Donepezil, an inhibitor of acetylcholinesterase (AChE) targeting the brain, is a common medication for Alzheimer's disease. Interestingly, a recent clinical study found that administration of this agent is associated with lower risk of hip fracture independently of falling, suggesting its direct effect on bone tissues as well. AChE has been reported to be involved in osteoblast function, but the role of AChE on osteoclastogenesis still remains unclear. We analyzed the effect of AChE and donepezil on osteoclastogenesis in vivo and in vitro. Cell-based assays were conducted using osteoclasts generated in cultures of murine bone marrow macrophages (BMMs) with receptor activator of nuclear factor-kappa B ligand (RANKL). The effect of donepezil was also determined in vivo using a mouse model of RANKL-induced bone loss. Recombinant AChE in BMMs cultured with RANKL further promoted RANKL-induced tartrate-resistant acid phosphatase (TRAP)-positive osteoclast differentiation. RANKL also upregulated AChE expression in BMMs. RNA interference-mediated knockdown of AChE significantly inhibited RANKL-induced osteoclast differentiation and suppressed gene expression specific for osteoclasts. AChE upregulated expression of RANK, the receptor of RANKL, in BMMs. Donepezil decreased cathepsin K expression in BMMs and the resorptive function of osteoclasts on dentine slices. Donepezil decreased RANK expression in BMMs, resulting in the inhibition of osteoclast differentiation with downregulation of c-Fos and upregulation of Id2. Moreover, administration of donepezil prevented RANKL-induced bone loss in vivo, which was associated with the inhibition of bone resorption by osteoclasts. AChE promotes osteoclast differentiation in vitro. Donepezil inhibits osteoclast function in vitro and prevents bone loss by suppressing bone resorption in vivo, suggesting the possibility that donepezil reduces fracture risk in patients with Alzheimer's disease.

Aim: Recent studies have demonstrated that netrin-1 plays a vital role in bone metabolism. Previous studies have shown that osteoblasts produce netrin-1 which affects osteoclast differentiation. However, the role of netrin-1 in osteoblast differentiation is not well understood. In this study, we explored the roles of netrin-1 in osteoblasts. Materials and Methods: Quantitative reverse-transcriptase polymerase chain reaction (qPCR), RNA interference for netrin receptors, the generation of netrin-1 plasmid, transfection of plasmids, and cell proliferation assay were performed. Results: During osteoblast differentiation by ascorbic acid, netrin-1 expression was significantly decreased. Gene expression related with osteoblast differentiation was down-regulated by netrin-1 treatment. We also found that osteoblast differentiation by bone morphogenetic protein-4 (BMP-4) was inhibited in the presence of recombinant netrin-1. Forced expression of both BMP-4 and netrin-1 significantly decreased alkaline phosphatase expression. On the other hand, Unc5b, neogenin, and A2b which belong to netrin receptors were expressed by osteoblasts. Moreover, alkaline phosphatase expression was significantly decreased by knockdown for the combination of two receptors among these receptors. Conclusion: Netrin-1 is involved in the regulation of osteoblast differentiation.

Limited mouth opening is a common health problem that interferes with eating, makes examination of the oral cavity difficult, and may increase the mortality rate during emergency intubation. Here we introduce a disease designated as masticatory muscle tendon-aponeurosis hyperplasia, which is a new clinical condition of limited mouth opening. Most oral surgeons and dentists are still unaware of this disease condition, thus increasing the risk of incorrect diagnosis as some other disease, such as temporomandibular joint disorder. We will review the clinical features, epidemiology, pathophysiology, etiology, diagnosis, treatment, and prognosis of this disease and also appraise the literature available on the subject.