钙结合蛋白S100A9在小鼠铜绿假单胞菌肺炎中的表达及意义

发布时间：2018-05-04 18:23

本文选题：S100A9钙结合蛋白 + S100A9mRNA　；参考：《泸州医学院》2013年硕士论文

【摘要】：目的：观察小鼠铜绿假单胞菌肺炎肺组织中钙结合蛋白S100A9mRNA表达，并同时测定对应小鼠血液中S100A9钙结合蛋白含量，观察肺组织的病理改变，探讨钙结合蛋白S100A9在铜绿假单胞菌所致小鼠肺部感染中的地位与作用。方法：小鼠36只，按随机数字表分为空白对照组12只、模型组小鼠24只。通过经环荚膜穿刺注射铜绿假单胞菌菌液建立肺部感染模型。空白对照组（经环荚膜注射等量的2个麦氏单位浓度（6×10~8cfu/ml）0.6ml的铜绿假单胞菌菌液，造模过程中死亡2只）、模型组（经环荚膜注射等量的2个麦氏单位浓度的铜绿假单胞菌菌液0.6ml，造模中死亡5只,分别在在3小时和9小时2个时间点各处死9和10只）。取小鼠右肺下叶HE染色观察肺组织病理，观察空白对照组，3h模型组及9h模型组的病理改变；使用普通PCR检测系统对三组小鼠肺组织中S100A9mRNAPCR反应进程中的产量进行实时监测,最终对实验数据进行分析,从而比较三组小鼠S100A9mRNA表达，并同步采用双抗体夹心酶联免疫吸附测定(ELISA)法分别测定三组小鼠血液中S100A9以及TNF-a的含量，比较三组小鼠血液中生成S100A9钙结合蛋白的差异及与TNF-a变化的相关系。结果：（1）病理观察：大鼠右下肺肺组织HE染色200倍光镜下病理结果显示，正常对照组（1组）肺泡结构较为均匀与清晰，有少许淋巴细胞与巨噬细胞侵润；模型组（3h组）主要为肺泡炎性改变，肺泡内及周围可见渗出，中性粒细胞和淋巴细胞，巨噬细胞浸润;模型组（9h组）肺泡炎性最严重，肺泡内及周围可见大量渗出，大量中性粒细胞和淋巴细胞，巨噬细胞浸润，病理炎性改变明显重于3h组。（2）普通PCR实验:本实验根据电泳后条带亮度的强弱来判断模板拷贝数的高低，或者是表达量的高低。实验结果得出s100A9mRNA在第3h，9h组中的条带亮度强度明显高于空白组，提示S100A9表达量模型组明显高于对照组,且经统计分析后3h与对照组差异具有统计学意义（p0.05)，与9h模型组比较（P0.05）差异有意义。（3）S100A9钙结合蛋白ELISA检测：结果显示3h，9h组外周血中S100A9的含量明显高于空白组，且以3h组增高最明显，3h组与空白组比较差异具有统计学意义（p0.05）与9h组比较（P0.05）有差异。（4）肿瘤坏死因子TNF-a的ELISA检测结果显示：3h，9h组外周血中TNF-a的含量明显高于空白组，差异具有统计学意义（p0.05）。（5）通过对S100A9钙结合蛋白与肿瘤坏死因子-a的相关性分析后，得出两者之间的变化趋势存在正相关，3h模型组r=0.77；9h模型组，，r=0.653.结论：（1）.钙结合蛋白S100A9参与小鼠铜绿假单胞菌肺部炎症反应。（2）.钙结合蛋白S100A9在小鼠铜绿假单胞菌肺炎中与经典炎性因子TNF-a存在密切相关。
[Abstract]:Objective: to observe the expression of calcium-binding protein (S100A9mRNA) in the lung tissues of mice with Pseudomonas aeruginosa pneumonia, and to detect the content of S100A9 calcium binding protein in the blood of corresponding mice, and to observe the pathological changes of lung tissue. To investigate the role of calcium binding protein (S100A9) in pulmonary infection induced by Pseudomonas aeruginosa (Pseudomonas aeruginosa) in mice. Methods: 36 mice were randomly divided into control group (n = 12) and model group (n = 24). Pulmonary infection model was established by injection of Pseudomonas aeruginosa through circular capsule. Blank control group (injected with the same amount of 2 McClone unit concentrations of 6 脳 10~8cfu/ml)0.6ml Pseudomonas aeruginosa solution through circular capsule), In the model group, two rats died in the process of modeling, and in the model group, 9 and 10 rats were killed at 3 hours and 9 hours, respectively, and 5 rats were killed in the model group (injection of equal amount of 0.6 ml of Pseudomonas aeruginosa per unit concentration of 2 McClomonas aeruginosa) through the capsule of the model, and 9 hours, respectively. The histopathology of lung tissue was observed by HE staining in the lower lobe of the right lung of mice, and the pathological changes were observed in the 3 h model group and 9 h model group of the blank control group, and the output of the S100A9mRNAPCR reaction process in the lung tissues of the three groups was monitored by ordinary PCR detection system in real time. Finally, the experimental data were analyzed to compare the expression of S100A9mRNA in the three groups of mice, and the contents of S100A9 and TNF-a in blood of the three groups were measured by double antibody sandwich enzyme-linked immunosorbent assay (Elisa). To compare the difference of S100A9 calcium binding protein in blood of three groups and its relationship with the change of TNF-a. Results (1) pathological observation: the pathological results showed that the alveolar structure of the right lower lung tissue of the rats was uniform and clear, with a few lymphocytes and macrophages infiltrating into the lung tissue of the right lower lung tissue by HE staining for 200 times under the light microscope. The results showed that the alveolar structure was more uniform and clear in the normal control group (1 group). In the model group (3 h), the alveolar inflammatory changes were mainly observed, the infiltration of neutrophils, lymphocytes and macrophages was observed in and around the alveoli, and the alveolar inflammation was the most serious in the model group (9 h), and a large amount of exudation was observed in and around the alveoli. A large number of neutrophils and lymphocytes, macrophages infiltration, pathological inflammatory changes were significantly more serious than the 3h group. (2) ordinary PCR experiment: this experiment according to the intensity of the band brightness after electrophoresis to judge the template copy number, or the high or low level of expression. The results showed that the luminance intensity of s100A9mRNA was significantly higher in the 3h / 9h group than in the blank group, suggesting that the expression of S100A9 in the model group was significantly higher than that in the control group. After statistical analysis, there was a significant difference between the control group and the control group at 3h, and the difference was significant compared with the model group at 9h (P0.05). The results showed that the content of S100A9 in the peripheral blood of the 3h / 9h group was significantly higher than that of the blank group. The results of ELISA detection of tumor necrosis factor (TNF-a) showed that the level of TNF-a in peripheral blood of the 3h group was significantly higher than that of the control group (P 0.05), and that of the control group was significantly higher than that of the control group (P < 0.05). By analyzing the correlation between S100A9 calcium binding protein and tumor necrosis factor-a, it was found that there was a positive correlation between the changes of S100A9 calcium binding protein and tumor necrosis factor-a in 3h model group. Conclusion: 1: 1. Calcium binding protein (S100A9) is involved in pulmonary inflammation of Pseudomonas aeruginosa in mice. Calcium binding protein S100A9 is closely related to the classical inflammatory factor TNF-a in mouse pseudomonas aeruginosa pneumonia.
【学位授予单位】：泸州医学院
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R563.1

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