3-(5′-hydroxymethyl-2′-furyl)-1-benzylindazole (YC-1), sGC inhibitor ODQ, non-competitive selective PDE inhibitor IBMX, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) were purchased from Sigma-Aldrich (St. Louis, MO, USA). The PKA inhibitor H89 were purchased from Enzo Life Sciences (Farmingdale, NY, USA). The sGC activator BAY 41-2272 and NF-κB inhibitor Ro 106-9920 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). The extracellular regulated MAP kinase (ERK) inhibitor U0126, p38 MAP kinases inhibitor SB202190, c-Jun N-terminal kinases (JNK) inhibitor SP600125, HIF-1α inhibitor CAY10585, p38 activator anisomycin, and phosphatidylinositol-3-kinases (PI3K) inhibitor wortmannin were purchased from Cayman Chemical (Ann Arbor, MI, USA).
Cay10585
Manufactured by Cayman Chemical
Sourced in United States
CAY10585 is a laboratory reagent supplied by Cayman Chemical. It is a solid compound intended for research use.
Automatically generated - may contain errors
Lab products found in correlation
Anisomycin, by Cayman Chemical (1 mentions)
Wortmannin, by Cayman Chemical (1 mentions)
Bay 41 2272, by Santa Cruz Biotechnology (1 mentions)
Ro 106 9920, by Santa Cruz Biotechnology (1 mentions)
Sb202190, by Cayman Chemical (1 mentions)
U0126, by Cayman Chemical (1 mentions)
Sp600125, by Cayman Chemical (1 mentions)
3 isobutyl 1 methyl xanthine (ibmx), by Merck Group (1 mentions)
Chetomin, by Santa Cruz Biotechnology (1 mentions)
Sq22536, by Merck Group (1 mentions)
Dbcamp, by Merck Group (1 mentions)
Forskolin, by Merck Group (1 mentions)
β actin, by Abcam (1 mentions)
Caspase 1, by Abcam (1 mentions)
Ham s f 12 nutrient mixture, by Thermo Fisher Scientific (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by GE Healthcare (1 mentions)
Modular incubator chamber mic 101, by Embrient Inc (1 mentions)
Dm αkg, by Merck Group (1 mentions)
7 protocols using cay10585
1
Inhibition of Signaling Pathways in Cell Viability Assay
2
Modulating Immune Responses via Cytokine and Signaling Pathways
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For IL-4 neutralization, mice were given 1 mg of either rat IgG or anti-mouse IL-4 Ab (11B11) intraperitoneally (i.p.) at day 7 p.i. For Notch inhibition, GSI Ly411,575 (Sigma) or dimethyl sulfoxide (DMSO) was given i.p. at 7 mg/kg of body weight daily from day 7 p.i. In the Notch ligand blockade experiment, 0.25 mg of purified hamster IgG (Leinco Technologies, St. Louis, MO) or hamster anti-mouse Jagged1 or Jagged2 Ab (BioLegend, San Diego, CA) was given to mice i.p. at day 0 and at days 4 and 8 p.i. HIF-1α signaling was blocked with inhibitors against HIF-1α accumulation (30 µM CAY10585) (Cayman Chemical, Ann Arbor, MI) or p300–HIF-1α complex formation (7.5 nM chetomin) (Santa Cruz Biotechnologies, Dallas, TX) for 3 days prior to harvesting DCs.
3
HIF-1α Stabilization and Autophagy Modulation
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For HIF-1α stabilization, PHD inhibitor IOX2 at 0.1 mM or dimethyloxalylglycine (DMOG; Cayman Chemical) at 0.1 mM was added along with infection, whereas inhibition of HIF-1α was achieved by treating the cells with 10 μM CAY10585 (Cayman Chemical) 30 min prior to infection. Autophagy was either blocked or induced 2 hpi and cells were analyzed 24 hpi, treating MDM with Spautin-1 at 10 μM or MRT68921 at 5 μM and PI-103 at 5 μM (Selleckchem Chemicals).
