Recombinant human il 1β protein

THP-1 (human monocyte) cells were purchased from the Shanghai Institute of Cell Biology, China. For osteoclast differentiation, 3 × 10⁴ THP-1 cells were seeded in each well of 48-well plates and cultured with osteoclast differentiation medium, according to the previous studies (Li et al., 2017 (link); Ren et al., 2019 (link)). In some osteoclastogenesis assays, THP-1 cells were treated with recombinant human IL-1β protein (R&D, United States) at the indicated concentration every 2 days. After 14 days of induction, cells were fixed with 4% PFA and stained with TRAP staining solution (Jiancheng Biotech, China). TRAP-positive cells with three or more nuclei were considered osteoclasts, and the number of osteoclasts in each well was counted under a microscope.

To induce osteoblast differentiation, a total of 1 × 10⁵ PDLSCs were seeded into each well of 6-well plates. Upon reaching a density of 70%, the PDLSCs were grown in osteogenic differentiation medium (Cyagen, United States) for 21 days. In some osteogenic induction assays, the PDLSCs were treated with recombinant human IL-1β protein (R&D, United States) at the indicated concentration every 2 days. After 21 days, the cells were fixed with 4% PFA in PBS and stained with 2% Alizarin red (pH = 4.2).

For adipogenic induction, PDLSCs were plated in 6-well plates. When the PDLSCs reached 100% confluence, the growth medium was changed to adipogenic differentiation medium (Cyagen, United States), and the manufacturer’s protocol was followed. In some adipogenic induction assays, PDLSCs were treated with recombinant human IL-1β protein (R&D, United States) at the indicated concentration every 2 days. After 21 days, the cells were fixed in 4% PFA for 30 min and stained with Oil red O.