本文關鍵詞： 寄生蟲免疫 細粒棘球蚴 免疫因子 腸粘膜免疫屏障　出處：《新疆醫(yī)科大學》2016年博士論文　論文類型：學位論文

【摘要】：背景:由細粒棘球絳蟲的幼蟲引起的棘球蚴病,是一種人畜共患的寄生蟲病,高發(fā)于我國及世界各地的畜牧區(qū),分布地域相當廣闊,目前已成為全球性的公共衛(wèi)生問題。人類及部分食草類動物(如綿羊等)因為誤食了由其終末宿主排出的蟲卵成為其中間宿主,蟲卵經(jīng)過胃,最終在十二指腸及小腸上段腸粘膜內(nèi)寄居,在此蟲卵孵化出六鉤蚴,穿過腸粘膜屏障入血并到達肝臟及其他器官,對機體的健康危害極大。既往研究發(fā)現(xiàn),細粒棘球蚴之所以能夠在機體內(nèi)長期生存,是其通過影響宿主Th1及Th2兩型細胞免疫應答平衡因子,誘導非保護性的Th2型免疫應答成為主導,利于細粒棘球蚴在體內(nèi)的生長。但是,腸粘膜免疫屏障系統(tǒng)是機體內(nèi)最大的淋巴免疫系統(tǒng),是細粒棘球蚴進入機體的第一道屏障,其對腸粘膜免疫屏障影響的相關研究鮮有報道,本研究旨在通過自然感染細粒棘球蚴的綿羊做為大動物研究對象,采取隨機對照實驗,應用酶聯(lián)免疫吸附測定(ELISA)及免疫組化等方法,分析Th1/Th2相關的部分免疫因子(IL-6,IL-10及IFN-γ)在外周血和小腸粘膜組織內(nèi)的水平變化;同時,應用阿苯達唑質(zhì)脂體治療自然感染細粒棘球蚴感染的綿羊,進一步評價細粒棘球蚴感染后對外周血及腸粘膜內(nèi)Th1/Th2所介導的免疫反應的影響;此外,通過對肝細粒棘球蚴患者與正常人群中幽門螺桿菌感染率的進行病例對照研究,以期從腸道微生態(tài)角度來探討,腸道微生態(tài)變化對細粒棘球蚴影響腸粘膜免疫屏障功能可能起到的作用。最終來研究,自然感染細粒棘球蚴對腸道粘膜免疫屏障可能發(fā)生的影響,為今后預防和治療細粒棘球蚴感染提供可行性研究方向。方法:用B超初檢和CT復檢的方法,從新疆巴音布魯克草原986只綿羊中,篩選出年齡在24-36月左右,自然感染細粒棘球蚴的雌性綿羊20只,按隨機數(shù)字表法,分為兩組,分別為使用阿苯達唑治療組和細粒棘球蚴感染組,以及按同法篩選出未感染組綿羊10只,做為正常對照組。嚴格隨機三盲方法,對上述三組實驗對象進行治療及飼養(yǎng)3月后,進行外周血采集、處死后小腸粘膜標本收集。用ELISA法對外周血中IL-6,IL-10及IFN-γ等免疫因子水平進行測定,分析細粒棘球感染及阿苯達唑質(zhì)脂體治療后外周血中免疫因子的變化;同時通過對小腸粘膜石蠟切片進行免疫組化染色,比較IL-6,IL-10及IFN-γ等免疫因子的陽性表達水平;此外,通過隨機選取臨床確診感染細粒棘球蚴患者51例及正常健康體檢人群30例成人做為對照組,使用化學發(fā)光法檢測幽門螺桿菌感染情況,進一步探討,腸道微生態(tài)對細粒棘球蚴作用于腸粘膜免疫屏障過程中的可能影響。結(jié)果:1)ELISA法對自然感染細粒棘球蚴綿羊外周血免疫因子IL-6、IL-10、IFN-γ進行測定提示:感染組上述3種免疫因子水平均高于對照組,但只有IL-10具有統(tǒng)計學意義(P0.001);免疫組化法對自然感染細粒棘球蚴綿羊腸粘膜內(nèi)免疫因子IL-6、IL-10、IFN-γ進行測定提示:感染組僅有IFN-γ明顯高于對照組,并有統(tǒng)計學意義(P0.005);2)阿苯達唑脂質(zhì)體治療干預后,ELISA法測定外周血Th2相關免疫因子IL-6,IL-10均有降低,但僅有IL-10具有統(tǒng)計學意義(P0.01);阿苯達唑脂質(zhì)體治療后,腸粘膜免疫組化結(jié)果示:腸粘膜內(nèi)Th1相關免疫因子IFN-γ保持在較高水平,并具有統(tǒng)計學意義(P.01),腸粘膜內(nèi)Th2相關免疫因子IL-10有所降低,但沒有統(tǒng)計學意義(P=0.893);3)用化學發(fā)光法對51例細粒棘球感染患者進幽門螺桿菌抗體檢測發(fā)現(xiàn)陽性率達74.51%,而對照組為50.00%,兩者具有統(tǒng)計學差異(P=0.025)。結(jié)論:本研究利用自然感染細粒棘球蚴綿羊做為研究對象,更接近同樣做為細粒棘球感染中間宿主的人類感染模式。研究發(fā)現(xiàn),自然感染細粒棘球蚴綿羊外周血中免疫反應主要以Th2型為主,而腸粘膜免疫反應可能以Th1型為主;行阿苯達唑質(zhì)脂體進行干預治療后結(jié)果提示:阿苯達唑質(zhì)脂體可增強機體體液及腸粘膜免疫屏障的Th1型免疫反應,對Th2類免疫因子有抑制作用;在對人細粒棘球蚴感染與幽門螺桿菌感染相關性病例對照研究結(jié)果提示:腸道微生態(tài)的變化可能對細粒棘球蚴感染后,腸粘膜免疫屏障功能改變具有影響,其中幽門螺桿菌可能與細粒棘球蚴存在協(xié)同感染可能,為進一步研究細粒棘球蚴如何影響腸粘膜屏障提供了一個可能的研究方向。
[Abstract]:Background: caused by Echinococcus larval hydatid disease is a zoonotic parasitic disease, high incidence in our country and around the world in the area of animal husbandry, the geographical distribution is very wide and has become a global public health problem. Human beings and some herbivorous animal (such as sheep) because eating eggs the discharge from the end host as its intermediate host, the eggs of the stomach, duodenum and small intestine in the final sojourn in the intestinal mucosa, the eggs hatched six larvae, through the intestinal mucosal barrier in the blood and reach the liver and other organs, great harm to the health of the body. The previous research found that Echinococcus granulosus has been able to long-term live in the body, it is through the type two Th1 and Th2 affect the host cellular immune response balance factor induced Th2 immune response of non protection become dominant, in favor of Echinococcus multilocularis in life Long. However, the intestinal mucosal immune barrier system is the largest lymphoid immune system, is the first barrier of Echinococcus granulosus into the body, the related research is rarely reported in its effect on intestinal mucosal immune barrier, through the research of natural infection of Echinococcus granulosus sheep as the object of animal research, a randomized controlled experiment, application enzyme linked immunosorbent assay (ELISA) and immunohistochemistry method, analysis of some immune factors related to Th1/Th2 (IL-6, IL-10 and IFN- gamma) levels in peripheral blood and intestinal mucosal tissues; at the same time, the application of albendazole liposome in treatment of natural infection of Echinococcus granulosus infection in sheep, immune the reaction further evaluation of Echinococcus granulosus infection after peripheral blood and intestinal mucosa mediated by Th1/Th2; in addition, the rate of Helicobacter pylori infection in patients with hepatic hydatid cyst and normal people. A case-control study was conducted, in order from the perspective of the intestinal microflora, intestinal mucosal barrier function of intestinal