牙周炎中M1型巨噬细胞与破骨细胞形成相关性研究

Correlation study of M1 macrophages and osteoclastogenesis in periodontitis

目的:探究牙周炎中M1型巨噬细胞与破骨细胞形成的相关性。方法:取2月龄C57BL/6J小鼠,丝线结扎建立右侧上颌牙周炎模型,左侧上颌作为自身对照。术后7 d收样,Micro-CT扫描分析牙槽骨吸收情况,苏木素-伊红(HE)、抗酒石酸酸性磷酸酶(TRAP)、CD80和诱导型一氧化氮合酶(iNOS)免疫荧光染色观察破骨细胞及M1型巨噬细胞数量及位置情况,实时定量RT-PCR检测白细胞介素(Il)-1β、Il-6、肿瘤坏死因子α(Tnf-α)、iNos等M1型巨噬细胞相关基因及破骨细胞相关免疫球蛋白样受体(Oscar)、基质金属蛋白酶9(Mmp9)、组织蛋白酶K(Ctsk)、Trap等破骨细胞相关基因表达,对M1型巨噬细胞与破骨细胞的相关基因表达、M1型巨噬细胞与破骨细胞的数量进行相关性分析,对M1型巨噬细胞与破骨细胞定位分布进行分析。结果:成功构建小鼠牙周炎模型,上颌牙周炎侧牙槽骨骨组织参数明显下降(P<0.05);牙周炎侧M1型巨噬细胞及破骨细胞数量明显增加(P<0.05);牙周炎侧Il-1β、Il-6、Tnf-α、iNos等M1型巨噬细胞相关基因及Oscar、Mmp9、Ctsk、Trap等破骨细胞相关基因表达较对照侧高(P<0.05);牙周炎中破骨细胞相关基因与M1型巨噬细胞相关基因表达水平呈显著正相关(P<0.05),M1型巨噬细胞数量与破骨细胞数量呈显著正相关(P<0.05);TRAP染色和iNOS免疫荧光共染结果显示,破骨细胞位于的牙槽骨表面周围有较多M1型巨噬细胞分布。结论:牙周炎时,M1型巨噬细胞和破骨细胞数量均显著增加,且呈显著正相关,牙槽骨周围的M1型巨噬细胞和牙槽骨表面的破骨细胞具有相邻的定位关系。

Objective:To investigate the correlation between M1 macrophages and osteoclastogenesis in periodontitis.Methods:Periodontitis model was established of the right maxillary on 2-month-old C57BL/6J mice,with left maxilla as control.Mice were sacrificed 7 days after surgery.Micro-CT was used to detect alveolar bone loss and histological staining was conducted to examine M1 macrophages and osteoclasts.The expression of genes related to M1 macrophages and osteoclastogenesis was detected by real-time RT-PCR.The correlation between numbers of M1 macrophages and osteoclasts,and location relation of them were analyzed.Results:Mouse periodontitis model was established successfully.Micro-CT results showed significant decrease of bone volume,BV/TV,Tb.N,Tb.Th and BMD of alveolar bone in right maxilla with periodontitis(P<0.05).And histological staining showed that number of M1 macrophages and osteoclasts significantly increased in maxilla with periodontitis(P<0.05).Expressions of M1 macrophages related genes(Il-1β,Il-6,Tnf-α,iNos),and osteoclastogenesis genes(Oscar,Mmp9,Ctsk,Trap)significantly elevated in periodontitis(P<0.05).Correlation analysis results indicated positive correlation between M1 macrophages and osteoclastogenesis in periodontitis.Co-staining results showed that osteoclast mainly located on alveolar bone surface with M1 macrophages around.Conclusions:In periodontitis,M1 macrophages and osteoclasts were increased and there was positive correlation between them,and there were many M1 macrophages around the alveolar bone surface where osteoclasts located.