4
Multifunctional Iron Chelator M30 Assay Protocol
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The multifunctional iron chelator, M30 (5-[N-methyl-N-propargylaminomethyl]-8-hydroxyquinoline) was synthesized and kindly provided by Varinel Inc. (Philadelphia, PA) [12 (link), 13 (link)]. CAY10585 (HIF-1α inhibitor) was purchased from Cayman Chemical (Ann Arbor, MI). Forskolin (AC agonist), SQ22536 (AC inhibitor), H89 (PKA inhibitor), and db-cAMP (stable cAMP analogue) were purchased from Sigma-Aldrich (St. Louis, MO). Antibodies against catalase (CAT), glutathione peroxidase 1 (GPx1), NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), caspase-1, and β-actin were ordered from Abcam (Cambridge, UK).
5
Fibroblast Cultivation from Keloid Tissue
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This study included 38 patients with keloids (Supplementary Table S4) undergoing surgical excision in the Yonsei University Health System, Seoul, Koris. Adjacent normal skin tissue samples (n ¼ 9) were also obtained. Fibroblast cultivation was performed using the dermal portion of keloid tissues obtained from five patients with keloid. Patients signed written informed consent. This study was approved by the institutional review board of Yonsei University Hospital (IRB: 4-2018-1024). The specimens were cut into 2-mm 3 sizes, placed in 60-mm culture dishes, and cultured in a mixture of DMEM (Gibco, Thermo Fisher Scientific, Waltham, MA) and Ham's nutrient mixture F12 (Gibco, Thermo Fisher Scientific) medium, by a 3:1 ratio. The supplements included 10% fetal bovine serum (Hyclone, GE Healthcare, Chicago, IL), 100 U/ml penicillin, 100 mg/ml streptomycin, 1 Â 10 À10 M cholera toxin, 0.4 mg/ml hydrocortisone, 5 mg/ml insulin, 5 mg/ml transferrin, and 2 Â 10 À11 M triiodothyronine. The outgrown cells were harvested, and the identities of cells were verified by vimentin and a-SMA expression. Cells were maintained in incubators at 37 C and exposed to normoxia (20% O 2 and 5% CO 2 ) or hypoxia (1% O 2 and 5% CO 2 ). In experiments involving inhibitors, cells were treated with 30 mM CAY10585 (Cayman Chemical, Ann Arbor, MI) or the same amount of DMSO as control for 24 hours.
6
Hypoxia-induced Adipocyte Differentiation
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Human SGBS preadipocytes (Wabitsch et al., 2001 (link)) were cultivated and differentiated to mature adipocytes as described previously (Geiger et al., 2011a (link)). Fully differentiated cells were exposed to hypoxia (of 1% O2, 5% CO2 and 94% N2), using a MIC-101 modular incubator chamber (Billups-Rothenberg, Inc., Del Mar, CA, USA), sealed and incubated at 37 °C. Adipocytes were cultured in the hypoxic environment for 3, 6 and 16 h while the control group was cultured under normoxic conditions (21% O2) prior to harvest. Reagents were obtained from Sigma–Aldrich (St. Louis, MO, USA) unless specified otherwise. For suppressing HIF-1α accumulation we used the HIF-1α inhibitor CAY10585 (Cayman Chemical) dissolved in DMSO and DMSO alone as a control.
7
Culturing LX2 Stellate Cells for Experiments
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The LX2 human stellate cell line (LX2 cells; Millipore Sigma, SCC064) was maintained in DMEM (Gibco, 11965-084) supplemented with 2% fetal bovine serum (FBS), sodium pyruvate (1 mM), and Penicillin-Streptomycin (100 U/mL). Unless otherwise mentioned, for cell culture experiments, LX2 cells were treated overnight with indicated treatments in Gln-free DMEM (Gibco, 11960-051) containing 10% FBS, 1 mM sodium pyruvate, and 100 U/mL Penicillin-Streptomycin. Reagents and inhibitors used in these studies included: TGFβ (PeproTech; AF-100-21C), CAY10585 (Cayman Chemical; 10012682), BCLA (Sigma-Aldrich; C9033), dm-αKG (Sigma-Aldrich; 349631), and DMOG (MedChemExpress; HY-15893).
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