2-(n-(7-nitrobenz-2-oxa-1-3-diazol-4-yl)amino)-2-deoxyglucose and Disease-Models--Animal

Abscisic acid (ABA) is a plant hormone active also in mammals where it regulates, at nanomolar concentrations, blood glucose homeostasis. Here we investigated the mechanism through which low-dose ABA controls glycemia and glucose fate. ABA stimulated uptake of the fluorescent glucose analog 2-NBDG by L6, and of [

Topics: 4-Chloro-7-nitrobenzofurazan; Abscisic Acid; AMP-Activated Protein Kinase Kinases; Animals; Cell Line; Deoxyglucose; Diabetes Mellitus; Disease Models, Animal; Glucose; Insulin; Male; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle, Skeletal; Myoblasts; Protein Kinases; Rats; Rats, Wistar; RNA, Small Interfering; TRPM Cation Channels

Extrahepatic bile duct cancer (cholangiocarcinoma) has a poor prognosis. Since surgical resection is the only way to prolong the patient's life, it is of critical importance to correctly determine the extent of lesions. However, conventional pre-operative assessments have insufficient spatial resolution for determining the surgical margin. A fluorescent contrast agent might provide a more precise measure to identify anomalies in biliary surface, when combined with probe-based confocal laser endomicroscopy (pCLE). We have previously shown that 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-L-glucose (2-NBDLG), a fluorescent derivative of L-glucose (fLG), is specifically taken up into spheroids consisting of cells showing heterogeneous nuclear-cytoplasm ratio, a feature of malignant cells in clinical settings. In addition, a combined use of 2-TRLG, a membrane-impermeable fLG, with 2-NBDLG visualized membrane integrity as well. We therefore explored in the present study the availability of the fLGs in vivo as a contrast agent for pCLE by using a hamster model of cholangiocarcinoma. Extrahepatic cholangiocarcinoma developed in mid common duct in ~20 % of the animals subjected to cholecystoduodenostomy with the ligation at the distal end of the common duct followed by injection of a carcinogen N-nitrosobis(2-oxopropyl)amine. After infusing bile duct with a solution containing 2-NBDLG and 2-TRLG, the lumen was surgically exposed and examined by pCLE. Fluorescence pattern characterized by bright spots and dark clumps was detected in the areas diagnosed with cholangiocarcinoma in later histopathology, whereas no such pattern was detected in control animals. These findings may form a basis for elucidating a potential availability of fLGs in imaging cholangiocarcinoma by pCLE.

Topics: 4-Chloro-7-nitrobenzofurazan; Animals; Bile Duct Neoplasms; Bile Ducts; Cholangiocarcinoma; Cricetinae; Deoxyglucose; Disease Models, Animal; Endoscopes; Female; Fluorescence; Mesocricetus; Microscopy, Confocal

We have reported earlier, an orally active insulin-like protein (ILP) from Costus igneus having potent hypoglycemic property in STZ-induced diabetic Swiss mice. The blood glucose level was reduced significantly within two hours after feeding ILP orally in an oral glucose tolerance test. The present study elucidates the mechanism underlying the hypoglycemic action of ILP. Mechanism of action of ILP was studied in differentiated L6 myotubes. 2-NBDG uptake stimulated by ILP was studied in differentiated L6 myotubes under normoglycemic, hyperglycemic and induced insulin resistant conditions. ILP treatment significantly increased 2-NBDG uptake in differentiated L6 myotubes. The levels of insulin signaling molecules IRS-1 and GLUT-4 were assessed in ILP treated L6 myotubes by immunoblot analysis of cytoplasmic and plasma membrane fractions respectively. Immunoblot analysis revealed an increase in cytoplasmic IRS-1 with a concomitant increase in GLUT-4 translocation to the plasma membrane in a time dependent manner. Toxicity studies of ILP were performed on normal as well as diabetic Swiss albino mice. ILP did not show any toxicity in the acute and sub-chronic toxicity studies in normal as well as diabetic Swiss albino mice. Mass spectrometry was carried out to identify ILP. MALDI TOF/TOF MS analysis of ILP revealed sequence homology with the predicted protein from Physcomitrella patens. Our study reveals that ILP acts via insulin signaling pathway and can be used as oral insulin mimetic.

