Figure 1.

Increased frequency of osteoclast progenitors (OCPs) in B6 mice with collagen-induced arthritis (CIA). (A) Experimental timeline for CIA model in B6 mice. To induce arthritis, mice were immunized with chicken collagen type II emulsified with complete Freund’s adjuvant (CFA) at day 0, followed by a booster dose at day 21 (CIA group). Likewise, CFA control group received only adjuvant, and control (ctrl) group was left untreated. Three time points during arthritis development were selected for analysis. (B) Gating strategy for flow cytometric analysis of OCPs in medullar compartment [distal tibia bone marrow (periarticular, PBM) and tarsometatarsal joints (TMT)], bearing the phenotype CD45⁺CD3^-B220^-NK1.1^-CD11b^-/loCD115⁺, and peripheral compartment [spleen (SPL) and peripheral blood (PBL)], bearing the phenotype CD45⁺CD3^-B220^-NK1.1^-CD11b⁺CD115⁺. Representative dot plots for PBM and SPL are shown. (C) Frequency of OCPs at different time points (25, 35, and 45 days after primary immunization) in analyzed tissues of ctrl, CFA, and CIA mice, determined using the previously described gating strategy. The OCP percentage of CD45⁺ population is presented. (D) Immunophenotyping of chemokine receptors (CCR2, CCR3, CCR5, CCR9, CXCR4, and CX3CR1) on PBM and SPL OCPs (35 days after primary immunization). The percentage of chemokine receptor-expressing OCPs of CD45⁺ population is presented. The experiments were repeated three times and pooled data are shown (n = 6–10 mice per group). Values are presented as medians (middle horizontal lines), boxes represent the interquartile range (IQR), whiskers represent 1.5 times the IQR. Statistically significant difference was determined at p < 0.05, Mann–Whitney U-test (D), or Kruskal–Wallis test (p-value marked on the graph) with Conover post-hoc test for group-to-group comparisons (C); correction for multiple comparisons (D) was performed using Holm–Bonferroni method; line denotes significant difference between groups. LY—CD3/B220/NK1.1.