The inducible nitric oxide synthase pathway promotes osteoclastogenesis under hypoxic culture conditions.

Bone homeostasis depends on the balance between bone resorption by osteoclasts and bone formation by osteoblasts. Bone resorption can become excessive under various pathological conditions, including rheumatoid arthritis. Our previous studies have shown that osteoclast formation is promoted under hypoxia. However, the precise mechanisms behind osteoclast formation under hypoxia have not been elucidated. The present study investigated the role of inducible nitric oxide synthase (iNOS) on osteoclast differentiation under hypoxia. Primary bone marrow cells obtained from mice were stimulated with receptor activator of nuclear factor-kappa B ligand and macrophage colony-stimulating factor to induce osteoclast differentiation. The number of osteoclasts was increased in the culture under hypoxia (oxygen concentration, 5%) compared with that in the culture under normoxia (oxygen concentration, 20%). iNOS gene and protein expression increased in the culture under hypoxia. Addition of an iNOS inhibitor under hypoxic conditions suppressed osteoclastogenesis. Addition of a nitric oxide (NO) donor to the normoxic culture promoted osteoclastogenesis. Furthermore, insulin-like growth factor 2 expression was significantly altered in both iNOS inhibition experiments and NO donor experiments. These data might provide clues to therapies for excessive osteoclastogenesis under several hypoxic pathological conditions, including rheumatoid arthritis.