Localized apoptosis of osteocytes, the tissue-resident cells within bone, occurs with fatigue microdamage and activates bone resorption. Osteoclasts appear to target and remove dying osteocytes, resorbing damaged bone matrix as well. Osteocyte apoptosis similarly activates bone resorption with estrogen loss and in disuse. Apoptotic osteocytes trigger viable neighbor (ie, bystander) osteocytes to produce RANKL, the cytokine required for osteoclast activation. Signals from apoptotic osteocytes that trigger this bystander RANKL expression remain obscure. Studying signaling among osteocytes has been hampered by lack of in vitro systems that model the limited communication among osteocytes in vivo (ie, via gap junctions on cell processes and/or paracrine signals through thin pericellular fluid spaces around osteocytes). Here, we used a novel multiscale fluidic device (the Macro-micro-nano, or Mμn) that reproduces these key anatomical features. Osteocytes in discrete compartments of the device communicate only via these limited pathways, which allows assessment of their roles in triggering osteocytes RANKL expression. Apoptosis of MLOY-4 osteocytes in the Mμn device caused increased osteocyte RANKL expression in the neighboring compartment, consistent with in vivo findings. This RANKL upregulation in bystander osteocytes was prevented by blocking Pannexin 1 channels as well as its ATP receptor. ATP alone caused comparable RANKL upregulation in bystander osteocytes. Finally, blocking Connexin 43 gap junctions did not abolish osteocyte RANKL upregulation, but did alter the distribution of RANKL expressing bystander osteocytes. These findings point to extracellular ATP, released from apoptotic osteocytes via Panx1 channels, as a major signal for triggering bystander osteocyte RANKL expression and activating bone remodeling. © 2020 American Society for Bone and Mineral Research.

中文翻译：

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凋亡骨细胞通过嘌呤能信号传导和 Pannexin 通道激活诱导旁观者产生 RANKL。

骨细胞（骨内的组织驻留细胞）的局部凋亡伴随疲劳微损伤发生并激活骨吸收。破骨细胞似乎靶向并去除垂死的骨细胞，同时吸收受损的骨基质。骨细胞凋亡类似地激活骨吸收，同时雌激素丢失和废弃。凋亡的骨细胞触发有活力的邻近（即旁观者）骨细胞产生 RANKL，这是破骨细胞活化所需的细胞因子。来自凋亡骨细胞的触发这种旁观者 RANKL 表达的信号仍然模糊不清。由于缺乏模拟体内骨细胞之间有限通信的体外系统（即，通过细胞过程上的间隙连接和/或通过骨细胞周围薄细胞外周液空间的旁分泌信号），研究骨细胞之间的信号传导受到了阻碍。这里，我们使用了一种新颖的多尺度流体装置（Macro-micro-nano，或 Mμn）来重现这些关键的解剖特征。设备的离散隔室中的骨细胞仅通过这些有限的途径进行通信，从而可以评估它们在触发骨细胞 RANKL 表达中的作用。Mμn 装置中 MLOY-4 骨细胞的凋亡导致相邻隔室中骨细胞 RANKL 表达增加，这与体内研究结果一致。通过阻断 Pannexin 1 通道及其 ATP 受体，可以防止旁观者骨细胞中的这种 RANKL 上调。单独的 ATP 在旁观者骨细胞中引起相当的 RANKL 上调。最后，阻断连接蛋白 43 间隙连接并没有消除骨细胞 RANKL 上调，但确实改变了表达 RANKL 的旁观者骨细胞的分布。这些发现指向细胞外 ATP，通过 Panx1 通道从凋亡的骨细胞释放，作为触发旁观者骨细胞 RANKL 表达和激活骨重塑的主要信号。© 2020 美国骨与矿物研究学会。