Glucocorticoid-Induced Osteocytic Cell Death in a Hypoxic Environment Is Associated with Necroptosis
Glucocorticoid-Induced Osteocytic Cell Death in a Hypoxic Environment Is Associated with Necroptosis
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DOI：
10.3390/biochem1020009
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发表时间：
2021-08
期刊：
影响因子：
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通讯作者：
S. Ueda;T. Ichiseki;Miyako Shimasaki;Hiroaki Hirata;N. Kawahara;Y. Ueda
中科院分区：
文献类型：
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作者：
S. Ueda;T. Ichiseki;Miyako Shimasaki;Hiroaki Hirata;N. Kawahara;Y. Ueda
糖皮质激素相关性股骨头坏死的基础病理生理学或预防策略尚未建立。在神经血管和心脏缺血性疾病中，坏死性下垂已被报道为细胞死亡的新概念。在这里，我们研究了坏死性下垂在体外糖皮质激素诱导的骨坏死中的作用，其可能的原因是缺血。加入1µM地塞米松(Dex)的小鼠成骨细胞(MLO-Y4)在1%O2(低氧)中培养，被认为与糖皮质激素诱导的骨坏死发生的体内环境(H-D应激环境)相似。用培养24 h的细胞(Dex(+)/低氧(+)组)，用受体相互作用蛋白(RIP)1和RIP3进行免疫荧光染色和Western blotting。此外，在Dex(+)/缺氧(+)细胞中加入NEC-1(NEC-1)，培养12 h和24 h后，用凋亡/坏死细胞检测试剂盒计数并比较细胞的凋亡数和坏死率。在Dex(+)/低氧(+)组，RIP1和RIP3均有表达。此外，在Western blotting中，NEC-1的加入减弱了它们的表达。服用NEC-1后，细胞死亡数量也有所减少。坏死性下垂被认为是导致骨细胞坏死的原因之一。使用坏死下垂抑制剂NEC-1，提示了一种可能的方法，即使在H-D应激环境中，在12小时内给药也可以防止骨细胞坏死。
Neither the underlying pathophysiology of nor prophylactic strategies for glucocorticoid-associated femoral head osteonecrosis have yet been established. In neurovascular and cardiac ischemic disorders, necroptosis has been reported as a new concept of cell death. Here we investigated the involvement of necroptosis in glucocorticoid-induced osteonecrosis in vitro, the putative cause of which is ischemia. Murine osteocytic cells (MLO-Y4) to which 1 µM dexamethasone (Dex) was added and were cultured in 1% O2 (hypoxia) are thought to resemble the in vivo environment in which glucocorticoid-induced osteonecrosis occurs (H-D stress environment). Using such cells cultured for 24 h (Dex(+)/hypoxia(+) group), immunofluorescent staining and Western blotting were performed with receptor-interacting protein (RIP) 1 and RIP3, which are necroptosis expression factors. In addition, the necroptosis inhibitor necrostatin-1 (Nec-1) was added to Dex(+)/hypoxia(+) and cultured for 12 h and 24 h. Then using an Apoptotic/Necrotic Cells Detection Kit the numbers of apoptotic and necrotic cells were counted and compared. In Dex(+)/hypoxia(+) group, expression of both RIP1 and RIP3 was found. Additionally, in Western blotting, the addition of Nec-1 attenuated their expression. A decrease in the number of cell deaths was also found following Nec-1 administration. Necroptosis has been implicated as a cause of death in osteocytic cell necrosis. Use of the necroptosis inhibitor, Nec-1, suggests a possible approach to preventing osteocytic cell necrosis even in an H-D stress environment when given within 12 h.