免疫抑制状态下新西兰大白兔肺部真菌感染模型建立的研究

发布时间：2018-03-03 11:42

本文选题：新型隐球菌　切入点：白念菌　出处：《第二军医大学》2008年硕士论文　论文类型：学位论文

【摘要】： 目的:建立适合影像学研究使用的新西兰大白兔的新型隐球菌、白念菌、烟曲霉菌肺炎动物模型,为进一步的临床、病理、影像学诊断对照研究打下基础。材料与方法:选择健康雄性新西兰大白兔72只,分为新型隐球菌组、白念菌组、烟曲霉组,每组中设立实验组18只,对照组6只,新型隐球菌对照组为未注射免疫抑制剂,而接种了新型隐球菌悬浮液,白念菌组和曲霉菌组对照组为注射了免疫抑制剂而注射了等量的生理盐水。新型隐球菌实验组和白念菌组使用相同的方法使兔达到免疫抑制状态:于实验第1-5天,每日耳缘静脉注射阿糖胞苷440mg/m~2,6天后隔日一次维持免疫抑制状态。第4日起,所有兔给予万古霉素15mg/kg、头孢他定150mg/kg静脉注射,每日1次;庆大霉素5mg/kg静脉注射,隔日1次预防细菌感染。于第6日行经皮气管内穿刺法分别接种配置好新型隐球菌0.4ml(浓度2.5×10~8个/ml)悬浮液和白念菌悬浮液0.2ml(浓度5×10~8个/ml)的悬浮液,白念菌对照注射等量的生理盐水。烟曲霉组于实验第1-5天、第8、9天,每日耳缘静脉注射阿糖胞苷525mg/m~2、第1-2天每日耳缘静脉注射甲基强地松龙5mg/kg,第4天起给予万古霉素15mg/kg、头孢他定150mg/kg静脉注射,每日1次;庆大霉素5mg/kg静脉注射,隔日1次预防细菌感染,于第2日行经皮气管内穿刺法接种配置好的0.4ml烟曲霉悬浮液(浓度为1×10~8个/ml),对照组注射等量的生理盐水。每组分别于实验前和接种前抽耳缘静脉血做血常规检查了解粒细胞值的变化。每组所用兔在实验开始时进行一次胸部CT检查,接种后每隔一天行胸部CT检查了解肺部的影像学表现,并于接种后每2-3天随机处死3-4只肺部有阳性表现兔(如有兔死亡量酌减),无菌操作下剖胸取出肺组织标本行真菌培养和病理检查,了解上述真菌在肺内的生长和致病情况。结果:1、三种组兔耳缘静脉血粒细胞计数在注射免疫抑制剂后明显降低,行配对t检验P＜0.05,有统计学意义。 2、新型隐球菌组的实验组接种成功率为100%(18/18),模型建立成功率为83.33%(15/18)。对照组模型建立成功率为33.33%(2/6),与实验组统计学有明显差异。 3、白念菌组的实验组接种成功率为100%(18/18),模型建立成功率为88.89%(16/18),对照组模型建立成功率为16.67%(1/6),与实验组统计学有明显差异。 4、烟曲霉组的实验组接种成功率为100%(18/18),模型建立成功率88.89%(16/18),对照组接种和模型成功率均为0%。 5、胸部CT检查主要异常表现为GGO、实变,可伴有支气管血管束增粗、结节和网格线影。结论:新西兰大白兔免疫抑制后使用经皮穿刺气管内接种法将新型隐球菌、白念菌和烟曲霉菌注入兔肺内并进行定期CT扫描的方法了解肺内病变情况可以建立适合影像学研究使用的上述三种真菌肺部感染的动物模型,此方法成功率高、具有可操作性、可重复性好,其中兔隐球菌肺炎模型的建立国内外尚无文献报道,为以后的影像病理对照研究打下了良好的基础。
[Abstract]:Objective: to establish a new animal model of Cryptococcus neoformans, white fungus and Aspergillus fumigatus, which is suitable for imaging study, and lay a foundation for further clinical, pathological and imaging diagnosis.
Materials and methods: 72 male healthy New Zealand rabbits, divided into groups of Cryptococcus neoformans, Candida albicans, Aspergillus fumigatus group, set up the experimental group with 18 rats in each group, 6 rats in the control group, the control group is not Cryptococcus neoformans immune inhibitor, and inoculated with Cryptococcus albicans suspension. A group of bacteria and Aspergillus group control group injected immunosuppressant and injected with normal saline. Cryptococcus neoformans and Candida albicans were the experimental group using the same method to make rabbit immune suppression: on the 1-5 day, the daily auricular intravenous cytarabine 440mg/m~2,6 days after every other day for maintenance immunosuppression state. On the fourth day, all rabbits treated with vancomycin 15mg/kg, ceftazidime 150mg/kg intravenous injection, 1 times a day; gentamicin 5mg/kg intravenous injection, 1 every other day to prevent the bacterial infection. On the sixth day after percutaneous tracheal puncture inoculation respectively configured Cryptococcus 0.4ml (concentration of 2.5 * 10~8 /ml) suspension and Candida albicans suspension 0.2ml (concentration of 5 * 10~8 /ml) suspension of Candida bacteria injected with saline control group. Aspergillus fumigatus in the 1-5 days, 8,9 