Effects of activated T cells on osteoclastogenesis depend on how they are activated

Introduction: Activated T cells are emerging as important regulators of osteoclast function in inflammatory diseases. Both pro- and anti-resorptive properties have been described. We reasoned that this reported variability of the effects of T cells on osteoclast formation depends on how T cells are activated in vitro. Methods: We harvested T lymphocytes from 5-week-old C57BL/6 mouse spleens. Activation was performed with anti-CD3 and -CD28 Ab (Abs), concanavalin A (Con A), phytohemagglutinin (PHA), or the superantigen Staphylococcal enterotoxin A (SEA). Osteoclastogenesis was induced by receptor activator of NF-κB ligand (RANKL) in the mouse macrophage cell line RAW 264.7 cells, or primary macrophage CD11b+ cells from mouse spleen. Cells were cultured with T cells or with their conditioned medium. Results: Co-culture of activated T lymphocytes with RAW 264.7 cells inhibited osteoclastogenesis but only when activated by Abs. This effect was CD4+-dependent. Conditioned medium from activated T lymphocytes with Abs consistently blocked osteoclastogenesis in RAW 264.7 and CD11b+ cells. T cells activated with SEA, Con A, and PHA had inconsistent effects on osteoclastogenesis. We then tested the role of interferon (IFN)-γ, a known inhibitor of osteoclastogenesis, in the effects of T cells on osteoclast formation. IFN-γ neutralizing antibody blocked the inhibitory effect of T-cell conditioned medium on osteoclastogenesis. Osteoclast precursors from IFN-γ receptor-null mice treated with 0.1% medium from activated T cells formed osteoclasts. However, higher doses of medium inhibited osteoclastogenesis, so that we cannot exclude that other factors besides IFN-γ may be involved. Conclusions: Available methods to activate T lymphocytes result in variable effects on osteoclastogenesis. IFN-γ is the main factor responsible for the inhibitory effects of activated T cells on osteoclast formation.