THP-1 cells in the exponential growth phase were seeded in six-well plates at a concentration of 1 × 10 6 cells/ml. After 24 h, PMA (at a concentration of 100ng/ml) was added to the cells. After 48 h of stimulation, the supernatants were collected and stored in a refrigerator until further analysis. The cell pellets were lysed, followed by protein extraction, and the extracted protein samples were stored at -80°C until further analysis. The RAW264.7 cells were also seeded in six-well plates at a concentration of 1 × 10 6 cells/ml. After 24 h, LPS (at a concentration of 100ng/ml) was added to stimulate the cells for 48 h, and the supernatants and the cell pellets were collected. Human-TNF-a-Forward:5'-CTTCTCGAACCCCGAGTGAC-3'
Human-TNF-a-Reverse:5'-ATGAGGTACAGGCCCTCTGA-3'
The serum levels of the cytokines interleukin (IL)-6, TNF-α, and IL-1 were quanti ed using a LegendPlex TM Multiplex cytokine bead-based assay (BioLegend, San Diego, CA) following the manufacturer's instructions. For each reaction, 25 µl of serum was diluted with 125 µl deionized water. All samples were run on an Accuri C6 ow cytometer (BD Biosciences, San Diego, CA) and acquired data were analyzed using LegendPlex v7.0 software.
Human-TNF-a-Reverse:5'-ATGAGGTACAGGCCCTCTGA-3'
The serum levels of the cytokines interleukin (IL)-6, TNF-α, and IL-1 were quanti ed using a LegendPlex TM Multiplex cytokine bead-based assay (BioLegend, San Diego, CA) following the manufacturer's instructions. For each reaction, 25 µl of serum was diluted with 125 µl deionized water. All samples were run on an Accuri C6 ow cytometer (BD Biosciences, San Diego, CA) and acquired data were analyzed using LegendPlex v7.0 software.