Differentiation Of RAW264 Cells Research Articles

The effect of mesenchymal stem cells on osteoclast precursor cell differentiation.

Transplantation of mesenchymal stem cells (MSCs) has been extensively studied in the field of regenerative medicine. Bone regeneration is achieved via the interaction of osteoblasts and osteoclasts. However, the influence of MSCs on osteoclasts is unknown. The purpose of this study was to investigate the effect of MSCs on the expression of genes for osteoclast differentiation factors using qPCR after indirect co-culture of MSCs and RAW264 cells. The numbers of osteoclasts after addition of soluble receptor activator of nuclear factor kappa B (NF-κB) ligand (sRANKL) were also compared. Expression of osteoprotegerin (OPG) by MSCs was significantly elevated in co-culture over time. The differentiation of RAW264 cells into mature osteoclasts following addition of sRANKL was significantly inhibited by co-culture with MSCs. Expression of RANK, colony stimulating factor 1 receptor, NF-κB, and nuclear factor of activated T-cell cytoplasmic 1 in RAW264 cells was significantly inhibited by co-culture with MSCs. Expression of OPG protein was higher in co-culture with RAW264 cells than in MSCs alone, and the expression level was clearly higher than that of RANKL. MSCs appeared to inhibit osteoclast differentiation via expression of OPG.

Direct effects of estrogen on differentiation and apoptosis of osteoclasts

Osteoclasts play a crucial role in bone resorption. Since osteoclast differentiation/activation is involved in orthodontic tooth movement at the compression sites, the investigation on osteoclasts is very important to the field of orthodontics. It is well known that estrogen has the protective effect on bone. However, the mechanisms by which estrogen prevents bone loss remain to be elucidated. Although estrogen was recently reported to induce apoptosis of osteoclasts, the precise mechanisms of estrogen-induced osteoclast apoptosis remained controversial with regard to whether estrogen affects osteoclasts directly or not. Here we investigated whether estrogen directly induces differentiation and apoptosis of osteoclasts in vitro using mouse monocytic RAW264 cells differentiated into osteoclasts by RANKL. It was observed that estrogen inhibited RANKL-induced osteoclast differentiation of RAW264 cells in a dose-dependent manner. Estrogen suppressed p38 phosphorylation while it enhanced ERK phosphorylation induced by RANKL, suggesting that modulation of MAPK signaling may be involved in inhibition of osteoclast differentiation by estrogen. Next, it was shown that estrogen dose-dependently augmented caspase-3 activation in osteoclasts differentiated from RAW264 cells by RANKL, demonstrating that estrogen directly enhanced apoptosis of osteoclasts. Estrogen-induced caspase-3 activation was attenuated by ICI 182,780, suggesting that the effects of estrogen on osteoclast apoptosis is mediated through estrogen receptors. Thus, these results suggest that estrogen may directly inhibit differentiation and induce apoptosis of osteoclasts.

The Effect of Methyl-β-cyclodextrin on the Differentiation of RAW264 Cells into Osteoclasts

The Effect of Methyl-β-cyclodextrin on the Differentiation of RAW264 Cells into Osteoclasts

Induction of mouse c- src in RAW264 cells is dependent on AP-1 and NF-κB and important for progression to multinucleated cell formation

c- src is known to play an essential role in osteoclastogenesis. We studied the regulatory mechanism as well as the significance of c- src induction in RANKL-induced differentiation of mouse monocytic RAW264 cells to TRAP-positive-multinucleated cells. We determined the genomic organization of the 5′-terminal region of mouse c- src. Mutational and biochemical analyses in the region 0.9 kb upstream of the transcription start site revealed that c-Fos and JNK pathways, in addition to NF-κB, participate in c- src induction in response to RANKL. On the other hand, when the expression of c- src was suppressed by introducing antisense src, the number of multinucleated cells formed was significantly reduced. Together, these findings show that the expression of c- src is under the control of AP-1 and NF-κB in the differentiation of RAW264 cells and that c- src plays an essential role at the stage of progression to multinucleated cell formation.