LPS and TNF-alpha Regulate Cytokine and BMP-2 Expression in Osteoblasts

Osteoblasts are responsible for bone formation and also regulate osteoclasts through expression of cytokines and growth factors. Therefore, they play an important role in normal bone physiology and development of pathology. In inflammatory conditions such as alveolar bone in periodontitis and synovial joints in rheumatoid arthritis, resident osteoblasts are bombarded with bacterial lipopolysaccharide (LPS) and pro-inflammatory cytokines, such as TNF-alpha. Objective: To investigate the effect of the addition of LPS and TNF-alpha to human and mouse osteoblast cell lines on RANKL, IL-15, IL-23 and BMP-2 expression. Method: Human (MG63) and mouse (MC3T3) osteoblast cells were cultured and either unstimulated or stimulated with 10 ng/ml LPS or 5 ng/ml human or mouse TNF-alpha respectively for 16 hours. Total RNA was purified and RT-PCR perfomed using specific primers for RANKL, IL-15, IL-23 and BMP-2. MC3T3 cells were also stimulated with 10 ng/ml LPS for 48 hours, culture supernatant collected and cytokines measured by ELISA. Results: 1)LPS and TNF-alpha upregulated RANKL and BMP-2 gene expression in the human and mouse osteoblast cell lines, although the effect of TNF-&alpha was greater than LPS. 2)Unstimulated MG63 cells did not express IL-15. LPS, but not TNF-alpha induced IL-15 expression. 3)LPS and TNF-alpha induced IL-23p19 mRNA expression by MG63 and MC3T3 cells. IL-23 expression was also detected in supernatant collected from LPS-stimulated osteoblasts, but not unstimulated osteoblasts. We report here, for the first time, the detection of IL-23 expression from osteoblast cells induced by soluble inflammatory factors. Conclusion: These results suggested that osteoblasts are not only bone generators, but also play an important role in innate immunity by producing cytokines and growth factors. The results are of value in understanding the role of osteoblasts in inflammatory bone destruction and will contribute to current therapeutic developments for periodontitis. This research was funded by Arthritis Research Campaign (Ref:ARC_13928,ARC_ARC-17212).