S. Ueda, T. Ichiseki, Miyako Shimasaki, Hiroaki Hirata, N. Kawahara, Y. Ueda
{"title":"缺氧环境中糖皮质激素诱导的骨细胞死亡与坏死性坏死有关","authors":"S. Ueda, T. Ichiseki, Miyako Shimasaki, Hiroaki Hirata, N. Kawahara, Y. Ueda","doi":"10.3390/biochem1020009","DOIUrl":null,"url":null,"abstract":"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.","PeriodicalId":72357,"journal":{"name":"BioChem","volume":"49 1","pages":""},"PeriodicalIF":0.0000,"publicationDate":"2021-08-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Glucocorticoid-Induced Osteocytic Cell Death in a Hypoxic Environment Is Associated with Necroptosis\",\"authors\":\"S. Ueda, T. Ichiseki, Miyako Shimasaki, Hiroaki Hirata, N. Kawahara, Y. Ueda\",\"doi\":\"10.3390/biochem1020009\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"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.\",\"PeriodicalId\":72357,\"journal\":{\"name\":\"BioChem\",\"volume\":\"49 1\",\"pages\":\"\"},\"PeriodicalIF\":0.0000,\"publicationDate\":\"2021-08-30\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"BioChem\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.3390/biochem1020009\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"BioChem","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.3390/biochem1020009","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
Glucocorticoid-Induced Osteocytic Cell Death in a Hypoxic Environment Is Associated with Necroptosis
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.