ABSTRACT
Objective To explore the inhibitive effect of BSP on Caspase dependent apoptosis of preosteoclasts RAW264.7 cells by bone sialoprotein (BSP).Methods RAW264.7 cells were treated with the inhibitors of signaling pathway molecules and/or BSP.The activities of caspase3/8/9 in RAW264.7 cells were detected with caspase3/7/8/9 kits and the interaction between Caspase-8 and integrin αVβ3 was detected by Co-IP.Results The activities of Caspase3/8/9 in BSP-treated RAW264.7 cells were decreased and the apoptosis of RAW264.7 cells were inhibited significantly.In addition,there was a positive correlation between caspase-8 activity and caspase-3 activity.There was interaction between caspase-8 and integrin αVβ3.Conclusion The anti-apoptotic effect of BSP on RAW264.7 cells is mainly through the regulation of exogenous apoptosis pathway and the apoptosis induced by caspase-8 is inhibited by the binding of BSP and αvβ3.
ABSTRACT
Objective To explore the key signaling pathways and molecules of bone sialoprotein (BSP) to promote proliferation and differentiation of preosteoclasts RAW264.7. Methods RAW264.7 cells were treated with BSP and the inhibitors of signaling pathway molecules. MTS and TRAP staining kits were used to evaluate the effects of proliferation and differentiation. The activity assay kits of Calcineurin, AKT, JNK, ERK and p38 were used to detect their activities. Results Inhibiting ERK and p38 can inhibit cell proliferation induced by BSP significantly, while inhibiting AKT, Calcineurin and PI3K can reduce the role of promoting the differentiation by BSP. With increasing treatment time of BSP, the activity of Calcineurin in RAW264.7 cells increased. Furthermore, the activity of AKT was maximum at 48 h after treated with BSP, while the activity of JNK, ERK and p38 were maximum at 24 h after treated with BSP. Conclusions BSP mainly regulates the ERK and p38 of MAPK pathway to promote RAW264.7 cell proliferation, and regulates AKT, PI3K and Calcineurin pathways to promote RAW264.7 cell differentiation.