低氧诱导因子-1α在小鼠铜绿假单胞菌肺炎中的作用研究
发布时间：2018-11-07 20:11
【摘要】：目的：通过研究低氧诱导因子-1α (Hypoxia inducing factor-1α)在铜绿假单胞菌肺炎模型小鼠肺组织中的表达情况，以及探讨HIF-1α与TNF-α、IL-8之间的相关性，推测HIF-1α在铜绿假单胞菌肺炎中的可能作用，以期为铜绿假单胞菌肺炎的治疗寻找到新思路。方法：1.将鼠龄6~8周、体重25~30g的清洁级健康雄性昆明小鼠120只按随机原则分成肺部感染组（PA组：60只）和正常对照组（生理盐水组：60只）。PA组：氯胺酮麻醉小鼠后固定，常规消毒铺巾，严格无菌操作，切开颈部皮肤，钝性分离至皮下组织，暴露上段气管，经气管内穿刺注入铜绿假单胞菌（PA）菌液（2个麦氏单位浓度、0.03ml）建立小鼠铜绿假单胞菌肺炎模型；正常对照组：操作同PA组，经气管内穿刺注入无菌生理盐水（0.03ml）作为对照，两组均于造模成功后第3、9、24小时随机分别抽取20只小鼠，予以留取标本后处死。2.收集各组不同时间点处死的小鼠血液，离心分离血清后，以酶联免疫吸附试验（enzyme-linked immunosorbnent assay，ELISA）检测血清中TNF-α、IL-8的浓度；取右肺经10%多聚甲醛固定后，切片行HE染色比较炎症的范围及程度，并行免疫组化染色检测肺组织中HIF-1α蛋白的表达水平；同时留取左侧肺组织行肺组织匀浆细菌培养及菌落计数。3.应用SPSS17.0软件对数据进行统计分析。结果：1、PA感染组小鼠肺组织病理改变在24小时最为严重，炎症范围广，程度重，3、9小时与之相比均较轻，，但均有明显的炎症改变；而对照组的肺泡及肺间质结构基本正常，无明显出血及炎细胞浸润。并且PA感染组3、9、24小时肺组织匀浆均培养出铜绿假单胞菌，菌落计数均105CFU/g，而对照组肺组织匀浆各时间点均未培养出细菌。2、PA感染组3、9、24小时小鼠肺组织中HIF-1α蛋白的表达均有明显的增高，其中细菌接种3小时后HIF-1α的表达就有升高，9小时继续升高，24小时最高。而对照组HIF-1α各时间点均未见明显表达。3、PA感染组3、9、24小时小鼠血清中TNF-α、IL-8的水平较对照组均有明显的增高，差异有统计学意义（P＜0.05）。其中TNF-α、IL-8的水平均在3小时开始上升，9小时继续升高，24小时最高。4、HIF-1α蛋白的表达与TNF-α、IL-8的浓度呈正相关（相关系数分别为0.827和0.974）。结论：1. HIF-1α参与了铜绿假单胞菌肺炎的炎症过程，且炎症愈重表达愈明显。2. HIF-1α是炎症细胞因子网络中的重要一环，它与TNF-α、IL-8等相互作用，促进炎症细胞合成及释放多种炎症因子，在铜绿假单胞菌肺炎的发生发展中起了重要作用。
[Abstract]:Objective: to study the expression of hypoxia inducible factor-1 伪 (Hypoxia inducing factor-1 伪 in lung tissue of mice with Pseudomonas aeruginosa pneumonia and to explore the correlation between HIF-1 伪 and TNF- 伪, IL-8. The possible role of HIF-1 伪 in Pseudomonas aeruginosa pneumonia was speculated in order to find a new idea for the treatment of Pseudomonas aeruginosa pneumonia. Method 1: 1. A total of 120 healthy male Kunming mice were randomly divided into two groups: lung infection group (PA group: 60 mice) and normal control group (normal saline group: 60). PA mice). Routine disinfecting and spreading towel, strict aseptic operation, incision of neck skin, blunt separation to subcutaneous tissue, exposure of upper trachea, puncture and injection of Pseudomonas aeruginosa (PA) solution (2 McClone unit concentrations) in trachea through trachea. 0.03ml) to establish the model of Pseudomonas aeruginosa pneumonia in mice. Normal control group: in the same operation as PA group, 20 mice were randomly selected at 24 hours after successful modeling and 20 mice were randomly selected from the two groups after puncture and injection of sterile saline (0.03ml) into the trachea. 2. 2. Blood samples were collected from mice killed at different time points. After centrifugation, the concentrations of TNF- 伪 and IL-8 in serum were detected by enzyme linked immunosorbent assay (enzyme-linked immunosorbnent assay,ELISA). After the right lung was fixed with 10% paraformaldehyde, HE staining was performed to compare the extent and degree of inflammation, and the expression of HIF-1 伪 protein in lung tissue was detected by immunohistochemical staining. At the same time, the left lung tissue was retained for the culture of bacteria and colony count of lung tissue homogenate. The data were analyzed by SPSS17.0 software. Results: 1 the pathological changes of lung tissue in PA infected mice were the most serious in 24 hours. The range of inflammation was wide and the degree of inflammation was severe. Compared with the control group, the pathological changes of lung tissue in the group of 3 ~ 9 hours were lighter, but there were obvious inflammatory changes. In the control group, the alveolar and interstitial structures were normal, no obvious bleeding and inflammatory cell infiltration. Pseudomonas aeruginosa was cultured in lung tissue homogenate of PA infection group at 24 hours, and colony counts were 105 CFU / g, but no bacteria were cultured in lung tissue homogenate of control group. The expression of HIF-1 伪 protein in lung tissue of 24 h mice was significantly increased. The expression of HIF-1 伪 was increased 3 hours after inoculation, and continued to increase at 9 h, and reached the highest level at 24 h. The levels of TNF- 伪 and IL-8 in the control group were significantly higher than those in the control group (P < 0. 05). The levels of TNF- 伪 and IL-8 began to rise at 3 hours, continued to increase at 9 hours, and reached the highest at 24 hours. The expression of HIF-1 伪 protein and TNF- 伪 in 24 hours were the highest. The concentration of IL-8 was positively correlated (correlation coefficient was 0.827 and 0.974, respectively). Conclusion 1. HIF-1 伪 was involved in the inflammatory process of Pseudomonas aeruginosa pneumonia, and the higher the inflammatory expression was, the more obvious. 2. HIF-1 伪 is an important link in the inflammatory cytokine network. It interacts with TNF- 伪, IL-8 and so on, promotes the synthesis and release of many inflammatory factors by inflammatory cells, and plays an important role in the occurrence and development of 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