Topics: 4-Chloro-7-nitrobenzofurazan; Animals; Blood Glucose; Costus; Deoxyglucose; Diabetes Mellitus, Experimental; Disease Models, Animal; Glucose Tolerance Test; Hypoglycemic Agents; Insulin; Mice; Muscle, Skeletal; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Tumor Necrosis Factor-alpha

Evidence indicates that central galanin is involved in regulation of insulin resistance in animals. This study investigates whether type 1 galanin receptor (GAL1) in the brain mediates the ameliorative effect of galanin on insulin resistance in skeletal muscles of type 2 diabetic rats. Rats were intracerebroventricularly (i.c.v.) injected with galanin(1-13)-bradykinin(2-9) amide (M617), a GAL1 agonist, and/or Akti-1/2, an Akt inhibitor, via caudal veins once per day for 10 days. Insulin resistance in muscle tissues was evaluated by glucose tolerance and 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxyglucose (2-NBDG) tests, peroxisome proliferator-activated receptor-γ (PPARγ), glucose transporter 4 (GLUT4) mRNA expression levels, Akt phosphorylation, and GLUT4 and vesicle-associated membrane protein 2 (VAMP2) concentration at plasma membranes in muscle cells. The results show that i.c.v. treatment with M617 increased glucose tolerance, 2-NBDG uptake, PPARγ levels, Akt phosphorylation, GLUT4 protein, and GLUT4 mRNA expression levels as well as GLUT4 and VAMP2 concentration at plasma membranes. All increases may be blocked by pretreatment with Akti-1/2. These results suggest that activated central GAL1 may trigger the Akt signaling pathway to alleviate insulin resistance in muscle cells. Therefore, the impact of galanin on insulin resistance is mediated mainly by GAL1 in the brain, and the GAL1 agonist may be taken as a potential antidiabetic agent for treatment of type 2 diabetes mellitus. © 2016 Wiley Periodicals, Inc.

Topics: 4-Chloro-7-nitrobenzofurazan; Animals; Blood Glucose; Body Weight; Bradykinin; Deoxyglucose; Diabetes Mellitus, Experimental; Disease Models, Animal; Galanin; Glucose Tolerance Test; Glucose Transporter Type 4; Insulin; Insulin Resistance; Male; Muscle, Skeletal; Peptide Fragments; PPAR gamma; Rats; Rats, Wistar; Receptor, Galanin, Type 1; Subcellular Fractions; Vesicle-Associated Membrane Protein 2

We demonstrate an optical strategy using intravital microscopy of dorsal skin flap window chamber models to image glucose uptake and vascular oxygenation in vivo. Glucose uptake was imaged using a fluorescent glucose analog, 2-[N-(7-nitrobenz-2-oxa-1,3-diaxol-4-yl)amino]-2-deoxyglucose (2-NBDG). SO2 was imaged using the differential absorption properties of oxygenated [HbO2] and deoxygenated hemoglobin [dHb]. This study was carried out on two sibling murine mammary adenocarcinoma lines, 4T1 and 4T07. 2-NBDG uptake in the 4T1 tumors was lowest when rates of delivery and clearance were lowest, indicating perfusion-limited uptake in poorly oxygenated tumor regions. For increasing rates of delivery that were still lower than the glucose consumption rate (as measured in vitro), both 2-NBDG uptake and the clearance rate from the tumor increased. When the rate of delivery of 2-NBDG exceeded the glucose consumption rate, 2-NBDG uptake decreased with any further increase in rate of delivery, but the clearance rate continued to increase. This inflection point was not observed in the 4T07 tumors due to an absence of low delivery rates close to the glucose consumption rate. In the 4T07 tumors, 2-NBDG uptake increased with increasing rates of delivery at low rates of clearance. Our results demonstrate that 2-NBDG uptake in tumors is influenced by the rates of delivery and clearance of the tracer. The rates of delivery and clearance are, in turn, dependent on vascular oxygenation of the tumors. Knowledge of the kinetics of tracer uptake as well as vascular oxygenation is essential to make an informed assessment of glucose demand of a tumor.

