The following reagents were used: Streptozotocin (Merck, Darmstadt, Germany); AdipoRon (Selleckchem, Houston, TX, USA); a triglyceride assay kit and a non-esterified free fatty acid assay kit (Jiancheng Bioengineering Institute, Nanjing, China); and dihydroethidium (Thermofisher, MA, USA). The following antibodies were used: Rabbit monoclonal to AdipoR1 (EPR6626,Abcam, Cambridge, UK); rabbit polyclonal to RNA-binding protein with multiple splicing (RBPMS; Abcam); rabbit monoclonal to glutamine synthetase (EPR13022(B),Abcam); chicken polyclonal to glial fibrillary acidic protein (GFAP; Abcam); rabbit polyclonal to EGR4 (Abcam); phospho-AMPKα rabbit monoclonal antibody (mAb) and AMPKα rabbit mAb (40H9/D5A2,Cell signaling, Boston, USA); phospho-acetyl CoA carboxylase (ACC) rabbit mAb and ACC rabbit mAb (D7D11/C83B10,Cell signaling); cleaved caspase-3 rabbit mAb (Cell signaling), β-actin (8H10D10,Cell signaling); donkey anti-rabbit AlexaFluor-594 and donkey anti-chicken Alexa 488 (Jackson ImmunoResearch, Philadelphia, USA). The following reagents were used: 5,6-chloromethyl-2ʹ,7ʹ-dichlorodihydrofluorescein diacetate (CM-H2DCFDA; Thermofisher); RNAimax (Thermofisher); Opti-MEM I (Thermofisher); small interfering RNA (siRNA; GenePharma, Shanghai, China); Cell Counting Kit-8 (CCK-8; Dojindo, Tokyo, Japan); and 4ʹ,6-diamidino-2-phenylindole (DAPI; Absin, Shanghai, China).
Adiporon
Manufactured by Selleck Chemicals
Sourced in United States
AdipoRon is a synthetic compound that acts as an agonist (activator) of the AdipoR1 and AdipoR2 receptors. These receptors are involved in the regulation of cellular processes related to energy metabolism and inflammation.
Automatically generated - may contain errors
Lab products found in correlation
Rnaimax, by Thermo Fisher Scientific (2 mentions)
Streptozotocin (stz), by Merck Group (1 mentions)
Dihydroethidium, by Thermo Fisher Scientific (1 mentions)
Glial fibrillary acidic protein (gfap), by Abcam (1 mentions)
Donkey anti rabbit alexafluor 594, by Jackson ImmunoResearch (1 mentions)
Opti mem 1, by Thermo Fisher Scientific (1 mentions)
Non esterified free fatty acid assay kit, by Nanjing Jiancheng (1 mentions)
Rbpms, by Abcam (1 mentions)
Cleaved caspase 3 rabbit mab, by Cell Signaling Technology (1 mentions)
Donkey anti chicken alexa 488, by Jackson ImmunoResearch (1 mentions)
Cell counting kit 8 cck 8, by Dojindo Laboratories (1 mentions)
β actin, by Cell Signaling Technology (1 mentions)
Triglyceride assay kit, by Nanjing Jiancheng (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
Ang 2, by Merck Group (1 mentions)
Alzet mini osmotic pump, by Durect (1 mentions)
Anti fibronectin ab2413, by Abcam (1 mentions)
Anti cleaved caspase 3 9661, by Cell Signaling Technology (1 mentions)
Anti bcl 2 af6139, by Affinity Biosciences (1 mentions)
Anti β actin 60008 1 ig, by Proteintech (1 mentions)
7 protocols using adiporon
1
Adiponectin Receptor Signaling in Diabetic Retinopathy
2
Isolation and Culture of MUSCs
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C2C12 cells were cultured in complete Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% foetal bovine serum (FBS; Gibco BRL, Grand Island, New York) in a 37% CO2 humidified incubator. Isolation and culture of MUSCs were performed as previously described.23 Briefly, the hindlimb muscle tissues were isolated and fully cut into pieces. Then, the pieces were then digested with collagenase I for 1 h and centrifuged at 1500 rpm for 5 min. The supernatant was discarded, and the precipitate was digested with trypsin for 15 min. The pellet was resuspended in a culture flask after centrifugation. The suspension was transferred to a new flask coated with Matrigel for 2.5 h, and AdipoRon (Selleck Chemicals, China) was used to activate APNrs.
