口腔颌面部细菌感染体外模型的建立  

Establishment of oral and maxillofacial bacterial infection model in vitro

作　　者：史庆怡 孔晨飞[2] 王晓峰[1] 张天夫[1] SHI Qing-yi;KONG Chen- fei;WANG Xiao- feng;et al(Department of Stomatology ,Chinx Japan Union Hospital ,Jilin University ,Changchun 130033 ,China)

机构地区：[1]吉林大学中日联谊医院口腔科,吉林长春130033 [2]吉林大学中日联谊医院科学研究中心,吉林长春130033

出　　处：《中国实验诊断学》2018年第4期704-708,共5页Chinese Journal of Laboratory Diagnosis

基　　金：2017年吉林省卫生厅项目;吉林省教育厅"十三五"科学技术项目(JJKH20170822KJ);吉林省直卫生专项项目(sczsy201525)

摘　　要：目的利用细菌脂多糖(LPS)刺激小鼠结缔组织成纤维细胞L929及小鼠巨噬细胞RAW264.7,建立体外口腔颌面部细菌感染及免疫应答模型,检测不同时间点炎症因子的表达情况。方法利用LPS刺激L929及RAW264.7细胞,采用CCK-8法测定LPS对细胞增殖的影响。QPCR检测肿瘤坏死因子TNF-α、细胞炎症因子、趋化因子等相关基因的表达变化情况。ELISA检测细胞上清液中IL-6水平变化。结果 CCK-8结果显示,LPS能够引起细胞增殖变化。QPCR实验结果显示TNF-α,IL-1,IL-5,IL-6,IL-8,IL-33的mRNA表达明显升高,而IL-10,IL-13,IL-23表达明显下降。CXCL2,CXCL10,FOXO3,GDF15表达量显著增加。ELISA结果显示50μg/mL LPS刺激24h、48h,L929细胞分泌IL-6含量分别为42.04pg/ml、45.52pg/mL;RAW264.7细胞分泌IL-6含量为25.90pg/ml,28.92pg/ml。100μg/ml LPS刺激24h、48h,L929细胞分泌IL-6的含量为18.97pg/ml、21.29pg/ml;RAW264.7细胞分泌IL-6含量为16.90pg/ml,18.21pg/ml。结论 LPS刺激L929及RAW264.7细胞能够引起细胞发生炎症反应,相关炎症因子及趋化因子的表达增加,证明成功建立了口腔颌面部感染体外模型。Objective Using bacterial lipopolysaccharide（lipopolysaccharide,LPS）to stimulate mouse connective tissue fibroblast L929 and mouse macrophage RAW264.7 to establish a bacterial infection and immune response model in oral and maxillofacial region and to detect the expression of inflammatory cytokines at different time points.Methods The L929 and RAW264.7 cells were stimulated by LPS and the effect of LPS on cell proliferation was determined by CCK-8.QPCR detected the mRNA expression changes of tumor necrosis factor TNF-α,cytokines,chemokines and other related gene.The changes of TNF-αand IL-6 in supernatant of cells were detected by ELISA.Results CCK-8 results showed that LPS could cause changes of cell proliferation.QPCR experimental showed that mRNA expressions of TNF-α,IL-1,IL-5,IL-6,IL-8,IL-23,IL-33 were significantly elevated while the expressions of IL-10,IL-13 decreased markedly.The expressions of CXCL2,CXCL10,FOXO3 and GDF15 were increased significantly.ELISA results showed that 50μg/mL LPS stimulated L929 cells 24 hand 48 h,and the secretion of IL-6 was 42.04 pg/mL and 45.52 pg/mL respectively,The levels of IL-6 secreted by RAW264.7 cells were 25.90 pg/ml and 28.92 pg/ml.The content of IL-6 was 18.97 pg/ml and 21.29 pg/ml in the L929 cells of 100μg/ml LPS stimulation 24 hand 48 h.The content of IL-6 in RAW264.7 cells was 16.90 pg/ml,18.21 pg/ml.Conclusion LPS stimulation of L929 and RAW264.7 cells can induce inflammatory response,increase the expression of related inflammatory factors and chemokines.These changes demonstrated a successful establishment of an in vitro model of oral and maxillofacial infection.

关 键 词：口腔颌面部感染 TNF-Α IL-6 LPS 

分 类 号：R782.3[医药卫生—口腔医学]