宿主免疫抑制对青蒿琥酯抗日本血吸虫作用的影响及日本血吸虫成虫自发荧光研究

发布时间：2018-01-01 17:07

本文关键词：宿主免疫抑制对青蒿琥酯抗日本血吸虫作用的影响及日本血吸虫成虫自发荧光研究　出处：《苏州大学》2009年硕士论文　论文类型：学位论文

【摘要】： 日本血吸虫病是一种严重危害我国人民健康、影响经济发展的寄生虫病。化疗作为血吸虫病综合性防治措施的重要组成部分,其目的在于通过杀灭人和哺乳动物体内的血吸虫,降低人、畜感染率和感染度,获得控制患病和消灭传染源的双重效果。我国和某些以病人为血吸虫病主要传染源的国家,在安全有效、使用方便的特效药问世之前,化疗已被列为主要措施之一,与灭螺并列为两大固定防治措施。青蒿琥酯(Artesunate)等青蒿素类药物对日本血吸虫病和曼氏血吸虫病的早期预防治疗效果肯定,并已在多次现场应用和抗洪救灾中发挥了积极有效的作用。本实验旨在研究青蒿琥酯的抗日本血吸虫作用与宿主免疫功能状态的关系,特别是用氢化可的松(Hydrocortisone)造模观察宿主免疫功能抑制状态对青蒿琥酯抗血吸虫作用的影响,以期为各种原因导致的机体免疫缺陷人群更加合理、安全、有效的用药提供参考和依据。另外,我们还利用荧光显微镜和激光共聚焦扫描显微镜对日本血吸虫成虫的自发荧光现象进行了观察和光谱学信息分析。第一部分宿主免疫抑制对青蒿琥酯抗日本血吸虫作用的影响目的:研究宿主免疫抑制对青蒿琥酯抗日本血吸虫作用的影响。方法:小鼠常规方法实验室感染日本血吸虫尾蚴,每鼠50±2条,并随机分为4组:对照组、氢化可的松组(HC组)、青蒿琥酯组(Art组)和氢化可的松+青蒿琥酯组(HC+Art组)。感染后第7d开始腹腔注射氢化可的松构建免疫抑制模型,25mg/kg.d,连续7d。感染后第14d一次灌胃给药青蒿琥酯混悬液,400mg/kg。感染7wk后剖杀所有小鼠取虫计数。同样造模方法于感染后第14d开始注射氢化可的松,第21d青蒿琥酯灌胃治疗,感染7wk后取虫做盐酸卡红染色制片。结果:青蒿琥酯在免疫抑制小鼠体内的杀虫作用较之于在免疫功能正常小鼠体内显著减弱,减虫率分别为19.92%和33.00%(P0.05);在免疫抑制小鼠模型中,使用青蒿琥酯治疗与否,其减虫率分别为19.92%和19.28%,并无显著性差异(P0.05)。盐酸卡红染色直观显示虫体肠管大体变化也提示,青蒿琥酯在免疫抑制模型中对虫体的损伤作用明显减弱。结论:青蒿琥酯的抗日本血吸虫效果与宿主的免疫功能状态有着密切关系,在宿主免疫功能抑制状态下青蒿琥酯甚至不能发挥有效的杀虫作用。宿主正常的免疫功能状态可能是青蒿琥酯充分发挥杀虫作用的重要前提之一。第二部分日本血吸虫成虫自发荧光观察及共聚焦λ扫描分析目的:研究日本血吸虫成虫自发荧光现象及共聚焦λ扫描特点,丰富日本血吸虫生物学和光谱学信息。方法:小鼠感染日本血吸虫尾蚴6wk后剖杀,收集成虫于无菌生理盐水中并分离雌雄虫体。荧光显微镜下分别以紫外光、蓝光、绿光等不同激发光源激发观察虫体自发荧光;激光共聚焦扫描显微镜下分别以405nm、458nm、476nm、488nm、514nm、543nm、633nm等7种激发光激发雄虫口吸盘部做自发荧光λ扫描分析。结果:不同激发光照射下日本血吸虫虫体能发出多种不同颜色的自发荧光,以蓝光激发的黄绿色荧光效果最佳,虫体大体结构清晰;λ扫描分别以405nm、488nm、514nm和543nm激发光激发样品,相应敏感的发射波长分别为490-510nm、550-570nm、560-580nm和590-600nm,其中488nm和514nm激发效果较好。458nm、476nm、633nm激发效果欠佳。结论:荧光显微镜下可观察日本血吸虫成虫自发荧光,最佳模式是以蓝光激发观察黄绿色荧光;日本血吸虫雄虫自发荧光光谱较宽,主要分布于500-600nm;488nm和514nm为较适宜的激发波长。
[Abstract]:Schistosomiasis is a serious harm to the health of our people, parasitic diseases affecting economic development. Chemotherapy as an important part of comprehensive schistosomiasis control measures, its purpose is to reduce people by killing human and mammalian in vivo, and the infection rate of Schistosoma infection rate of livestock, to gain control of double effect of illness and eliminate the source of infection. Our country and some to the patient as the main infection sources of schistosomiasis in the country, safe and effective, convenient use before the advent of medicine, chemotherapy has been listed as one of the main measures, and the snail is one of two major fixed control measures. Artesunate (Artesunate) etc. artemisinins at early stage of schistosomiasis and the prevention and treatment of schistosomiasis mansoni a positive effect, and has several field application and flood relief has played a positive and effective role. This experiment aims to study the Anti Japanese artesunate The relationship between schistosome and host immune function, especially the use of hydrocortisone (Hydrocortisone) effect of anti schistosomiasis effect model to observe the host immune function inhibition of artesunate, in order for all causes of immunodeficiency crowd is more reasonable, safe, and provide reference for effective medication. In addition, we also use fluorescence microscopy confocal laser scanning microscopy and autofluorescence of adult Schistosoma japonicum were analyzed to observe and spectroscopic information.
The effect of artesunate against Schistosoma japonicum in the first part of the host immunosuppression
Objective: To study the effect of host immunosuppression on the effect of artesunate against Schistosoma japonicum.
