Briefly, THP-1 human monocytes (TIB-202; ATCC) were cultured in RPMI-1640 (189-02025; Wako) supplemented with 10% fetal bovine serum (175012; Nichirei), penicillin (100 U/mL) (168-23191; Wako), and streptomycin (100 mg/mL) (168-23191; Wako). F. nucleatum was cultured as previously described (43 (link)). THP-1 monocytes were pretreated with 100 μM FAC (RES20400-A7; Sigma-Aldrich), 100 μM DFO (D9533; Sigma-Aldrich), or the indicated concentration of calyculin A (038-14453; Wako) for the indicated time, followed by stimulation with 100 ng/mL LPS (tlrl-eblps; InvivoGen) for the indicated time. THP-1 monocytes were differentiated into macrophages using 6 ng/mL phorbol 12-myristate 13-acetate (AG-CN2-0010; AdipoGen) for 48 hours. Differentiated cells were pretreated with 100 μM FAC, 100 μM DFO, or 5 μM TAK-242 (13871; CAYMAN) for the indicated times, followed by treatment with F. nucleatum at a multiplicity of infection of 10 for 3 hours.
Tak 242
Manufactured by Cayman Chemical
Sourced in United States
TAK-242 is a synthetic small molecule that functions as a toll-like receptor 4 (TLR4) inhibitor. It is used as a research tool to study the role of TLR4 signaling in various biological processes and disease models.
Automatically generated - may contain errors
Lab products found in correlation
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
Calyculin a, by Fujifilm (1 mentions)
Tlrl eblps, by InvivoGen (1 mentions)
Penicillin, by Fujifilm (1 mentions)
Streptomycin, by Fujifilm (1 mentions)
Rpmi 1640, by Fujifilm (1 mentions)
Fetal bovine serum (fbs), by Nichirei Biosciences (1 mentions)
Wortmannin, by Merck Group (1 mentions)
Lps escherichia coli 055 b5, by Merck Group (1 mentions)
12 o tetradecanoylphorbol 13 acetate tpa, by Merck Group (1 mentions)
Tumor necrosis factor (tnf), by Thermo Fisher Scientific (1 mentions)
U0126, by Cell Signaling Technology (1 mentions)
Pam3csk4, by InvivoGen (1 mentions)
Flexstation multilabel plate reader, by Molecular Devices (1 mentions)
Pmetluc2, by Takara Bio (1 mentions)
Lipopolysaccharide lps, by Beyotime (1 mentions)
Roscovitine, by Fujifilm (1 mentions)
Myriocin, by Cayman Chemical (1 mentions)
Fatty acid free bovine serum albumin, by Fujifilm (1 mentions)
Ro 31 8220, by Santa Cruz Biotechnology (1 mentions)
24 protocols using tak 242
1
THP-1 monocyte immune response assay
2
Inhibition of Inflammatory Pathways
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Unless otherwise stated, cells were treated with the final concentration of 10 μM CFMB [(2S)-2-(4-chlorophenyl)-3,3-dimethyl-N-(5-phenylthiazol-2-yl)butanamide] (Calbiochem) (ChemSpider ID: 24656891) (EC50 of ~0.7 μM)34 (link), 10 μM AR420626 [N-(2,5-Dichlorophenyl)-4-(furan-2-yl)-2-methyl-5-oxo-1,4,5,6,7,8-hexahydro-quinoline-3-carboxamide] (Glixx Laboratories) (EC50 of ~0.7 μM)33 , 100 ng/mL Escherichia coli 055:B5 LPS (Sigma), 10 ng/mL TNF (Gibco) and 200 nM 12-O-Tetradecanoylphorbol-13-acetate (TPA) (Sigma), 100 ng/mL PAM3CSK4 (Invivogen). 500 ng/mL of Pertussis Toxin (PT) (Invitrogen), 10 μM YM25489039 (link), 2 μM Wortmannin (Sigma), 10 μM U0126 (Cell Signalling) and 2 μM TAK-242 (Cayman Chemical)35 (link)36 (link).
3
NF-κB Activation by Red Blood Cell-Derived Extracellular Vesicles
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Monocytes (2x104 cells/well) were seeded in 96-well plates in RPMI-1640 containing 10% FBS were transfected with the NF-kB-responsive luciferase reporter construct (NF-kB pMetLuc 2) and control plasmid (pMetLuc 2) (Clontech, Mountain View, CA, USA) in RPMI-1640 containing 10% FBS and incubated for 24 h. Cells were pretreated with or without TAK-242 (1 µM; Cayman Chemical Company, Ann Arbor, MI, USA) and then incubated with RBC 0-EVs or RBC 21-EVs for 24 h at 37°C in a 5% CO2 atmosphere. The medium containing luciferase was collected for each treatment and was incubated with luciferin. Luminescence emitted from the luciferin cleavage was monitored using the Flexstation™ multilabel plate reader (Molecular Devices, San Jose, CA, USA).
