低氧对炎性环境下髓核细胞凋亡的作用及机制  

Effect and mechanism of hypoxia on nucleus pulposus cell apoptosis in infammatory environment

作　　者：李磊[1] 王一范 施强慧 钟华建 曹佳实 Li Lei;Wang Yifan;Shi Qianghui;Zhong Huajian;Cao Jiashi(Department of Orthopaedics,Quzhou Hospital(Quzhou People's Hospital),Wenzhou Medical University,Quzhou 324000,Zhejiang,China;Department of Health Management,Changzheng Hospital,Naval Medical University,Shanghai 200003,China;Department of Orthopaedics,63680th Hospital of Chinese PLA,Jiangyin 214400,Jiangsu,China;Department of Orthopaedics,Changzheng Hospital,Naval Medical University,Shanghai 200003,China;Department of Naval Medical Center,Naval Medical University,Shanghai 200433,China)

机构地区：[1]温州医科大学附属衢州医院(衢州市人民医院)骨科,衢州324000 [2]海军军医大学长征医院健康管理科,上海200003 [3]中国人民解放军63680部队医院骨科,江阴214400 [4]海军军医大学长征医院骨科,上海200003 [5]海军军医大学海军特色医学中心,上海200433

出　　处：《脊柱外科杂志》2024年第1期37-45,共9页Journal of Spinal Surgery

摘　　要：目的探讨低氧对炎性环境下髓核细胞凋亡的影响及作用机制。方法体外培养大鼠原代髓核细胞,采用低氧小室进行低氧干预,采用促炎因子肿瘤坏死因子-α(TNF-α)刺激髓核细胞模拟炎性环境。通过流式细胞术及TUNEL染色检测髓核细胞凋亡情况,通过蛋白质印迹法检测凋亡相关蛋白表达水平,明确低氧对髓核细胞凋亡的影响;随后通过荧光定量PCR及酶联免疫吸附试验评估炎性环境下髓核细胞内炎性反应情况,并通过蛋白质印迹法分析髓核细胞内NLRP3炎性小体激活水平;最后采用NLRP3炎性小体激动剂QS-21,通过回复实验明确NLRP3炎性小体在低氧调控髓核细胞凋亡中的作用。结果低氧对正常环境下髓核细胞凋亡无明显影响,但能抑制炎性环境下髓核细胞凋亡。在炎性环境下,髓核细胞炎性细胞因子白细胞介素(IL)-1β、IL-8、IL-18的表达及分泌增加,细胞内NLRP3炎性小体激活,而低氧干预能缓解上述改变。回复实验表明,NLRP3炎性小体激动剂QS-21可削弱低氧对炎性环境下髓核细胞凋亡的抑制作用。结论低氧通过抑制NLRP3炎性小体激活缓解炎性环境下髓核细胞凋亡。Objective To investigate the effect and mechanism of hypoxia on nucleus pulposus cell apoptosis in inflammatory environments.Methods Primary rat nucleus pulposus cells were cultured in vitro.Hypoxic intervention was performed using a hypoxic chamber,and the pro-inflammatory factor tumor necrosis factor-α(TNF-α)was used to simulate the inflammatory environment.In order to determine the effect of hypoxia on nucleus pulposus cell apoptosis,flow cytometry and TUNEL staining were performed to detect the nucleus pulposus cell apoptosis,and the expression of apoptosis-related proteins was measured by Western blotting.Subsequently,the inflammatory response in nucleus pulposus cells in inflammatory environment was evaluated by real-time PCR and ELISA,and the NLRP3 inflammasome activation in nucleus pulposus cells was estimated through Western blotting.Finally,the NLRP3 inflammasome agonist QS-21 was used to clarify the role of NLRP3 inflammasome in hypoxia regulating nucleus pulposus cell apoptosis in inflammatory environments.ResultsHypoxia significantly inhibited nucleus pulposus cell apoptosis in inflammatory environments while showing no effect on nucleus pulposus cells in normal conditions.When cultured in inflammatory conditions,the expression and secretion of inflammatory cytokines interleukin(IL)-1β,IL-8,and IL-18 by nucleus pulposus cells were increased,accompanied with NLRP3 inflammasome activation,while hypoxia intervention significantly alleviated those changes in nucleus pulposus cells.Rescue assay showed that NLRP3 inflammasome agonist QS-21 significantly weakened the inhibitory effect of hypoxia on nucleus pulposus cell apoptosis in inflammatory environment.Conclusion Hypoxia alleviates nucleus pulposus cell apoptosis in inflammatory environment by inhibiting NLRP3 inflammasome activation.

关 键 词：细胞低氧 炎症 椎间盘退行性变 细胞凋亡 大鼠 

分 类 号：R342.2[医药卫生—基础医学]