Cytokine regulation and the signaling mechanism of osteoclast inhibitory peptide-1 (OIP-1/hSca) to inhibit osteoclast formation

The osteoclast (OCL) is the primary bone resorbing cell. OCL formation and activity is regulated by local factors produced in the bone microenvironment. We recently identified OCL inhibitory peptide-1 (OIP-1/ hSca) as a novel inhibitor of OCL formation and bone resorption that is produced by OCLs. OIP-1 is a glycosylphosphatidyl-inositol (GPI)-linked membrane protein (16 kDa) related to the mouse Ly-6 family of hematopoietic proteins. OIP-1 mRNA is expressed in human OCL precursors, granulocyte-macrophage colony-forming unit (GM-CFU), bone marrow cells, and osteoblast cells. We used cycle-dependent reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, which further demonstrated that interferon-γ (IFN-γ) strongly enhanced OIP-1/hSca mRNA expression in bone marrow cells and GM-CFU. Similarly, interleukin (IL)-1β also enhanced OIP-1 mRNA expression in GM-CFU. To determine the participation of OIP-1 in IFN-γ inhibition of OCL formation, we tested the capacity of a neutralizing antibody specific to OIP-1 c-peptide to inhibit IFN-γ's effects on OCL-like cell differentiation of mouse macrophages, RAW 264.7 cells. Anti-OIP-1 c-peptide specific antibody partially neutralized IFN-γ inhibition of OCL differentiation. Furthermore, OIP-1 inhibited phospho-c-Jun (p-c-Jun) kinase activity in RAW 264.7 cells. However, OIP-1/ hSca did not affect NF-κB activation in these cells. Western blot analysis further demonstrated that OIP-1 significantly decreased TNF receptor associated factor 2 (TRAF-2) expression in RAW 264.7 cells. However, OIP-1 had no effect on TRAF-6 expression in these cells. These data show that IFN-γ enhances OIP-1/hSca expression in OCL precursors, GM-CFU, and that OIP-1 inhibits OCL formation through suppression of TRAF-2 and p-c-Jun kinase activity.