本文编号：2317445
[Abstract]:Objective: to study the expression of hypoxia inducible factor-1 伪 (Hypoxia inducing factor-1 伪 in lung tissue of mice with Pseudomonas aeruginosa pneumonia and to explore the correlation between HIF-1 伪 and TNF- 伪, IL-8. The possible role of HIF-1 伪 in Pseudomonas aeruginosa pneumonia was speculated in order to find a new idea for the treatment of Pseudomonas aeruginosa pneumonia. Method 1: 1. A total of 120 healthy male Kunming mice were randomly divided into two groups: lung infection group (PA group: 60 mice) and normal control group (normal saline group: 60). PA mice). Routine disinfecting and spreading towel, strict aseptic operation, incision of neck skin, blunt separation to subcutaneous tissue, exposure of upper trachea, puncture and injection of Pseudomonas aeruginosa (PA) solution (2 McClone unit concentrations) in trachea through trachea. 0.03ml) to establish the model of Pseudomonas aeruginosa pneumonia in mice. Normal control group: in the same operation as PA group, 20 mice were randomly selected at 24 hours after successful modeling and 20 mice were randomly selected from the two groups after puncture and injection of sterile saline (0.03ml) into the trachea. 2. 2. Blood samples were collected from mice killed at different time points. After centrifugation, the concentrations of TNF- 伪 and IL-8 in serum were detected by enzyme linked immunosorbent assay (enzyme-linked immunosorbnent assay,ELISA). After the right lung was fixed with 10% paraformaldehyde, HE staining was performed to compare the extent and degree of inflammation, and the expression of HIF-1 伪 protein in lung tissue was detected by immunohistochemical staining. At the same time, the left lung tissue was retained for the culture of bacteria and colony count of lung tissue homogenate. The data were analyzed by SPSS17.0 software. Results: 1 the pathological changes of lung tissue in PA infected mice were the most serious in 24 hours. The range of inflammation was wide and the degree of inflammation was severe. Compared with the control group, the pathological changes of lung tissue in the group of 3 ~ 9 hours were lighter, but there were obvious inflammatory changes. In the control group, the alveolar and interstitial structures were normal, no obvious bleeding and inflammatory cell infiltration. Pseudomonas aeruginosa was cultured in lung tissue homogenate of PA infection group at 24 hours, and colony counts were 105 CFU / g, but no bacteria were cultured in lung tissue homogenate of control group. The expression of HIF-1 伪 protein in lung tissue of 24 h mice was significantly increased. The expression of HIF-1 伪 was increased 3 hours after inoculation, and continued to increase at 9 h, and reached the highest level at 24 h. The levels of TNF- 伪 and IL-8 in the control group were significantly higher than those in the control group (P < 0. 05). The levels of TNF- 伪 and IL-8 began to rise at 3 hours, continued to increase at 9 hours, and reached the highest at 24 hours. The expression of HIF-1 伪 protein and TNF- 伪 in 24 hours were the highest. The concentration of IL-8 was positively correlated (correlation coefficient was 0.827 and 0.974, respectively). Conclusion 1. HIF-1 伪 was involved in the inflammatory process of Pseudomonas aeruginosa pneumonia, and the higher the inflammatory expression was, the more obvious. 2. HIF-1 伪 is an important link in the inflammatory cytokine network. It interacts with TNF- 伪, IL-8 and so on, promotes the synthesis and release of many inflammatory factors by inflammatory cells, and plays an important role in the occurrence and development of 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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