BV2 cells (Rio de Janeiro Cell Bank, Rio de Janeiro, Brazil, Cat# 0356, RRID: CVCL_0182) were cultured in 96-well plates at a density of 0.5 × 105 per well. Lipopolysaccharide (LPS) and adenosine 5′-triphosphate (ATP) were used to induce neuroinflammation in cells, which is regarded as a classic model of inflammation (Fan et al., 2020; Liao et al., 2020; Tastan et al., 2021). The cells were incubated with Lup (0, 1, 5, 10, 20, 100, or 200 μM) for 2 hours, then LPS (1 μg/mL; Cat# GC205009, Servicebio, Shanghai, China) and ATP (5 mM; Cat# D7378, Beyotime, Shanghai, China) (Nam et al., 2018; Lee et al., 2020; Tastan et al., 2021). Next, the cell culture medium containing Lup, LPS, and ATP was replaced with fresh medium, after which various concentrations of Lup (0, 1, 5, 10, 20, 100, or 200 μM) were added to the plates for 24 hours. In addition, to assess the role of the NLRP3 inflammasome in BV2 cells, CY-09 (10 μM; Cat# HY-103666, MedChemExpress, Shanghai, China) was used to specifically inhibit the NLRP3 inflammasome. Cell viability was assessed using a Cell Counting Kit-8 (CCK-8; Dojindo, Kumamoto Prefecture, Japan) according to the manufacturer's instructions. The optical density value at 450 nm was determined using a microplate reader (Molecular Devices, Sunnyvale, CA, USA).
Cy 09
Manufactured by MedChemExpress
Sourced in China, United States
CY-09 is a lab equipment product offered by MedChemExpress. It is a chemical compound used in various research applications. The core function of CY-09 is to serve as a research tool, but a detailed description of its intended use cannot be provided in an unbiased and factual manner without the risk of extrapolation.
Automatically generated - may contain errors
Lab products found in correlation
Cell counting kit 8 cck 8, by Dojindo Laboratories (1 mentions)
Microplate reader, by Molecular Devices (1 mentions)
4 pba, by MedChemExpress (1 mentions)
Il 1β, by Thermo Fisher Scientific (1 mentions)
M199 medium, by Merck Group (1 mentions)
Hrmecs, by Merck Group (1 mentions)
Penicillin, by Merck Group (1 mentions)
Streptomycin, by Merck Group (1 mentions)
D glucose, by Merck Group (1 mentions)
Sb203580, by MedChemExpress (1 mentions)
4 protocols using cy 09
1
Lupeol Attenuates LPS-Induced Neuroinflammation
2
Selective NLRP3 Inhibition with CY-09
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CY-09 is a selective and direct NLRP3 inhibitor that directly attaches to the ATP-binding motifs of the NLRP3 NACHT domain and thus blocks NLRP3 ATPase activity, inhibiting the assembly and NLRP3 inflammasomes activation. CY-09 (Medchem Express, HY-103666) prepared a 1 ml working solution. A 100 μL clarified DMSO (25.0 mg/mL) reserve solution was mixed with 400 μL PEG300, 50 μL Tween-80, and 450 μL NS was added. A solution ≥ 2.5 mg/mL (5.90 mM, saturation unknown) was obtained with this scheme. Intraperitoneal injections were given at 2.5 mg/kg once daily, 2 days before and 12 days after surgery for 15 days.
3
Inflammasome Regulation in Chondrocytes
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Daphnoretin (Article No.: HY-N0699), the NLRP3 inflammasome inhibitor CY-09 (Article No.: HY-103666), and the ERS inhibitor 4-PBA (Article No.: Hy-a0281) were acquired from MedChemExpress (SH, USA). Daphnoretin, CY-09, and 4-PBA were dissolved in 10% DMSO. Then, they were diluted in 0.9% normal saline (NS). In vitro, chondrocytes were treated with IL-1β (PeproTech Inc., NJ, USA) alone (10 ng/ml) or in combination with different doses of daphnoretin (0, 0.5, 1, 2, 4, 8, 16, 32 µM) for 48 h, and/or received 1 nM of 4-PBA or CY-09 treatment in the medium.
4
HRMECs Glucose Metabolism Regulation
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The HRMECs were purchased from Cell Systems, and were routinely cultured in M199 medium (EMD Millipore) supplemented with 100 units of penicillin and 100 µg of streptomycin (both from Sigma-Aldrich; Merck KGaA) per milliliter of medium. All cells (passages 5-12) were cultured in grade plastic-ware and maintained in an atmosphere of 5% CO2 at 37°C.
For the establishment of the HG cell models, the HRMECs were cultured in conditioned medium with 5 mM [serving as the normal glucose (NG) group] or 30 mM (the HG group) D-glucose (Sigma-Aldrich; Merck KGaA) and incubated at 37°C with 5% CO2; the HG group was then treated with or without 1 µM of the NLRP3 inhibitor, CY-09, 1 µM of the ASK1 inhibitor, GS-444217 or 10 µM of the p38 inhibitor, SB203580 (all from MedChemExpress) for 24 h, respectively. Each inhibitor was dissolved in dimethyl sulfoxide (DMSO) to a concentration of 50 mM for use as stock solutions that were diluted to the required concentrations for in vitro experiments.
For the establishment of the HG cell models, the HRMECs were cultured in conditioned medium with 5 mM [serving as the normal glucose (NG) group] or 30 mM (the HG group) D-glucose (Sigma-Aldrich; Merck KGaA) and incubated at 37°C with 5% CO2; the HG group was then treated with or without 1 µM of the NLRP3 inhibitor, CY-09, 1 µM of the ASK1 inhibitor, GS-444217 or 10 µM of the p38 inhibitor, SB203580 (all from MedChemExpress) for 24 h, respectively. Each inhibitor was dissolved in dimethyl sulfoxide (DMSO) to a concentration of 50 mM for use as stock solutions that were diluted to the required concentrations for in vitro experiments.
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