Nox4 expression in osteo-progenitors controls bone development in mice during early life.

Tightly regulated and cell-specific NADPH-oxidases (Nox) represent one of the major sources of reactive oxygen species (ROS) signaling molecules that are involved in tissue development and stem cell self-renewal. We have characterized the role of Nox4 in osteo-progenitors during postnatal bone development. Nox4 expression in bone and ROS generation were increased during early osteoblast differentiation and bone development. Stromal osteoblastic cell self-renewal, proliferation and ROS production were significantly lower in samples from whole-body Nox4 knockout mice (Nox4) and conditional knockout (CKO) mice with depletion of Nox4 in the limb bud mesenchyme compared with those from control mice (Nox4), but they were reversed after 9 passages. In both sexes, bone volume, trabecular number and bone mineral density were significantly lower in 3-week old CKO and Nox4 mice compared with Nox4 controls. This was reflected in serum levels of bone formation markers alkaline phosphatase (ALP) and procollagen 1 intact N-terminal propeptide (P1NP). However, under-developed bone formation in 3-week old CKO and Nox4 mice quickly caught up to levels of control mice by 6-week of age, remained no different at 13-week of age, and was reversed in 32-week old male mice. Osteoclastogenesis showed no differences among groups, however, CTX1 reflecting osteoclast activity was significantly higher in 3-week old male CKO and Nox4 mice compared with control mice, and significantly lower in 32-week old Nox4 mice compared with control mice. These data suggest that Nox4 expression and ROS signaling in bone and osteoblastic cells coordinately play an important role in osteoblast differentiation, proliferation and maturation.