Colivelin

For 48 hours, 2×10⁵ CD11b⁺Ly6G⁻ cells isolated from the BM of either EGFP‐Spp1‐knock‐in reporter or WT mice were cultured in 100 pmol/L Colivelin (R&D Systems, Minneapolis, MN).

Melatonin was purchased from Sigma-Aldrich Chemical Co. (St. Louis, MO, USA). Fluorescein sodium was acquired from Alcon Laboratories, Inc. (Fort Worth, TX, USA). Colivelin was obtained from R&D Systems (Minneapolis, MN, USA). The antibodies for Western blot and immunofluorescence in this study are listed in Supplementary Table S1.

Pexmetinib was from Selleck Chemicals (Houston, Texas, USA). DMSO were purchased from Sigma-Aldrich (St. Louis, MO, USA). α-MEM (Eagle’s minimal essential medium with Alpha Modification), dulbecco’s modified eagle medium (DMEM), fetal bovine serum (FBS), and penicillin/streptomycin were purchased from Gibco-BRL. CCK-8 (Cell Counting Kit-8) was obtained from Dojindo Molecular Technology (Kumamoto, Japan). Recombinant soluble mouse M−CSF, mouse RANKL and Colivelin were obtained from R&D Systems (Minneapolis, MN, USA). All experiments were performed in the absence of visible light to prevent photosensitivity. Pexmetinib was diluted in cell culture medium so that DMSO comprised < 0.1% of the total volume. Meanwhile, 0.1% DMSO as a control group in vitro. Specific antibodies against ERK (extracellular signal-regulated kinase, 1:1000), JNK (c-Jun N-terminal kinase, 1:1000), p38 (1:1000), phosphorylated p-ERK (Thr202/Tyr204, 1:1000), p-JNK (Thr183/Tyr185, 1:1000), p-p38 (Thr180/Tyr182, 1:1000), STAT3 (1:1000), p-STAT3 (Tyr705, 1:1000), NFATc1 (1:1000) and GAPDH (1:10000) were obtained from Cell Signaling Technology (Boston, USA), and antibodies against TRAP (1:1000), MMP-2 (1:1000) and MMP-9 (1:1000) were purchased from Abcam (Cambridge, UK). TRAP staining kit and all other reagents were purchased from Sigma-Aldrich, unless otherwise stated.