CONICET | Buscador de Institutos y Recursos Humanos

During rheumatoid arthritis, macrophages differentiate into osteoclasts and bone destruction occurs. We demonstrated that extracts from mouse placenta (EPs), containing Th1 and Th2 cytokines; regulate the osteoclast differentiation of RAW 264.7 macrophages. Now we analyze, whether IL-10, TGF-beta, as well as the NFATc1 transcription factor are involved in such effect. Placental extracts were obtained at 7 days of pregnancy (EP7). RAW cells were cultured with RANKL and M-CSF (RM) in the presence or absence of EP7, and neutralizing antibodies (NA) against IL-10 and TGF-beta; the differentiation of the cells was determined by assessing multinucleated, tartrate resistent acid phosphatase positive cells (TRAP+), using a commercial kit; and matrix metalloprotease (MMP) activity by zymogram. The effect of EP7 on NFATc1 expression /phosphorylation was assessed by western blot. EP7 inhibited the RM-induced MMP activity and TRAP+ (inhibition vs RM: 68%, p<0.001 and 91%, p<0.001, respectively). Pre-incubation of RAW cells with a-IL-10 NA reverted the inhibitory effect of EP7 on MMP activity (a-IL10+RM vs a-IL10+RM+EP7, p>0.05) while the inhibition of TRAP+ was not affected (a-IL10+RM vs a-IL10+RM+EP7, p<0.01). Pre-incubation with a-TGF-beta NA reverted the inhibitory effect of EP7 on both, the MMP activity and the TRAP+ (a-TGF-beta+RM vs a-TGF-beta+RM+EP7, p>0.05). EP7 inhibited in 79 % the expressión of NFATc1 in cells cultured during 4 days with RM (p<0.001); nevetheless, the NFATc1 dephosphorylation induced by 2 h-treatment of the cells with RM was not affected by EP. We propose that the downregulation of NFATc1 expression by EP7 is involved in the inhibition of the RM-induced osteoclast differentiation of RAW cells. TGF-beta participates in the inhibition of MMP and TRAP+. The inhibitory effect of IL-10 on MMP activity was not enough to inhibit cell differentiation, since a-IL-10 NA was not able to revert the inhibition by EP7 of RM-induced TRAP+.