4
HMGB1 Modulation by Glycyrrhizic Acid
5
Inhibition of Lipid Signaling Pathways
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PA was purchased from Nacalai Tesque (Kyoto, Japan). Fatty-acid-free bovine serum albumin (BSA) was purchased from FUJIFILM Wako Chemicals (Osaka, Japan). TAK-242, an inhibitor of toll-like receptor 4 (TLR4), and myriocin, an inhibitor of serine palmitoyl-transferase (SPT), a key enzyme for ceramide de novo synthesis, were purchased from Cayman Chemical (Ann Arbor, MI, USA). Ro 31-8220, a potent inhibitor of protein kinase C (PKC), was obtained from Santa Cruz Biotechnology (Dallas, TX, USA). Roscovitine, a selective cyclin-dependent kinase (CDK) inhibitor of CDK1 and CDK2, was purchased from FUJIFILM Wako Chemicals. The inhibitors were reconstituted in dimethyl sulfoxide.
6
Inhibition of ERK and TLR4 Pathways in UV-Exposed Cells
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Prior to UV irradiation, cells were pre-treated with various concentrations of the ERK inhibitor U0126 (Sigma-Aldrich, MO, USA) or TLR4 inhibitor TAK242 (Cayman Chemicals, MI, USA). Immediately before irradiation, the culture medium was removed and cells were washed twice in PBS. After UVA exposure, the cells were cultured in fresh media and incubated for an additional amount of time before being collected for further analysis.
7
Fluid Shear Stress Regulates Ngf Expression
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To assess Ngf expression, MC3T3-E1 cell cultures were subjected to fluid shear stress of 1 mL/min for 0 (control), 10, 15, 30, and 60 min using a previously developed microfluidics fluid flow device46 (link) coupled to a standard peristaltic pump with 1.6 mm (ID) polyethylene tubing. To determine the requirement of the NF-κB signaling pathway following loading, osteoblast cultures were supplemented with either 5 μM BAY 11-7082 (Sigma) or 50 μM pyrrolidine dithiocarbamate (PDTC, Sigma) 1 h before loading for 60 min at 1 mL/min. To determine the role of TLR4 signaling, cultures were supplemented with either 5 or 10 μM TAK-242 (Cayman Chemicals) 2 h before loading for 60 min at 1 mL/min. Cells were harvested 1 h following loading in all inhibitor experiments. Control groups were not loaded and/or treated with DMSO (vehicle).
8
Investigating HMGB1 and RAGE Signaling
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Puromycin dihydrochloride (#P9620) was purchased from Sigma-Aldrich; Merck KGaA. TLR4 antagonist TAK-242 (#13871) and RAGE antagonist FPS-ZM1 (#11909) were purchased from Cayman Chemical Co. Chicken anti-HMGB1 polyclonal antibody (#326052233) was purchased from Shino-Test. Control shRNA plasmid-A (#sc-108060), HMG-1 shRNA plasmid (#sc-37982-SH), and anti-RAGE antibody (anti-mouse, monoclonal, sc-80652) were purchased from Santa Cruz Biotechnology, Inc. HMGB1 antibody (anti-mouse, monoclonal, GTX628834) was purchased from GeneTex, Inc. Anti-phospho-p44/42 MAPK antibody (p-ERK; anti-rabbit, monoclonal, #4370), anti-p44/42 MAPK antibody (ERK; anti-rabbit, monoclonal, #4695), horseradish peroxidase (HRP)-conjugated IgG antibody (goat anti-rabbit, monoclonal, #7074), HRP-conjugated IgG antibody (goat anti-mouse, monoclonal, #7076), and Alexa Fluor 488-conjugated IgG (H+L) F(ab′)2 fragment (goat anti-rabbit, monoclonal, #4412) were purchased from Cell Signaling Technology, Inc. Anti-TLR4 antibody (anti-rabbit, polyclonal, #ab13556), anti-β-actin antibody (anti-mouse, monoclonal, #ab49900), anti-CGRP antibody (anti-goat, polyclonal, #ab36001), and Alexa Fluor 647-conjugated IgG H&L antibody (donkey anti-goat, monoclonal, #ab150135) were purchased from Abcam.
9
TAK-242 Administration for Wound Healing
10
Inhibition of TLR4 and C5a Receptor in IgG Response
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