Two-month-old C57BL/KsJ- db/db male mice were purchased from Charles River Laboratories and randomly divided into 4 groups with 6 mice in each group. Mice in the control group were fed with standard chow diet. The mice in other groups were fed with MCD diet (Research Diets, A02082002BR) for 4 weeks to induce NASH. For exenatide treatment group, the mice were injected subcutaneously with exenatide (Baxter Pharmaceutical Solutions, Deerfield, IL, USA) at a dose of 20 µg/kg/day subcutaneously from the beginning of week 2 to the end of week 4. For exendin 9-39 treatment group (exenatide + exendin 9-39), the mice were further injected intraperitoneally with exendin 9-39 (Sigma-Aldrich, St. Louis, MO, USA) at a dose of 20 µg/kg/day (17 (link)) on the basis of exenatide administration. Mice in the control and NASH group were administrated with normal saline instead. After 4 weeks, blood samples were collected by retro-orbital bleeding, and the serum was collected after centrifugation (3,000 rpm, 10 min). Then the animals were sacrificed via CO2 inhalation for the collection of liver samples. The animal study was reviewed and approved by The First Affiliated Hospital of Harbin Medical University (Approval Number: 2019/02/213).
Exendin 9 39
Manufactured by Merck Group
Sourced in United States, China, Sao Tome and Principe
Exendin 9-39 is a synthesized peptide compound used as a research tool in laboratory settings. It functions as a glucagon-like peptide-1 (GLP-1) receptor antagonist, which can be utilized to study the effects of GLP-1 signaling in various experimental models.
Automatically generated - may contain errors
Lab products found in correlation
Liraglutide, by Novo Nordisk (3 mentions)
Glp 1, by Merck Group (3 mentions)
Liraglutide, by Merck Group (2 mentions)
Forskolin, by Merck Group (2 mentions)
Exendin 4, by Merck Group (2 mentions)
Penicillin, by Thermo Fisher Scientific (2 mentions)
Streptomycin, by Thermo Fisher Scientific (2 mentions)
Esi 09, by Merck Group (1 mentions)
Ly294002, by Merck Group (1 mentions)
Rbmecs, by Cell Applications (1 mentions)
Exenatide, by Merck Group (1 mentions)
C57bl ksj db db male mice, by Charles River Laboratories (1 mentions)
Elisa reader, by Dynex (1 mentions)
Adenosine, by Merck Group (1 mentions)
Mdl 12 330a, by Merck Group (1 mentions)
Carbachol, by Merck Group (1 mentions)
Biie0246, by Bio-Techne (1 mentions)
Ce3f4, by Bio-Techne (1 mentions)
Gs6201, by Bio-Techne (1 mentions)
3 isobutyl 1 methyl xanthine (ibmx), by Merck Group (1 mentions)
21 protocols using exendin 9 39
1
Exenatide Attenuates NASH in db/db Mice
2
Vagus Nerve Stimulation and GLP-1 in Diabetes
3
Effects of GLP-1 Analogs and Inhibitors
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Rats were randomized into five different groups: (i) control (HEPES buffer); (ii) liraglutide (3.2 nmol/L, Novo Nordisk); (iii) Exendin9-39 (0.3 nmol/L, Sangon Biotech, Shanghai, China); (iv) LY294002 (10 μmol/L, Sigma); and (v) Exendin9-39 + liraglutide (Exendin9-39 0.3 nmol/L, liraglutide 3.2 nmol/L).
4
Pharmacological Inhibition of PKA and EPAC
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Atropine and exendin (9–39) from Sigma-Aldrich (St. Louis, MO, USA) were prepared in solution using culture medium. Additionally, lithocholic acid and triamterene from same supplier were prepared in dimethyl sulfoxide (DMSO) as stock solutions. The PKA inhibitor 14–22 amide (PKA I) is myristoylated at the N-terminus, which enhances its cell permeability (Tocris Bioscience, Bristol, UK). H89 is a potent selective inhibitor of PKA (Sigma-Aldrich). ESI-09 (Sigma-Aldrich) has been shown to block EPAC activity and function. Other reagents from indicated suppliers were all of analytical grade.
5
GLP-1 Receptor Antagonist Exendin 9-39 in [6]-Gingerol-Induced Insulin Release
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To further confirm the role of GLP-1 in [6]-Gingerol augmented glucose-induced insulin release, we employed Exendin 9-39 (Ext9) (Sigma-Aldrich, USA), a potent GLP-1 receptor antagonist, at a dose 300 pmol/kg/min. Ext9 was administered through a femoral vein catheter continuously for 1 h, while the animal subjects were kept under sodium pentobarbital anesthesia. Saxagliptin (SxLn), an inhibitor of dipeptidyl peptidase-4 (DPP-4), was administered 4 h before the glucose load, orally at a dose of 10 μmol/.
