小鼠模型感染大片形吸虫研究平台初探

发布时间：2018-07-12 16:55

本文选题：大片形吸虫 + 囊蚴培养　；参考：《广西大学》2015年硕士论文

【摘要】：片形吸虫病主要侵害宿主的肝胆系统,且该病早期诊断困难,即便后期用药驱除了虫体,对肝脏造成的损伤也较难修复。因此探明大片形吸虫感染后肝脏损伤的相关机制,对丰富大片形吸虫和宿主之间的相互作用机制及感染宿主的肝脏治疗和康复具有极其重要的意义。本研究内容包括：大片形吸虫囊蚴实验室繁育方案的优化完善,大片形吸虫-昆明小鼠感染模型的建立以及感染昆明小鼠肝脏病理学的初步研究；这些研究旨在为大片形吸虫肝脏免疫和组织损伤修复等下游研究夯实基础。首先完善优化实验室人工培养大片形吸虫囊蚴体系,并对大片形吸虫在中间宿主体内的发育特点进行观察。分别以1,3,7条毛蚴感染2～3mm,5～7mm两种体型大小的小土蜗螺,在室温26℃时,感染后10～17天可见母雷蚴,28天可见子雷蚴体内充满尾蚴,30～33天可见尾蚴释放并结囊为囊蚴。对比分析不同大小的螺感染不同数量的毛蚴最后产囊蚴率结果,得出最佳培养方案与肝片形吸虫囊蚴培养不同,大片形吸虫囊蚴培养选用3-5mm大小的螺,每螺感染3条毛蚴时产囊蚴量最多。以5、15、30和50个囊蚴经口感染小鼠,于不同时期分别剖检以观察虫体分布、回收、发育情况,以及肝脏的病理变化。结果显示：感染后5天内便可在肝实质探查到虫体,35-42天内童虫完成从肝实质向胆管移行,但各感染组均未发现成虫；大片形吸虫在昆明小鼠体内异位寄生现象发生频率较高,肺部、口腔、膈肌、前肢皮下等部位均可发现虫体；受感染昆明小鼠肝脏可见弥散出血、化脓、脂变、纤维化、坏死、硬变等明显肉眼病理变化,病变程度随感染强度的增加而加重。表明大片形吸虫-昆明小鼠模型适宜用于童虫期的研究,15个囊蚴的感染强度是动态观察昆明小鼠宿主状态及相关研究的适宜感染剂量；小鼠为大片形吸虫的非适宜宿主,不宜作为研究反刍动物(适宜宿主)感染大片形吸虫动物模型。观察人工感染大片形吸虫小鼠的肝脏组织学病理变化显示：感染后7天,肝脏肉眼可见轻微损伤,HE染色可见肝细胞排列开始紊乱；感染后21～42天,肝脏损失逐渐加重,显微镜下可观察到肝细胞坏死、结缔组织增生、炎性细胞的增生浸润等典型的肝组织病理学改变。检测感染大片形吸虫的小鼠血清中碱性磷酸酶(ALP)、谷草转氨酶(AST)、谷丙转氨酶(ALT)、总蛋白(TP)、白蛋白(ALB)、球蛋白(GLOB)及白球比(A/G)的变化,显示AST在感染后第3天开始升高,21～56天显著升高(P0.01),49天达峰值；ALT于感染后14天开始升高,21～35天开始显著升高(P0.01),49天达峰值；ALP呈现感染后14～21天升高,35～49天下降,56天又开始回升的波动变化;GLOB于感染后21～56天总体呈上升趋势；ALB自感染后呈现下降趋势,感染后49天达最低值；表明：血清肝脏损失相关因子的变化与肝脏组织病变动态发展相一致；AST.ALT可作为感染急性期指标,ALB.GLOB和A/G比值的范围可评估大片形吸虫病所引起的肝损伤程度,联合检测这些生化指标对大片形吸虫病诊断及预后具有重要意义。
[Abstract]:Fasciosis mainly infringes the liver and bile system of the host, and it is difficult to diagnose the early stage of the disease. Even if the later drug expelling the insect body, the damage to the liver is difficult to repair. Treatment and rehabilitation are of great significance. This study includes the optimization of laboratory breeding programs for large Fasciola cercariae, the establishment of large Fasciola - Kunming mice infection model and the preliminary study of liver pathology in Kunming mice; these studies are aimed at repairing the liver immunity and tissue damage of large Fasciola. In the first step, we should improve the foundation of the study of the lower reaches of the lower reaches. First, improve the artificial cultivation of large tracer cercariae in the laboratory, and observe the development characteristics of large tracts in the middle host. The 1,3,7 cercariae infect 2 to 3mm, 5 to 7mm, and two kinds of small snail size. At room temperature 26, the larvae can be seen for 10~17 days after infection. On the 28 day, the cercariae were full of cercariae in the cercariae, the cycariae were released on the 30~33 day and cysts were cysts. Compared and analyzed the results of the final production of cercariae in different numbers of different sizes of the cercariae, the best training scheme was different from the culture of the liver slice form of the cercariae. The 3-5mm size snails were selected for the cultivation of the large fasciolus cercariae, and 3 hairs per snails were infected. The amount of cercariae was the most. The mice infected with 5,15,30 and 50 cercariae were infected in different periods to observe the distribution, recovery, development, and pathological changes of the liver. The results showed that the insect body could be detected in the liver parenchyma within 5 days after infection, and in 35-42 days, the children migrated from the liver parenchyma to the bile duct, but all the infected groups were all infected. No adult worms were found. The occurrence of ectopic parasitism in Kunming mice was high, and the body was found in the lungs, the mouth, the diaphragm and the subcutaneous parts of the forelimb. The liver of the infected Kunming mice showed diffuse hemorrhage, pyogenic, fat, fibrosis, necrosis and hard change, and the degree of pathological changes increased with the intensity of infection. It was added and aggravated. It showed that the model of large bugs - Kunming mice was suitable for the study of the stage of childhood. The infection intensity of the 15 cycariae was a dynamic observation of the host state of Kunming mice and the appropriate dose of infection. Histopathological changes of liver in mice with artificial infection of large tracts showed that 7 days after infection, the liver was slightly damaged in the naked eye, the liver cell arrangement began to be disorganized by HE staining, and the liver damage was gradually increased on the 21~42 day after infection. The necrosis of hepatocytes, connective tissue proliferation and inflammatory cells increased under the microscope. The changes in serum alkaline phosphatase (ALP), glutamic pyruvic aminotransferase (AST), glutamic pyruvic aminotransferase (ALT), total protein (TP), albumin (ALB), globulin (GLOB) and white ball ratio (A/G) in mice infected with Fasciola Fasciola showed that AST increased at third days after infection and increased significantly (P0.01), 49 (P0.01), 49. The peak value of Tianda, ALT began to rise on the 14 day after infection, the 21~35 days began to rise significantly (P0.01), 49 Tianda peak, ALP showed an increase of 14~21 days after infection, 35~49 days, and the 56 days began to rebound, and the overall increase trend was 21~56 days after infection; ALB showed a downward trend after infection and the lowest value of 49 Tianda after infection. The changes in serum liver loss related factors are consistent with the dynamic development of liver tissue lesions, and AST.ALT can be used as an indicator of acute infection, and the range of ALB.GLOB and A/G ratios can be used to assess the degree of liver damage caused by large Fasciola. The combined detection of these biochemical indexes is important for the diagnosis and prognosis of large Fasciola. Righteousness.
【学位授予单位】：广西大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S855.99

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