Stimulation of bone formation in cortical bone of mice treated with a receptor activator of nuclear factor-κB ligand (RANKL)-binding peptide that possesses osteoclastogenesis inhibitory activity.
Stimulation of bone formation in cortical bone of mice treated with a receptor activator of nuclear factor-κB ligand (RANKL)-binding peptide that possesses osteoclastogenesis inhibitory activity.
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用核因子 - κB配体（RANKL）结合肽的受体激活剂治疗的小鼠的皮质骨刺激骨形成，该肽具有破骨构成抑制活性。
DOI：
10.1074/jbc.m112.426080
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发表时间：
2013-02-22
期刊：
影响因子：
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通讯作者：
Yasuda H
中科院分区：
文献类型：
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作者：
Furuya Y;Inagaki A;Khan M;Mori K;Penninger JM;Nakamura M;Udagawa N;Aoki K;Ohya K;Uchida K;Yasuda H
背景：已知RANKL结合肽WP 9 QY（W 9）抑制破骨细胞生成。结果：W 9对小鼠皮质骨有合成代谢作用。W 9结合RANKL和分化的成骨细胞，产生自分泌因子如BMP-4。结论：RANKL信号通路参与了W 9诱导的成骨细胞分化。意义：RANKL通路可能是成骨细胞分化的一种新机制。到目前为止，甲状旁腺激素是临床上唯一可用的骨合成代谢药物。开发此类药物的主要困难是缺乏对成骨细胞分化和骨形成调节机制的阐明。在这里，我们报告了一种已知通过阻断核因子-κB配体（RANKL）受体激活剂-RANK信号传导在体内消除破骨细胞分化的肽（W 9），我们惊讶地发现该肽在体内表现出骨合成代谢作用。W 9皮下给药3次/天，持续5天，可显著增加小鼠皮质骨的骨矿物质密度。组织形态计量学分析显示股骨远端干骺端破骨细胞生成减少，股骨干骨形成显著增加。我们的研究结果表明，W 9发挥骨合成代谢活性。为了阐明这种活性所涉及的机制，我们研究了W 9对MC 3 T3-E1（E1）细胞中成骨细胞分化/矿化的影响。茜素红染色显示，W 9显著增加碱性磷酸酶（成骨细胞的标志酶）的活性和矿化。在W 9处理的E1细胞中，几种成骨相关因子的基因表达增加。加入W 9激活E1细胞中的p38 MAPK和Smad 1/5/8，并且W 9在体外显示出与BMP-2协同的成骨刺激活性和异位骨形成。E1细胞中RANKL表达的敲低降低了W 9的作用。此外，在碱性磷酸酶测定中，W 9对RANKL缺陷型成骨细胞的作用较弱。综上所述，我们的研究结果表明，这种肽可能有助于治疗骨疾病，W 9通过RANKL对成骨细胞的作用并伴随着几种自分泌因子的产生来实现其骨合成代谢活性。
Background: A RANKL-binding peptide WP9QY (W9) is known to inhibit osteoclastogenesis. Results: W9 showed an anabolic effect on cortical bone in mice. W9 bound RANKL and differentiated osteoblasts with production of autocrine factors like BMP-4. Conclusion: Signaling through RANKL is involved in part in the W9-induced osteoblast differentiation. Significance: The RANKL pathway could be a novel mechanism in osteoblast differentiation. To date, parathyroid hormone is the only clinically available bone anabolic drug. The major difficulty in the development of such drugs is the lack of clarification of the mechanisms regulating osteoblast differentiation and bone formation. Here, we report a peptide (W9) known to abrogate osteoclast differentiation in vivo via blocking receptor activator of nuclear factor-κB ligand (RANKL)-RANK signaling that we surprisingly found exhibits a bone anabolic effect in vivo. Subcutaneous administration of W9 three times/day for 5 days significantly augmented bone mineral density in mouse cortical bone. Histomorphometric analysis showed a decrease in osteoclastogenesis in the distal femoral metaphysis and a significant increase in bone formation in the femoral diaphysis. Our findings suggest that W9 exerts bone anabolic activity. To clarify the mechanisms involved in this activity, we investigated the effects of W9 on osteoblast differentiation/mineralization in MC3T3-E1 (E1) cells. W9 markedly increased alkaline phosphatase (a marker enzyme of osteoblasts) activity and mineralization as shown by alizarin red staining. Gene expression of several osteogenesis-related factors was increased in W9-treated E1 cells. Addition of W9 activated p38 MAPK and Smad1/5/8 in E1 cells, and W9 showed osteogenesis stimulatory activity synergistically with BMP-2 in vitro and ectopic bone formation. Knockdown of RANKL expression in E1 cells reduced the effect of W9. Furthermore, W9 showed a weak effect on RANKL-deficient osteoblasts in alkaline phosphatase assay. Taken together, our findings suggest that this peptide may be useful for the treatment of bone diseases, and W9 achieves its bone anabolic activity through RANKL on osteoblasts accompanied by production of several autocrine factors.