磨损颗粒刺激下小鼠骨髓来源巨噬细胞调控白细胞介素-34表达的实验研究  

Experimental evaluation of the regulation of IL-34 expression in mouse BMMs stimulated by wear particles

作　　者：刘子歌 今出真司 张晨[1] 宋国瑞 陈德胜[1] LIU Zige;IMADE Shinji;ZHANG Chen;SONG Guorui;CHEN Desheng(Department of Orthopedics,General Hospital of Ningxia Medical University,Yinchuan 750004,China;Department of Orthopedics,Shimane University Hospital,Shimane Izumo 693-8501,Japan)

机构地区：[1]宁夏医科大学总医院骨科,银川750004 [2]日本岛根大学附属医院矫形外科,日本岛根出云693-8501

出　　处：《中国医科大学学报》2021年第6期496-501,506,共7页Journal of China Medical University

基　　金：国家自然科学基金(81560364,81760405,81760395,82060408);宁夏自然科学基金重点项目(2018AAC02013);宁夏医科大学校级课题重点项目(XZ2018014)。

摘　　要：目的探讨白细胞介素-34(IL-34)在体外是否可以促进破骨细胞形成以及在磨损颗粒刺激下小鼠骨髓来源巨噬细胞(BMMs)中IL-34的体外表达量和相关机制。方法提取小鼠BMMs原代细胞,采用抗酒石酸酸性磷酸酶(TRAP)染色检测IL-34或巨噬细胞集落刺激因子(M-CSF)在核因子κB受体激活剂(RANKL)存在下处理BMMs后破骨细胞的形成。酶联免疫吸附(ELISA)及实时聚合酶链反应(RT-PCR)法测定不同浓度下钴铬钼合金颗粒(CoCrMo)刺激下小鼠BMMs中IL-34的表达量。RT-PCR和Western blotting法检测丝裂原活化蛋白激酶(MAPK)信号转导通路抑制剂干预下IL-34及其磷酸化蛋白的表达量。结果IL-34和M-CSF对于促进体外破骨细胞的生成具有相同的作用。ELISA和RT-PCR结果显示,CoCrMo磨损颗粒以时间和浓度依赖趋势增加BMMs中IL-34的表达水平。Western blotting结果进一步解释BMMs中IL-34的表达量的升高可能与细胞外调节蛋白激酶(ERK)和c-Jun氨基末端激酶(JNK)磷酸化水平的增加有关。结论磨损颗粒通过影响ERK和JNK磷酸化水平调控IL-34的表达,参与体外破骨细胞的形成。Objective To investigate whether interleukin 34(Il-34)can promote osteoclast formation in vitro,and to assess in vitro expression of IL-34 in bone marrow macrophages(BMMs)stimulated by wear particles and related mechanisms.Methods Mouse BMMs were extracted from primary cells and stained with anti-tartrate acid phosphatase,to identify cells producing IL-34 or macrophage colony stimulating factor(M-CSF).Incubation of bone marrow with nuclear factor kappa B receptor activator promotes formation of osteoclasts from BMMs.Enzyme-linked immunosorbent assay(ELISA)and real-time polymerase chain reaction(RT-PCR)were used to quantitate IL-34 expression in small BMMs stimulated with different concentrations of cobalt-chromium-molybdenum alloy particles(CoCrMo).RTPCR and Western blotting were used to detect IL-34 expression and phosphorylation under the intervention of a MAPK signaling pathway inhibitor.Results IL-34 and M-CSF could promote osteoclast formation in vitro.ELISA and RT-PCR results showed that CoCrMo wear particles can increase IL-34 expression in BMMs in a time-and concentration-dependent manner.Western blotting indicated that these particles might increase ERK and JNK phosphorylation.Conclusion Wear particles may modulate the pathological process of in vitro osteoclast formation involving IL-34,by up-regulating ERK and JNK phosphorylation.

关 键 词：白细胞介素-34 破骨细胞 骨质疏松症 丝裂原活化蛋白激酶信号转导通路 

分 类 号：R392[医药卫生—免疫学]