3
Dietary Modulation of AdipoRon Effects
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Four-week old mice fed with the LP, NP and HP diets were treated via oral gavage of DMSO (as control) or AdipoRon according to a 2 × 3 factorial design. AdipoRon (Selleck Chemicals, Houston, TX, USA) dissolved in DMSO (Sigma) was suspended in 0.5% carboxymethylcellulose and was then administered orally to each group of mice (n = 6) at a dose of 50 mg/kg per day. This dosage was observed to have beneficial effects on metabolism, lifespan, and anti-apoptosis [[33] (link), [34] (link), [35] (link)]. Mice were sacrificed for sampling 28 days after treatment.
4
Ang II-Induced Hypertension and AdipoRon
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Male, 8–12 week-old C57BL/6J background mice were used in the present study. To induce hypertension, mice (n=5) were subcutaneously infused with Ang II (Sigma-Aldrich; Merck KGaA) at a 1,500 ng/kg/min dose for 14 days with ALZET mini osmotic pumps (DURECT Corporation, Cupertino, CA, USA), and in the sham group mice were treated with saline (n=5). Blood pressure was measured non-invasively every day by the tail-cuff method (Softron BP-98A; Softron Co., Ltd., Tokyo, Japan). To determine the role of APN receptor in hypertensive vascular injury, Ang II-infused mice received APN receptor agonist AdipoRon (n=5, 30 mg/kg/day; Selleck Chemicals, Houston, TX, USA) via a gavage tube. The experiments were performed in adherence with the National Institutes of Health guidelines on the Use of Laboratory Animals and were approved by the Animal Care and Use Committee of Shanghai University of Traditional Chinese Medicine (Shanghai, China).
5
Antibody Selection and Chemical Treatments for Metabolic Regulation
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The following commercial antibodies were used in our study. Anti-ADIPOR1 (ab126611), anti-AMPKα1 (ab32047), anti-collagen I (ab34710), and anti-fibronectin (ab2413) were obtained from Abcam. Anti-phospho-PERK (Thr980) (3179), anti-phospho-AMPKα (Thr172) (2535), anti-CHOP (2895S), anti-GRP78 (3177S), and anti-cleaved caspase 3 (9661S) were purchased from Cell Signaling Technology. Anti-PERK (AF5304) and anti-Bcl-2 (AF6139) were from Affinity. Anti-CHOP (GB34710) was from Servicebio. Anti-BAX (60267-1-Ig) and anti-β-actin (60008-1-Ig) were obtained from Proteintech. All secondary antibodies used for western blot analysis were purchased from Abcam. AdipoRon was from SELLECK. AICAR, compound C, and TUDCA (S3654) were also purchased from SELLECK.
6
Cell Culture and Reagent Preparation
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The CNE-2 and S18 cell lines were kindly gifted by Professor Chaonan Qian at SYSUCC. HNE2, 5-8F, C666-1 and 6-10B cells were from the Central South University Advanced Research Center (Changsha, Hunan, China). HNE2, 5-8F and 6-10B cells were cultured in RPMI-1640 medium, CNE-2 and S18 cells were cultured in Dulbecco's modified eagle medium containing 4.5 mg/mL glucose, all supplemented with 10% fetal bovine serum (Gibco, Carlsbad, CA, USA), 100 U/mL penicillin and 100 ug/mL streptomycin (Hyclone, Logan, UT, USA). Cells were maintained in a humidified atmosphere of 5% CO2 at 37 °C. The cell line was authenticated via deoxyribonucleic-acid profiling using short tandem repeat analysis.
Recombination human full-length adiponectin was dissolved in deionized water to prepare a working stock solution of approximately 0.5 mg/mL (BioVendor, Brno, Czech Republic). AdipoRon was dissolved in DMSO to prepare a working stock solution of approximately 50 mM (Selleck Chemicals, Houston, TX, USA). Compound C was purchased from MedChem Express (Monmouth Junction, NJ, USA), and was prepared as a stock concentration at 10 mM in DMSO and stored at − 80 °C.
Recombination human full-length adiponectin was dissolved in deionized water to prepare a working stock solution of approximately 0.5 mg/mL (BioVendor, Brno, Czech Republic). AdipoRon was dissolved in DMSO to prepare a working stock solution of approximately 50 mM (Selleck Chemicals, Houston, TX, USA). Compound C was purchased from MedChem Express (Monmouth Junction, NJ, USA), and was prepared as a stock concentration at 10 mM in DMSO and stored at − 80 °C.
7
AdipoRon Bioavailability in Bone Cells
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AdipoRon was purchased from Selleck Chemicals, China. MC3T3‐E1 and RAW264.7 cell lines were provided by State Key Laboratory of Oral Disease, West China Hospital of Stomatology, Sichuan University.
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