Methods: the conventional method of laboratory mice infected with Schistosoma japonicum cercariae per mouse, 50 + 2, and randomly divided into 4 groups: control group, hydrocortisone group (HC group), artesunate group (Art group) and hydrocortisone plus artesunate group (HC+Art group). 7d after infection of peritoneal injection of hydrocortisone construction the model of immune suppression, 25mg/kg.d, continuous 7d. infection 14d after an intragastric administration of artesunate suspension, 400mg/kg. infection after 7wk killed all mice. The same insect count model after infection after 14d injection of hydrocortisone, 21d artesunate orally treated, 7wk infection after worm hydrochloric acid carmine staining producer.
Results: compared with the insecticidal effect of artesunate in immunosuppressive mice significantly reduced the immune function in normal mice, the worm reduction rates were 19.92% and 33% (P0.05); immune suppression in the mouse model, the use of artesunate in the treatment or not, the worm reduction rates were 19.92% and 19.28%, there was no significant difference (P0.05). Display card hydrochloride red staining insect intestinal gross changes also suggest that artesunate in immunosuppressive effect on body injury model was significantly decreased.
Conclusion: the effect of artesunate against Schistosoma japonicum and the immune function of the host is closely related to the inhibitory condition of artesunate can not even play an effective role in killing the host immune function. Normal immunity of the host may be one of the important premise of the antischistosomal effect of artesunate.
The second part of the autofluorescence of adult Schistosoma japonicum and the analysis of CO focal lambda scanning
Objective: To study the characteristics of the spontaneous fluorescence of Schistosoma japonicum and the characteristics of the co focal lambda scanning, and to enrich the biological and spectroscopic information of Schistosoma japonicum.
Methods: mice infected with cercariae of Schistosoma japonicum 6wk after slaughter, was collected in sterile saline and separating the male and female worms. Under fluorescence microscope respectively with UV light, blue light, green light and different excitation observation body autofluorescence; confocal laser scanning microscope respectively by 405nm, 458nm, 476nm, 488nm, 514nm. 543nm, 633nm and other 7 kinds of excitation male oral sucker of spontaneous fluorescence analysis of lambda scanning.
Results: the different excitation light of Schistosoma japonicum from physical autofluorescence in different colors, with yellow green fluorescence effect of blue light excitation best body generally clear structure; scanning lambda respectively by 405nm, 488nm, 514nm and 543nm excitation emission wavelength sensitive samples, respectively 490-510nm, 550-570nm, 560-580nm and 590-600nm, in which 488nm and 514nm better results.458nm, 476nm, 633nm excitation effect is poor.
Conclusion: Observation of Schistosoma japonicum adult spontaneous fluorescence under fluorescence microscope, the best model is based on the observation of blue light yellow green fluorescence; Schistosoma japonicum by autofluorescence spectrum is wide, mainly distributed in the 500-600nm; 488nm and 514nm as the excitation wavelength is more appropriate.

【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R285;R392

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