429373.fig.002a
(a)
429373.fig.002b
(b)
Figure 2: OC-iTcREG suppress osteoclast differentiation and actin ring formation: (a) OC-iTcREG were generated from polyclonal wt, IL-10−/−, IL-6−/−, or IFN-γ −/− splenic T cells and cocultured with osteoclast precursors in the presence of RANKL and M-CSF for 5 days. The T cells were then removed and the adherent cells were assayed for TRAP activity using a fluorescent substrate ELF97. IFN-γ and IL-6 were found to be necessary for OC-iTcREG anti-osteoclastogenic activity. (b) Osteoclasts are differentiated on bovine bone slices and then cultured for 24 hrs with wt, IL-10−/−, IL-6−/−, or IFN-γ −/− OC-iTcREG. The T cells were then removed and the osteoclasts were stained with fluorophore-conjugated phalloidin to assay for actin ring formation. IFN-γ was shown to be required for the OC-iTcREG to suppress actin ring formation.