Analysis of signaling pathways involved in polarization of osteoclasts.

(1)我们之前已经建立了一个检测破骨细胞形成的培养系统。在本研究中，我们还利用体外形成的破骨细胞样细胞建立了一种新的吸收测定系统。因此，现在有可能在一系列实验中检查不同化合物对破骨细胞募集和由相同的破骨细胞形成窝的影响。(2)利用op/op小鼠来源的成骨细胞，我们发现M-CSF不仅对破骨细胞前体的增殖，而且对它们向破骨细胞的分化都是必不可少的。(3)破骨细胞置于牙本质切片或塑料培养皿上，形成足小体（肌动蛋白环）的环状结构。环状结构对应于骨吸收破骨细胞的清晰区。肌动蛋白环的形成和破坏与破骨细胞的窝形成活性密切相关。(4) p60^<c-src>、PI-3激酶、局灶黏附激酶和小GTP结合蛋白rho介导的信号通路主要参与破骨细胞肌动蛋白环的形成。已经提出降钙素受体与蛋白激酶A和蛋白激酶c相关的途径偶联。我们研究了破骨细胞样细胞中降钙素作用的信号通路。我们发现降钙素引起的细胞骨架的剧烈改变主要是通过蛋白激酶A途径介导的。

(1) We have previously established a culture system for examining osteoclast formation. In the present study, a new resorption assay system was also established using osteoclast-like cells formed in vitro. Therefore, it is now possible to examine the effect of various compounds on the recruitment of osteoclasts and pit formation by the same osteoclasts in a series of experiments.(2) Using op/op mouse-derived osteoblastic cells, we found that M-CSF is indispensable not only for proliferation of osteoclast precursors but also for their differentiation into osteoclasts.(3) When osteoclasts were placed on dentine slices or plastic dishes, they formed the ringed structure of podosomes (actin rings). The ringed structure is shown to correspond to clear zones of bone resorbing osteoclasts. The Formation and disruption of the actin ring were correlated well with pit forming activity of osteoclasts.(4) It was shown that signaling pathways meditated by p60^<c-src>, PI-3 kinase, focal adhesion kinase and small GTP binding protein, rho, are essentially involved in actin ring formation by osteoclasts. It has been proposed that calcitonin receptors are coupled to the pathways associated with both protein kinase A and protein kinase C.We examined the signaling pathway of calcitonin action in osteoclast-like cells. We found that calcitonin-induced dramatic alteration in the cytoskeleton is mainly mediated by the protein kinase A pathway.