MIP-1α and IL-6 do not increase RANKL expression in human bone marrow and RANK-Fc does not inhibit MIP-1α–stimulated osteoclast formation.

(A) Cells were incubated for 48 hours with media, 1,25-(OH)₂D₃ (10⁻⁸ M), IL-6 (100 pg/mL), MIP-1α (200 pg/mL), or a combination of MIP-1α and IL-6. The cells were then collected and the lysate subjected to Western blot analysis. 1,25-(OH)₂D₃ increased RANKL expression in human bone marrow cultures. IL-6, MIP-1α, or IL-6 plus MIP-1α did not significantly increase RANKL expression in human bone marrow. Expression of β-actin is shown as a control. There was no effect on β-actin expression. (B) CFU-GM–derived cells were prepared and then treated with RANKL (50 ng/mL) or MIP-1α (200 pg/mL) in the presence of varying concentrations of recombinant RANK-Fc (0-100 ng/mL). RANK-Fc significantly decreased RANKL-stimulated osteoclast formation in human marrow cultures. In contrast, RANK-Fc modestly affected or did not affect osteoclast-like cell formation stimulated by MIP-1α. High concentrations of RANK-Fc (100 ng/mL) decreased basal osteoclast formation in these cultures. Results represent the mean ± SEM for quadruplicate determinations for a typical experiment. Similar results were seen in 2 independent experiments.