T6167923

We assessed macrophage transcripts from THP + GF alone and THP + GF + SMV. We utilized simvastatin (a synthetic derivative of a fermentation product from Aspergillus terreus) as a representative statin. Briefly, macrophage cultures were pretreated with simvastatin (MedChemExpress, 10µM, 18 h prior to polarization protocol), followed by complete media change before initiating CCL2 treatment (as shown above). Notably, simvastatin was replenished at each time point of introducing new media throughout the protocol. MyD88 homodimerization inhibitor, T6167923 [30 (link)] (MedChemExpress, 10 µM, 18 h prior to polarization protocol).

The BV2 mouse microglia cell line (FH0355) and HT22 mouse hippocampal neuron cell line (FH1027) was both purchased from Shanghai FuHeng Biology Co., Ltd. in China. On the first day, BV2 cells were maintained in Dulbecco’s modified Eagle’s medium (DMEM) (Gibco) supplemented with 10% fetal bovine serum (FBS) (Sigma–Aldrich, MO, USA) and 1% penicillin/streptomycin (Gibco) in humidified 5% CO₂ air environment at 37 °C. On the second day, the BV2 cell culture medium was replaced, and the cells were divided into 4 groups: 1) the Control group (BV2), in which BV2 cells were incubated with medium only for 24 h; 2) the Heme group (BV2 + Heme) in which BV2 cells were treated with 40 μM hemin (H9039, Sigma, USA) for 24 h; 3) the MyD88 inhibition group (BV2 + Heme + T6167923) in which BV2 cells were treated with 40 μM hemin and 2 μM MyD88 inhibitor (T6167923, #HY-19744, MedChemExpress, USA) for 24 h; and 4) the NF-κB inhibition group (BV2 + Heme + JSH-23), in which BV2 cells were treated with 40 μM hemin and 20 μM NF-κB inhibitor (JSH-23, #S7351, Selleck, USA) for 24 h. Then, the BV2 cells were harvested for subsequent experiments.