Primary astrocytes were stimulated with 500 ng/mL of rHMGB1 (Cat# 1690-HMB; R&D) in the presence or absence of 10 nM atractylenolide I (TLR4 antagonist; Cat# A2737; Sigma) or 10 nM C29 (TLR2 inhibitor; Cat# 363600-92-4, MCE, Monmouth Junction, NJ, USA) dissolved in 0.1% dimethyl sulfoxide for 24 hours. The culture supernatants were then collected, and the cells were subjected to lysis with a buffer (Beyotime). The lysates were centrifuged at 10,000 × g for 15 minutes. The PGE2 concentration in cell supernatants and lysates was measured using a PGE2 high-sensitivity enzyme-linked immunosorbent assay (ELISA) kit (Arbor Assays, Ann Arbor, MI, USA) according to the manufacturer’s instructions.
Atractylenolide 1
Manufactured by Merck Group
Sourced in United States
Atractylenolide I is a chemical compound found in various plant species. It is a naturally occurring sesquiterpene lactone. Atractylenolide I is commonly used as a reference standard and research tool in analytical chemistry and pharmacological studies.
Automatically generated - may contain errors
2 protocols using atractylenolide 1
1
Astrocyte HMGB1 Stimulation and TLR Modulation
2
Atractylenolide Analysis by HPLC
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Samples were analyzed by a reverse-phase HPLC system (Shimadzu 20A, Kyoto, Japan) that consisted of an autosampler (SIL-20A), a binary pump (LC-20AD), and a photodiode array detector (SPD-20A) and was equipped with a YMC-Triart C18 column (5 μm × 4.6 mm × 250 mm) (YMC, Kyoto, Japan). Gradient flows for the two-solvent system (solvent A, 0.05% phosphoric acid in water; solvent B, acetonitrile) were as follows: 85% A/15% B at 0 min, 85% A/15% B at 5 min, 50% A/50% B at 15 min, 50% A/50% B at 20 min, 40% A/60% B at 25 min, 40% A/60% B at 30 min, 15% A/85% at 35 min, 15% A/85% at 40 min, 85% A/15% at 42 min, and 85% A/15% at 45 min. The flow rate of the mobile phase was 1.0 ml/min with an injection volume of 10 μl. Detection was performed at 220 nm for atractylenolide III (Sigma, St. Louis, MO, USA) or at 280 nm for atractylenolide I (Sigma).
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