Topics: 4-Chloro-7-nitrobenzofurazan; Animals; Breast Neoplasms; Cell Line, Tumor; Deoxyglucose; Disease Models, Animal; Female; Fluorescent Dyes; Hypoxia; Kinetics; Mice; Neoplasm Metastasis; Oxygen Consumption; Regional Blood Flow; Sulfur Dioxide; Time Factors

Alzheimer's disease is characterized by β-amyloid accumulation in the central nervous system. As β-amyloid is neurotoxic in culture, we have explored the mechanisms of toxicity in the search for therapeutic targets for Alzheimer's disease and now identify a key role for poly(ADP-ribose) polymerase in β-amyloid-induced neuronal death. Exposure of hippocampal neuronal/glial co-cultures to β-amyloid peptides activates the glial nicotinamide adenine dinucleotide phosphate oxidase, followed by predominantly neuronal cell death. β-amyloid exposure caused the progressive loss of mitochondrial membrane potential in astrocytes, accompanied by transient mitochondrial depolarizations caused by reversible openings of the mitochondrial permeability transition pore. The transients were absent in cultures from cyclophilin D knockout mice, leaving the slow depolarization available for study in isolation. β-amyloid exposure decreased both nicotinamide adenine dinucleotide fluorescence and oxygen consumption, while provision of mitochondrial substrates reversed the depolarization, suggesting that substrate supply was limiting. Poly(ADP-ribose) polymerase is activated by oxidative stress and consumes nicotinamide adenine dinucleotide, decreasing substrate availability. β-amyloid exposure caused accumulation of the poly(ADP-ribose) polymerase product, poly-ADP-ribose polymers, in astrocytes. Inhibition of either poly(ADP-ribose) polymerase or of the nicotinamide adenine dinucleotide phosphate oxidase prevented the appearance of poly-ADP-ribose polymers and the mitochondrial depolarization. Exposure of co-cultures to β-amyloid for >8 h decreased nicotinamide adenine dinucleotide and mitochondrial membrane potential and increased cell death in neurons, all of which were prevented by poly(ADP-ribose) polymerase inhibitors. Poly-ADP-ribose polymers increased with age in the brains of the TASTPM Alzheimer mouse model. We conclude that β-amyloid-induced neuronal death is mediated by poly(ADP-ribose) polymerase in response to oxidative stress generated by the astrocytic nicotinamide adenine dinucleotide phosphate oxidase.

Topics: 4-Chloro-7-nitrobenzofurazan; Age Factors; Alzheimer Disease; Amyloid beta-Peptides; Analysis of Variance; Animals; Animals, Newborn; Astrocytes; Calcium; Cell Death; Coculture Techniques; Deoxyglucose; Disease Models, Animal; Glial Fibrillary Acidic Protein; Glucose; Hippocampus; Membrane Potential, Mitochondrial; Mice; Mice, Transgenic; NAD; NADPH Oxidases; Neurons; Oxygen Consumption; Peptide Fragments; Poly(ADP-ribose) Polymerases; Rats; Rats, Sprague-Dawley; Tetradecanoylphorbol Acetate

To study the effect of the Chinese compound prescription Chaihu Shugan Tang (CHSGT) on the excitability in the cerebral cortex and hippocampus (different brain regions) of pentetrazole (PTZ)-kindled chronic epileptic rats.. To establish the model of chronic kindling rats intraperitoneal injected with pentylenetet. Fully kindled rats were randomized into control and experimental groups for intragastric administration of normal saline (control, model), Sodium Valproate and CHSGT at the high, medium and low doses for 4 consecutive weeks. The content of 2-NBDG, the glutamate (Glu) and the aspartate (Asp) in different brain regions of rats were detected by fluorescence imaging techniques and HPLC assay respectively.. CHSGT at the high, medium and low doses all significantly decreased the content of 2-NBDG, the Glu and the Asp in different brain regions of chronic epileptic rats (P < 0.01).. CHSGT can inhibit the excitability in different brain regions of PTZ-induced epileptic rats, by decreasing the level of excitatory neurotransmitter maybe one of its antiepileptic mechanisms.

Topics: 4-Chloro-7-nitrobenzofurazan; Animals; Aspartic Acid; Brain; Chronic Disease; Deoxyglucose; Disease Models, Animal; Drugs, Chinese Herbal; Epilepsy; Glutamic Acid; Hippocampus; Humans; Kindling, Neurologic; Male; Pentylenetetrazole; Random Allocation; Rats; Valproic Acid