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目的探讨氯离子通道5(chloride channel 5,CLC5)在模拟失重下磷代谢异常致骨丧失的作用机制,为失重性骨代谢异常的防治提供数据参考。方法①利用回转器对IDG-SW3骨细胞模拟失重,将骨细胞分为模拟失重组(MG组)和失重对照组(MG-CON组),2 d后采用Real-time PCR检测牙本质基质蛋白1(dentinmatrix protein 1,DMP1)和CLC5在IDG-SW3骨细胞中的表达情况;②利用尾部悬吊法对小鼠模拟失重,将10只1月龄C57BL/6雌性小鼠随机分为悬尾组(SUS组)和空白对照组(CON组),每组5只,4周后取两组小鼠胫骨制作石蜡切片进行CLC5免疫组织化学染色;③2月龄DMP1基因敲除鼠和DMP1转基因鼠各5只(体重为20~25g),制备胫骨石蜡切片,进行CLC5免疫组织化学染色。结果 Realtime PCR检测结果显示2D回转后DMP1和CLC5在IDG-SW3骨细胞中的表达量均升高,差异均有统计学意义(均P<0.05);免疫组织化学染色显示CLC5在CON组和SUS组胫骨细胞胞质和基质中均有表达,且SUS组表达量明显高于CON组,差异有统计学意义(P<0.05);CLC5在DMP1基因敲除鼠和DMP1转基因鼠骨细胞中的表达也较CON组升高,差异均有统计学意义(均P<0.05)。结论模拟失重下CLC5参与了DMP1对磷代谢的调控,但其具体作用机制有待于进一步研究。
Objective To investigate the mechanism of chloride channel 5 (CLC5) on bone loss due to abnormal phosphorus metabolism under simulated weightlessness, and to provide a reference for the prevention and treatment of abnormal bone metabolism. METHODS: ①Simulated weightlessness of IDG-SW3 osteoblasts was established by using a gyrator. The osteoblasts were divided into simulated weightlessness group (MG group) and weightless control group (MG-CON group). Real-time PCR was used to detect dentin matrix protein (DMP1) and CLC5 in IDG-SW3 cells. ② The tail suspension method was used to simulate the weightlessness in mice, and 10 female C57BL / 6 mice aged 1 month were randomly divided into four groups (SUS group) and blank control group (CON group), 5 rats in each group. After 4 weeks, two groups of mice tibias were taken to make paraffin sections for CLC5 immunohistochemical staining; ③ 2-month-old DMP1 knockout mice and DMP1 transgenic mice Each of 5 (body weight of 20 ~ 25g), preparation of tibial paraffin sections, CLC5 immunohistochemical staining. Results The results of Realtime PCR showed that the expression of DMP1 and CLC5 in IDG-SW3 cells increased after 2D rotation, the difference was statistically significant (all P <0.05). Immunohistochemical staining showed that the expression of CLC5 in CON group and SUS The expression of CLC5 in the bone marrow of DMP1 knockout mice and DMP1 transgenic mice was significantly higher than that of CON group (P <0.05) Also higher than CON group, the difference was statistically significant (P <0.05). Conclusion CLC5 participates in the regulation of phosphorus metabolism under simulated weightlessness, but its specific mechanism remains to be further studied.