairpin (sh)RNAs with simultaneous expression of human mutant INSR transgenes. Results All expressed mutant INSR bound to all antibodies tested. Eight mutants showed antibody-induced autophosphorylation, while co-treatment with antibody and insulin increased maximal phosphorylation compared with insulin alone. After knockdown of mouse Insr and expression of mutant INSR in 3T3-L1 adipocytes, two antibodies (83-7 and 83-14) activated signalling via protein kinase B (Akt) preferentially over signalling via extracellular signalregulated kinase 1/2 (ERK1/2) for seven mutants. These antibodies stimulated glucose uptake via P193L, S323L, F382V and D707A mutant INSRs, with antibody response greater than insulin response for D707A. Conclusions/interpretation Anti-INSR monoclonal antibodies can activate selected naturally occurring mutant human insulin receptors, bringing closer the prospect of novel therapy for severe insulin resistance caused by recessive mutations.

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Measurement of Glucose Uptake in Cultured Cells

Cited by 115 publications

References 50 publications

Beneficial Effect of Pistachio Consumption on Glucose Metabolism, Insulin Resistance, Inflammation, and Related Metabolic Risk Markers: A Randomized Clinical Trial

Beneficial Effect of Pistachio Consumption on Glucose Metabolism, Insulin Resistance, Inflammation, and Related Metabolic Risk Markers: A Randomized Clinical Trial

Methyl 6‐Amino‐6‐deoxy‐d‐pyranoside‐Conjugated Platinum(II) Complexes for Glucose Transporter (GLUT)‐Mediated Tumor Targeting: Synthesis, Cytotoxicity, and Cellular Uptake Mechanism

Evaluation of anti-insulin receptor antibodies as potential novel therapies for human insulin receptoropathy using cell culture models

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