Receptor activator of nuclear factor-kappa B (RANK) ligand (RANKL) binds RANK on the surface of osteoclast precursors to trigger osteoclastogenesis. Recent studies have indicated that osteocytic RANKL has an important role in osteoclastogenesis during bone remodelling; however, the role of osteoblastic RANKL remains unclear. Here we show that vesicular RANK, which is secreted from the maturing osteoclasts, binds osteoblastic RANKL and promotes bone formation by triggering RANKL reverse signalling, which activates Runt-related transcription factor 2 (Runx2). The proline-rich motif in the RANKL cytoplasmic tail is required for reverse signalling, and a RANKL(Pro29Ala) point mutation reduces activation of the reverse signalling pathway. The coupling of bone resorption and formation is disrupted in RANKL(Pro29Ala) mutant mice, indicating that osteoblastic RANKL functions as a coupling signal acceptor that recognizes vesicular RANK. RANKL reverse signalling is therefore a potential pharmacological target for avoiding the reduced bone formation associated with inhibition of osteoclastogenesis.

Excessive exposure to glucocorticoids causes osteoporosis in children and adults. Occlusal disharmony is known to induce an increase in serum corticosteroid levels in murine models, but the influence of occlusal disharmony-induced stress on the bone mass during the growth period has not yet been clarified. The purpose of this study was to investigate whether occlusal disharmony-induced stress decreases bone mass. Five-week-old C57BL/6J male mice were used. A 0.5-mm increase in the vertical height of occlusion was used to induce occlusal disharmony for a period of 7 days. Serum corticosterone levels were significantly higher on post-induction day 7, with radiological evidence of osteopenia of the third lumbar vertebra and long bones of the hind limbs. Osteopenia was associated with a reduction of the mechanical properties of the tibia and femur, with significant suppression of bone formation parameters and an increase in bone resorption parameters, as evaluated by bone histomorphometric analysis of the tibial/femur metaphysis. Our findings at the level of bones were supported by our assessment of serum markers of systemic metabolism. Therefore, occlusal disharmony-induced stress may lead to osteopenia and reduce the mechanical strength of bone through an increase in serum glucocorticoid levels in mice.Occlusal disharmony frequently occurs during growth due to mixed dentition, as the primary dentition is changing to permanent dentition. Several lines of evidence indicate that occlusal disharmony increases serum corticosteroid levels in young animals. As the serum corticosteroid level is considered to be a marker of 'stress' in rodents, it is plausible that occlusal disharmony would cause stress 1-4 . The high frequency of bruxism (teeth grinding) during the period of growth further suggests the possible existence of occlusal disharmony-induced stress in childhood [5][6][7] . By contrast, it is well known that excessive exposure to glucocorticoids causes osteopenia 8 . As an example, the use of steroid medications in children for the treatment of immune-related diseases, such as asthma and skin allergies, commonly results in bone-related side effects, including fractures and slowed bone growth 9 . Given the significant effects of stress-induced hormone on bone metabolism 10 , evaluation of the effect of occlusal disharmony-induced stress on skeletal tissue is warranted.Our aim in this study was to use a mouse model of occlusal disharmony to evaluate the effects of occlusal disharmony-induced stress on bone mass and bone mineral density, as well as on the mechanical properties of the long bones and lumbar vertebrae, which are expected to be differentially affected by mechanical loading [11][12][13] . ResultsIncrease in serum corticosterone level with occlusal disharmony. Serum corticosterone levels were measured at baseline and after the induction of occlusal disharmony (Fig. 1). Serum corticosterone levels were significantly higher in the disharmony than in the control group on post-induction days 1, 3 and 7 (Fig. ...

IntroductionWe designed OP3-4 (YCEIEFCYLIR), a cyclic peptide, to mimic the soluble osteoprotegerin (OPG), and was proven to bind to RANKL (receptor activator of NF-κB ligand), thereby inhibiting osteoclastogenesis. We recently found that another RANKL binding peptide, W9, could accelerate bone formation by affecting RANKL signaling in osteoblasts. We herein demonstrate the effects of OP3-4 on bone formation and bone loss in a murine model of rheumatoid arthritis.MethodsTwenty-four seven-week-old male DBA/1J mice were used to generate a murine model of collagen-induced arthritis (CIA). Then, vehicle or OP3-4 (9 mg/kg/day or 18 mg/kg/day) was subcutaneously infused using infusion pumps for three weeks beginning seven days after the second immunization. The arthritis score was assessed, and the mice were sacrificed on day 49. Thereafter, radiographic, histological and biochemical analyses were performed.ResultsThe OP3-4 treatment did not significantly inhibit the CIA-induced arthritis, but limited bone loss. Micro-CT images and quantitative measurements of the bone mineral density revealed that 18 mg/kg/day OP3-4 prevented the CIA-induced bone loss at both articular and periarticular sites of tibiae. As expected, OP3-4 significantly reduced the CIA-induced serum CTX levels, a marker of bone resorption. Interestingly, the bone histomorphometric analyses using undecalcified sections showed that OP3-4 prevented the CIA-induced reduction of bone formation-related parameters at the periarticular sites.ConclusionThe peptide that mimicked OPG prevented inflammatory bone loss by inhibiting bone resorption and stimulating bone formation. It could therefore be a useful template for the development of small molecule drugs for inflammatory bone loss.Electronic supplementary materialThe online version of this article (doi:10.1186/s13075-015-0753-8) contains supplementary material, which is available to authorized users.

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