microflora changes of Echinococcus granulosus may play a role. To study the final, natural infection of Echinococcus granulosus may occur on intestinal mucosal immune barrier impact for future prevention and treatment of Echinococcus granulosus infection in the direction of the feasibility study provide. Methods: a re examination of the ultrasound examination and CT at the beginning of 986 sheep from Xinjiang Bayinbuluke grassland, screened at the age of 24-36 months, natural infection of Echinococcus granulosus in female 20 sheep were randomly divided into two groups, respectively using albendazole treatment group and Echinococcus granulosus the infection group, and according to the same method to filter out non infection group of 10 sheep, as normal control group. Three strictly random blind method, the three groups were treated by feeding and after March. Collect peripheral blood, after the death of intestinal mucosa were collected. Using ELISA method in peripheral blood IL-6, immune factors on levels of IL-10 and IFN- gamma were measured to analysis the changes of immune factor in peripheral blood of Echinococcus infection and albendazole liposome after treatment; at the same time through the intestinal mucosa paraffin sections immunohistochemical staining, IL-6, IL-10 and IFN- gamma serum expression level; in addition, the randomly selected clinical diagnosis of Echinococcus granulosus infection in 51 cases of patients and 30 cases of adult normal healthy people as the control group. The further study, the detection of Helicobacter pylori infection using chemiluminescence method, may affect the intestinal micro ecology of Echinococcus granulosus effect on intestinal mucosal barrier in the process. Results: 1) ELISA method on natural infection of Echinococcus granulosus sheep peripheral blood immune factors IL-6, IL-10, IFN- were determined by gamma tip: infection group The above 3 kinds of immune factors were higher than those in control group, but only IL-10 was statistically significant (P0.001); IL-10 immunohistochemical method on natural infection of Echinococcus granulosus sheep gut mucosal immune factors IL-6, IFN-, gamma determination tip: IFN- gamma group only significantly higher than the control group, with statistical significance (P0.005); 2) liposomal albendazole treatment after intervention, determination of peripheral blood Th2 related immune factors IL-6 ELISA, IL-10 had decreased, but only IL-10 was statistically significant (P0.01); albendazole liposome in the treatment of intestinal mucosa after immunohistochemical results are shown: in the intestinal mucosa of Th1 related immune factors IFN- gamma keep at a high level, and has statistical significance (P.01), in the intestinal mucosa of Th2 related immune factor IL-10 decreased, but not statistically significant (P=0.893); 3) by chemiluminescence method in 51 cases of Echinococcus infection in patients with Helicobacter pylori antibody detection The positive rate was 74.51%, and 50% in the control group, both had statistical difference (P=0.025). Conclusion: This study using the natural infection of Echinococcus granulosus sheep as research object, to do the same pattern as the human infection of Echinococcus infection of intermediate host. The study found that the natural infection of Echinococcus granulosus sheep immunity in peripheral blood the reaction mainly Th2 type and intestinal mucosal immune response may be dominated by Th1; for albendazole liposome intervention after treatment showed that albendazole liposome can enhance the humoral immune response to Th1 and intestinal mucosal immune barrier, inhibits Th2 immune factor; on Echinococcus granulosus infection with Helicobacter pylori infection related case-control study results suggested that the changes of intestinal microflora of Echinococcus granulosus infection, intestinal mucosal barrier function has the effect of changing, Helicobacter pylori may have synergistic infection with Echinococcus granulosus, which provides a possible direction for further research on how Echinococcus granulosus affects intestinal mucosal barrier.

【學位授予單位】：新疆醫(yī)科大學
【學位級別】：博士
【學位授予年份】：2016
【分類號】：R532.32

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