days, daily auricular intravenous cytarabine 525mg/m~2, the 1-2 day of ear vein injection of methylprednisolone 5mg/kg, fourth days for vancomycin 15mg/kg, ceftazidime 150mg/kg intravenous injection, 1 times a day; gentamicin 5mg/kg intravenous injection, 1 times a day to prevent bacterial infection, second underwent percutaneous tracheal puncture with a configured 0.4ml of Aspergillus fumigatus suspension (concentration of 1 * 10~8 /ml), the control group was injected with normal saline. Each group before experiment and inoculation before pumping ear vein blood for blood routine examination to understand the changes of neutrophils. The value of each rabbit at the start of a chest CT examination after inoculation Every other day to understand the chest CT examination of lung imaging performance, and in every 2-3 days after inoculation were sacrificed positive rabbits 3-4 lungs (such as rabbit mortality reduction), aseptic operation thoracotomy lung tissue specimens for fungal culture and pathological examination, understand the growth and pathogenicity of fungi in the lungs.
Results: 1, three groups of rabbit ear vein blood granulocyte count decreased significantly after injection of immunosuppressive agents, and paired t test was P < 0.05, with statistical significance.
2, the success rate of inoculation in Cryptococcus neoformans group was 100% (18/18), and the success rate of model establishment was 83.33% (15/18). The success rate of the control group was 33.33% (2/6), which was significantly different from that of the experimental group.
3, the inoculation success rate of the experimental group was 100% (18/18), the success rate of the model establishment was 88.89% (16/18), and the success rate of the control group was 16.67% (1/6), which was significantly different from that of the experimental group.
4, the successful rate of inoculation in the experimental group of Aspergillus fumigatus was 100% (18/18), the success rate of the model was 88.89% (16/18), and the success rate of inoculation and model in the control group was 0%.
5, the main abnormalities of the chest CT examination were GGO, real changes, accompanied by bronchovascular bundle thickening, nodules and grid lines.
Conclusion: New Zealand white rabbits were immunized after inhibition of using percutaneous intratracheal inoculation method of Cryptococcus neoformans, method of Candida albicans and Aspergillus fumigatus injected into rabbit lung and regular CT scan can be established for the understanding of pulmonary lesions imaging study using the above three kinds of pulmonary fungal infection animal model, the success rate of this method high maneuverability, good repeatability, has not been reported at home and abroad to establish rabbit model of cryptococcal pneumonia, imaging and pathologic for later control study to lay a good foundation.

【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R519;R-332

【参考文献】