6
Fluorescence Imaging of GLP-1 Binding
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Rat brain microvascular endothelial cells (RBMECs; Cell Applications, San Diego, CA, United States) were cultured on a cover slip in rat brain endothelial cell growth medium. After reaching confluency, cells were pre-treated with vehicle (PSS buffer), exendin 9-39 (10 nM) or H89 (Sigma-Aldrich, St. Louis, MO, United States) (10 μM) for 30 min or forskolin (Sigma-Aldrich, St. Louis, MO, United States) (10 μM) for 5 min. Cells were then incubated with GLP-1-FAM at either 10, 100 or 1,000 pM for 3 min at 37°C. DAPI was used to stain the nucleus. After fixation and washing three times with cold TBST (5 min each time), cell images were captured using confocal microscope and fluorescence intensity measured using ImageJ software1 . FAM signal from surface binding of FAM was examined by incubating cells with GLP-1-FAM at 4 or 37°C for 3 min, or free FAM (10, 100, and 1,000 pM) at 37°C for 3 min.
7
MTT Assay for PASMC Proliferation
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The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to assess proliferation of the PASMCs. In brief, after serum starvation for 48 h, proliferation of the PASMCs was induced by PDGF-BB (20 ng/mL) in DMEM supplemented with 1% FBS in 96-well plates at a density of 2 × 104 cells per well, with or without 1–10 nM liraglutide (Novo Nordisk, Novo Alle, Bagsvaerd, Denmark) pre-treatment (1 h). The GLP-1 receptor antagonist 500 nM exendin (9–39) (Sigma–Aldrich, St. Louis, MO, USA) was added to the cells for 30 min prior to treatment with liraglutide. After induction for 24 h, 48 h, and 72 h, MTT (0.5 mg/mL) was added to the medium for 4 h. The culture medium was then removed, and the cells were dissolved in isopropanol and shaken for 10 min. The amount of MTT formazan was quantified at 540 and 630 nm using an enzyme-linked immunosorbent assay (ELISA) reader (DYNEX Technologies, Denkendorf, Germany).
8
Characterization of Recombinant Soluble Adenylyl Cyclase
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EP45 was produced by Genscript (NJ, USA) at ≥ 97% purity and with C-terminal amidation. Purity and identity were confirmed in-house by reversed-phase high-performance liquid chromatography (RP-HPLC) and electron spray mass spectroscopy (ESMS), respectively. All cAMP analogs were from the BIOLOG Life Science Institute (Bremen, Germany). GLP-1, glucagon, exendin-4, exendin(9–39), GIP, PYY(1–36), PYY(3–36), forskolin, IBMX, adenosine, DDA, MDL-12,330 A, carbachol, ATP, 4-acetoxybenzyl alcohol (4-Abn-OH), and ESI-05 were from Sigma-Aldrich (St. Louis, MO). CE3F4, GS6201, and BIIE0246 were from Tocris Biosciences (Minneapolis, MN). LRE1 and HEK293 cells stably expressing recombinant sAC were from Profs. L. R. Levin and J. Buck (Weill Cornell Medical College, New York, NY).
9
Trichothecene Exposure in B6C3F1 Mice
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B6C3F1 mice (10–11 weeks, female, Comparative Medicine Center of Yangzhou University) were individually housed in the room with a normal 12 h light/dark cycle. The room temperature and relative humidity were 20–23 °C and 40–60%, respectively. All guidelines for animal experiments followed the Institutional Animal Care and Use Committee at Nanjing Agricultural University. Type B trichothecenes were tested using a Liquid Chromatograph Mass Spectrometer and by conducting Elemental Analysis with a purity of more than 98%. SP (R&D Systems, Inc), GLP-1 and GLP-1R antagonist Exendin9-39 (Sigma-Aldrich, St. Louis, MO, USA) were dissolved in PBS. The NK-1R antagonist Emend® (Merck & Co, Inc) was dissolved in 1% DMSO in filter-sterilized PBS.
10
Metformin's Glucose Regulation Mechanism
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Overnight fasted mice were treated either with saline (vehicle) or M400, M200 and M60, respectively(Glucophage; Merck, France) administered by oral gavage. Thirty min later animals were given D-glucose either orally at a dose 3 g/kg or intraperitoneally at a dose 1 g/kg and subjected to GTT. The degree of glucose intolerance was quantified as AUC. Insulin levels were measured in plasma obtained from blood samples collected before and 30 min after glucose administration, using the Sensitive rat insulin RIA kit (Merck Millipore, USA). Compound C (20 mg/kg body weight; Abcam), Exendin 9–39 (0.1 mg/kg body weight; Sigma Aldrich) and MK-801 (0.3 mg/kg body weight; Sigma Aldrich), i.e. the pharmaceutical antagonists of the AMPK, GLP1R and NMDAR, respectively, were administered via i.p. injections 30 min (Compound C) or 10 min (Exendin 9–39, MK-801) before metformin.
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