Pseudomonas aeruginosa lipopolysaccharide induces osteoclastogenesis through a toll-like receptor 4 mediated pathway in vitro and in vivo.

OBJECTIVES: Bacterial infections near bone result in localized inflammatory osteolysis, a significant complication of chronic ear infections. While many bacterial products may be involved, lipopolysaccharide (LPS) has been implicated as a major mediator of inflammation and osteolysis. However, the mechanisms by which LPS promotes bone resorption have not been clearly established. There is no consensus on whether LPS acts directly or indirectly on osteoclast precursors (bone marrow monocytes [BMM]) to induce bone resorption. In light of the role of Pseudomonas aeruginosa, in chronic ear infections, we investigated the effects of P. aeruginosa LPS on osteoclastogenesis in vivo and in vitro. METHODS: Wild-type C57BL/6J and toll-like receptor 4 knock-out (TLR4-/-) mice received subcutaneous calvarial injections of 250 mug of P. aeruginosa LPS or phosphate buffered saline (PBS) only (n = 5 per group). Osteoclastic bone resorption was assessed histologically. The effect of P. aeruginosa LPS on bone resorption was assessed in vitro using combinations of BMMs and osteoblasts with and without functional toll-like receptor 4 (TLR4). RESULTS: In vivo, P. aeruginosa LPS induced robust osteolysis, and this effect was completely abrogated in mice lacking expression of TLR4. In vitro, P. aeruginosa LPS failed to induce development of osteoclasts directly in BMMs. However, P. aeruginosa LPS did stimulate osteoclastogenesis in BMM-osteoblast cocultures. CONCLUSIONS: P. aeruginosa LPS acts indirectly through osteoblasts to induce bone resorption. Optimal osteoclastogenesis in vitro required functional TLR4 expression in both BMMs and osteoblasts.

Nitric oxide synthase I mediates osteoclast activity in vitro and in vivo.

Bone resorption is responsible for the morbidity associated with a number of inflammatory diseases such as rheumatoid arthritis, orthopedic implant osteolysis, periodontitis and aural cholesteatoma. Previous studies have established nitric oxide (NO) as a potentially important mediator of bone resorption. NO is a unique intercellular and intracellular signaling molecule involved in many physiologic and pathologic pathways. NO is generated from L-arginine by the enzyme nitric oxide synthase (NOS). There are three known isoforms of NOS with distinct cellular distributions. In this study, we have used mice with targeted deletions in each of these isoforms to establish a role for these enzymes in the regulation of bone resorption in vivo and in vitro. In a murine model of particle induced osteolysis, NOS I-/- mice demonstrated a significantly reduced osteoclast response. In vitro, osteoclasts derived from NOS I-/- mice were larger than wild type controls but demonstrated decreased resorption. Although NOS I has been demonstrated in osteoblasts and osteocytes as a mediator of adaptive bone remodeling, it has not previously been identified in osteoclasts. These results demonstrate a critical role for NOS I in inflammatory bone resorption and osteoclast function in vitro.