p130Cas, Crk-associated substrate (Cas), is an adaptor/scaffold protein that plays a central role in actin cytoskeletal reorganization. We previously reported that p130Cas is not tyrosine-phosphorylated in osteoclasts derived from Src-deficient mice, which are congenitally osteopetrotic, suggesting that p130Cas serves as a downstream molecule of c-Src and is involved in osteoclastic bone resorption. However, the physiological role of p130Cas in osteoclasts has not yet been confirmed because the p130Cas-deficient mice displayed embryonic lethality. Osteoclast-specific p130Cas conditional knockout (p130Cas DOCL-) mice exhibit a high bone mass phenotype caused by defect in multinucleation and cytoskeleton organization causing bone resorption deficiency. Bone marrow cells from p130Cas DOCL-mice were able to differentiate into osteoclasts and wild-type cells in vitro. However, osteoclasts from p130Cas DOCL-mice failed to form actin rings and resorb pits on dentine slices. Although the initial events of osteoclast attachment, such as b3-integrin or Src phosphorylation, were intact, the Rac1 activity that organizes the actin cytoskeleton was reduced, and its distribution was disrupted in p130Cas DOCL-osteoclasts. Dedicator of cytokinesis 5 (Dock5), a Rho family guanine nucleotide exchanger, failed to associate with Src or Pyk2 in osteoclasts in the absence of p130Cas. These results strongly indicate that p130Cas plays pivotal roles in osteoclastic bone resorption.
This study aimed to evaluate the effect of phytic acid (IP6), used as etchant, on resin-dentin bond strength, smear layer removal, and the viability of pulpal cells. Flat dentin surfaces with smear layer were etched with 1% IP6 for 60, 30, or 15 s; in the control group 37% phosphoric acid (PA) was used. Dentin surfaces were rinsed, blot-dried, and bonded with an etch-and-rinse adhesive, followed by composite build-ups. The specimens were subjected to tensile testing after 24 h of water storage at 37°C, and failure modes were determined using scanning electron microscopy. The effectiveness of IP6 to remove the smear layer was observed using scanning electron microscopy. To evaluate the effect on pulpal cells, solutions of 0.1 and 0.01% IP6 and of 3.7 and 0.37% PA were prepared and rat pulpal cells were treated with these solutions for 6 and 24 h. Cell viability was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The results demonstrated that all application times of IP6 produced bond-strength values that were significantly higher than that of the control. Phytic acid effectively removed the smear layer and plugs, thus exposing the collagen network. Phytic acid had a minimal effect on pulpal cells, whereas PA resulted in a marked decrease in their viability.
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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