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目的:利用耻垢分枝杆菌(M.smegmatismc2155)建立C57BL/6小鼠结核病模型。方法:每天以高剂量(5×107CFU)耻垢分枝杆菌给C57BL/6小鼠腹腔注射,连续感染4周,检测耻垢分枝杆菌对小鼠的致病性。分别于2周和4周处死小鼠,无菌条件下解剖小鼠取肺、脾脏组织匀浆,进行组织内细菌活力检测;通过嗜酸性染色进行分枝杆菌的鉴定;同时进行病理切片的制备,观察肺和脾脏组织的病理变化;最后进行菌体DNA的提取和基因检测,根据上述指标确定小鼠结核病模型的建立是否成功。结果:腹腔感染小鼠2周后,模型组小鼠只有脾脏组织匀浆液出现抗酸染色阳性菌落,肺部组织未见阳性菌落。腹腔感染小鼠4周后,模型组小鼠肺、脾脏组织匀浆液中均可见大量抗酸染色阳性的菌落;组织病理学观察结果显示:小鼠肺组织主要表现为以中性粒细胞为主的炎性病变;基因检测结果表明:模型组小鼠肺组织匀浆液中可检测到耻垢分枝杆菌特异性3-磷酸甘油醛脱氢酶(gap)基因,而脾脏组织未扩增出耻垢分枝杆菌特异性基因。结论:通过腹腔注射无致病性耻垢分枝杆菌方法,成功建立C57BL/6小鼠结核病发生模型,为结核分枝杆菌与宿主相互作用研究提供安全的疾病模型。
OBJECTIVE: To establish a C57BL / 6 mouse tuberculosis model by using M. smegmatism c2155. Methods: C57BL / 6 mice were injected intraperitoneally with high dose (5 × 107CFU) Mycobacterium smegmatis for 4 weeks. The pathogenicity of Mycobacterium smegmatis to mice was tested. Mice were sacrificed at 2 weeks and 4 weeks respectively, and the mice were dissected from the lungs and spleens under aseptic conditions to detect the activity of bacteria in the tissues. Mycobacterium was identified by eosinophilic staining. At the same time, pathological sections were prepared , Observe the pathological changes of lung and spleen tissue; finally DNA extraction and genetic testing of cells, according to the above indicators to determine the establishment of the mouse model of tuberculosis was successful. Results: Two weeks after intraperitoneal infection in mice, only the spleen homogenates showed positive acid-fast staining colonies and no positive colon in lung tissues. After 4 weeks of intraperitoneal infection in mice, a large number of colonies positive for acid-fast staining were found in the homogenate of lung and spleen of mice in model group. Histopathological observation showed that the main manifestations of lung tissue in mice were neutrophil Of the inflammatory lesions; gene test results showed that: model group mice lung tissue homogenate can detect mycobacterium smegmatis-specific glyceraldehyde-3-phosphate dehydrogenase (gap) gene, while the spleen did not amplify the shame Mycobacterium smegmatis-specific gene. Conclusion: The model of tuberculosis in C57BL / 6 mice was successfully established by intraperitoneal injection of non-pathogenic Mycobacterium smegmatis and provided a safe disease model for the study on the interaction between Mycobacterium tuberculosis and host.