MODULATION OF OSTEOCLAST DIFFERENTIATION BY PLACENTAL CYTOKINES

CUSTIDIANO A.; SACERDOTI F.; DE LEÓN, R.; GENTILE, T.; CANELLADA A.

BUENOS AIRES

Congreso; First French - Argentine Immunology Congress; 2010

Sociedad Argentina de Inmunología & French Society of Immunology

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 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 enought to inhibit cell differentiation, since a-IL-10 NA was not able to revert the inhibition by EP7 of RM-induced TRAP+.