Development of the cell culture system to examine the mechanism of the action on bone-resorbing factors in vitro.

使用各种体内和体外系统的研究表明，破骨细胞前体存在于骨髓细胞中，并表明破骨细胞是通过它们的融合形成的。近年来，骨髓细胞长期培养的实验表明，未成熟的单核细胞在1 α，25-二羟维生素D_3[1 α，25-（OH）_2D_3]的诱导下发生融合，形成具有破骨细胞特征的多核细胞。因此，在本研究中，我们开发了细胞培养系统，以检查骨吸收因子在体外的作用机制。为此，我们分别研究了1 α，25-（OH）_2D_3和4种1 α，25-（OH）_2D_3- 26，23-内酯在小鼠骨髓单个核细胞培养中介导多核细胞形成的能力。23（S），25<-9><-7>（S）-和23（R），25（R）-1alpha，25-（OH）_2D_3- 26，23-内酯也能增加多核细胞的数目，而23（S），25（R）-和23（R），25（S）-1alpha，25-（OH）_2D_3- 26，23-内酯则不能。此外，后两种非对映异构体抑制1 α，25-（OH）_2D_3对多核细胞形成的刺激作用，而23（S），25（S）-和23（R），25（R）-1 α，25-（OH）_2D_3- 26，23-内酯则无此作用。这些多核细胞对降钙素有反应，并含有抗酒石酸酸性磷酸酶，这两者都是破骨细胞的特征。这些结果表明，在小鼠骨髓培养中形成的多核细胞满足大多数破骨细胞的标准。

Studies using a variety of in vivo and in vitro systems have indicated that osteoclast precursors are present among bone marrow cells and have suggested that osteoclasts are formed by their fusion. More recent reports arising from experiments employing long term culture of bone marrow cells state that immature monocytes may undergo fusion induced by lalpha,25-dihydroxyvitamin D_3[lalpha,25-(OH)_2D_3], resulting in multinucleated cells with several characteristics of osteoclasts. Therefore,in this study, we have developed the cell culture system to examine the mechanism of the action on bone-resorbing factor in vitro. For this purpose, we have examined separately the ability of lalpha,25-(OH)_2D_3 and the four lalpha,25-(OH)_2D_3-26,23-lactones to mediate the formation of multinucleated cells in mouse bone marrow mononuclear cell cultures. The addition of lalpha,25-(OH)_2D_3 to these cultures dose-dependently stimulated the formation of multinuleated cells over a range of lo^<-9>- lo^<-7>M.The 23(S),25(S)- and 23(R),25(R)-lalpha,25-(OH)_2D_3-26,23-lactones also increased the number of multinucleated cells, whereas the 23(S),25(R)- and 23(R),25(S)-lalpha,25-(OH)_2D_3-26,23-lactones failed to do so. In addition, these latter two diastereoisomers inhibited the lalpha,25-(OH)_2D_3 stimulation of multinucleated cell formation, although the 23(S),25(S)- and 23(R),25(R)-lalpha,25-(OH)_2D_3-26,23-lactones did not. These multinuleated cells responded to calcitinin and contained tartrate-resistant acid phosphatase, both of which are characteristic of osteoclasts. These results suggest that the multinucleated cells formed in the mouse marrow culture satisfy most of the criteria of osteoclasts.