[Effect of staphylococcal lipoteichoic acid on differentiation of RAW264.7 cells into osteoclasts].

Objective: To investigate the effect of staphylococcal lipoteichoic acid (LTA-sa) on RAW264.7 cells differentiation into osteoclasts. Methods: RAW264.7 cells were cultured with LTA-sa of 100 ng/mL (group A), LTA-sa of 200 ng/mL (group B), LTA-sa of 400 ng/mL (group C), receptor activator of nuclear factor κB (NF-κB) ligand (RANKL) of 100 ng/mL as positive control (group D), and equal volume of PBS as blank control (group E) respectively for 5 days. And then, tartrate resistant acid phosphatase staining (TRAP) was used to detect the formation of osteoclast-like cells, Image-Pro Plus 6.0 software to measure the areas of bone resorption pits in Corning Osteo Assay Surface (COAS) wells, and MTT assay to observe the proliferation activity of RAW264.7 cells in group A, B, C, and E. Results: After cultured for 5 days, the formation of osteoclast-like cells and bone resorption pits were observed in all groups. The number of osteoclast-like cells and the area of bone resorption pits in groups A, B, C, and D were more than those in group E. And with the increased concentration of LTA-sa, the indexes in groups A, B, and C increased gradually, but were lower than those in group D, and differences were significant between groups ( P<0.05). At 5 days after culture, there was no significant difference in absorbance value among the experimental groups (groups A, B, C, and E) ( P>0.05). Conclusion: LTA-sa has promoting effect on RAW264.7 cells differentiation into osteoclasts.

Molecular mechanism of osteoclast differentiation induced by staphylococcal lipoteichoic acid / 实用医学杂志

Objective To investigate the molecular mechanism of osteoclast differentiation induced by staphylococcal lipoteichoic acid (LTA-sa). Methods Raw264.7 cells were treated with LTA-sa in a concentration of 200 ng/mL for 0, 5, 10, 20, 40, 60 min and 0, 1, 2, 3 days respectively, and the proteins in signaling pathways associated with osteoclast differentiation were measured with western blot. In addition, Raw264.7 cells were treated with different concentrations of LTA-sa (100, 200 and 400 ng/mL) and PBS for 0, 1, 2, 3 days, the expression of TNF-α, IL-1α and IL-6 was detected with Enzyme linked immunosorbent assay (ELISA). Results (1)Western blot showed that, under stimulation of LTA-sa, IκB-α decreased at 5 min and 10 min, while the phosphorylation of nuclear factor κB increased at 10 min . In addition , NFATc1 increased in 2 and 3 days gradually. The above results were statistically analyzed, and the difference was significant in statistics (P < 0.001). (2)ELISA showed that the expression of IL-6 increased in 2 and 3 days along with the increasing concentration and prolonging stimulation time of LTA-sa. Data were statistically analyzed, the difference was significant in statistics (P < 0.001). Conclusion LTA-sa promotes osteoclast differentiation through the NF-κB signaling pathway and the secretion of IL-6.