Osteoclast formation and bone resorption are inhibited by megakaryocytes
Osteoclast formation and bone resorption are inhibited by megakaryocytes
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DOI：
10.1016/j.bone.2006.06.004
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发表时间：
2006-11-01
期刊：
影响因子：
4.1
通讯作者：
Compston, J. E.
中科院分区：
文献类型：
--
作者：
Beeton, C. A.;Bord, S.;Compston, J. E.
以前有报道称，在体外向成骨细胞中加入巨核细胞（MK）导致成骨细胞胶原和骨保护素（OPG）的产生增加，表明MK在骨形成中的作用。为了进一步研究这种作用，我们研究了MK对破骨细胞形成和活性的影响。人破骨细胞由从外周血分离的CD 14单核细胞产生，并在存在M-CSF和sRANKL的情况下在牙本质和磷酸钙基质上培养。从分离自人外周血或脐带血的CD 34+细胞产生MK，并在液体培养基中培养6天，之后分离成熟的MK（CD 61阳性细胞）并加入单核细胞培养物中。培养6天和9天后，对形态学和TRAP染色鉴定的破骨细胞数量进行计数。在第0天向破骨细胞培养物中加入MK抑制破骨细胞形成的数量1.9倍（p > 0.003），而在第3天加入MK对破骨细胞数量没有影响。当与破骨细胞共培养时，MK的存在从第0天起抑制吸收8.7倍（p > 0.004），但当与破骨细胞共培养时从第3天起仅抑制吸收3.1倍（p > 0.01）。在剂量反应实验中，发现1-10%的MK加入单核细胞培养物引起最大的再吸收抑制。用CD 61阴性细胞（非MK）处理类似的破骨细胞培养物，以证实破骨细胞形成和活性的抑制是由MK特异性引起的。用细胞不可渗透膜进行的实验表明，细胞与细胞的接触和可溶性因子的释放都参与介导这些效应。这些结果表明MK抑制破骨细胞的形成和活性。当MK和破骨细胞从第0天起共培养时，观察到最显著的效果。提示MK主要作用于破骨细胞前体。(C)2006年爱思唯尔公司All rights reserved.
It has been previously reported that addition of megakaryocytes (MKs) to ostcoblasts in vitro results in increased osteoblastic collagen and osteoprotegerin (OPG) production, suggesting a role for MKs in bone formation. To further investigate this role, we have studied the effects of MKs on osteoclast formation and activity. Human osteoclasts were generated from CD14 monocytes isolated from peripheral blood and cultured in the presence of M-CSF and sRANKL on dentine and calcium phosphate substrates. MKs were generated from CD34+ cells isolated from either human peripheral blood or cord blood and cultured in liquid medium for 6 days, after which time maturing MKs (CD61 positive cells) were isolated and added to monocyte cultures. After 6 and 9 days of culture, the number of osteoclasts identified morphologically and by TRAP staining was counted. Cells were removed and the area of resorption was identified by von Kossa staining and quantitatively assessed by image analysis.The addition of MKs to osteoclast cultures at day 0 inhibited the number of osteoclasts formed 1.9-fold (p > 0.003), whereas addition at 3 days had no effect on osteoclast number. The presence of MKs inhibited resorption 8.7-fold when co-cultured with osteoclasts from day 0 (p > 0.004), but only by 3.1-fold when co-cultured from day 3 (p > 0.01). In dose-response experiments, it was found that 1-10% of MKs added to monocyte cultures elicited the greatest inhibition of resorption. Similar osteoclast cultures were treated with CD61 negative cells (non-MKs) to confirm that the inhibition of osteoclast formation and activity was specifically due to MKs. Experiments with a cell-impermeable membrane indicated that both cell to cell contact and release of soluble factor(s) were involved in mediating these effects. These results show that MKs inhibit osteoclast formation and activity. The most pronounced effects were seen when MKs and osteoclasts were co-cultured from day 0. suggesting that MKs act primarily on osteoclast precursors.(C) 2006 Elsevier Inc. All rights reserved.