本文選題：旋毛蟲 + Toll樣受體　； 參考：《吉林大學》2017年碩士論文

【摘要】：旋毛蟲病對人和動物健康、社會、經濟產生重大影響的食源性人獸共患寄生蟲病,該病是由寄生于哺乳動物橫紋肌細胞內寄生線蟲旋毛蟲引起的。在我國及全世界廣泛分布,給動物養(yǎng)殖業(yè)造成嚴重的經濟損失,對社會公共衛(wèi)生產生嚴重威脅。旋毛蟲在長期進化過程中形成了通過抑制宿主免疫反應而逃避宿主免疫系統攻擊的生存機制,使其得以順利侵襲與寄生,使宿主處于旋毛蟲感染的危險境地。這種免疫逃避使旋毛蟲免疫預防失效。免疫逃避已成為當前旋毛蟲有效免疫預防制劑研制的一個難以逾越的瓶頸。Toll樣受體(TLRs)在眾多的宿主免疫防御系統扮演了重要角色,TLRs激動劑作為抗旋毛蟲免疫逃避制劑具有無法估量的發(fā)展前景。本研究前期通過灌胃感染旋毛蟲,利用實時熒光定量PCR對旋毛蟲感染的不同階段的TLRs表達量進行檢測,同時ELISA法檢測Th1/Th2相關細胞因子白細胞介素-2(IL-2)、白細胞介素-4(IL-4)、白細胞介素-10(IL-10)、干擾素-γ(INF-γ)表達。結果表明:旋毛蟲感染期間,旋毛蟲以及其分泌產物能夠不同程度的調節(jié)宿主的免疫應答,在腸期階段TH1反應占優(yōu)勢,隨后新生幼蟲在骨骼肌期間TH2反應占優(yōu)勢。在感染初期,TLR1、TLR3、TLR4、TLR7和TLR9表達上調,新生幼蟲階段,幾乎所有的TLRs表達均下調,這表明新生幼蟲能夠抑制大多數TLRs表達和信號傳導。對宿主免疫應答具有廣泛的抑制作用。旋毛蟲能夠調節(jié)TLRs的表達以及其信號傳導通路。不同感染階段的TLRs表達對相關的信號傳導通路調控也不同,說明Toll樣受體在旋毛蟲感染期間發(fā)揮重要作用,旋毛蟲感染能夠誘導條件性Th1、Th2反應,調節(jié)T淋巴細胞和細胞因子產生,通過調節(jié)Toll樣受體表達來調控宿主對寄生蟲的免疫反應�；谏鲜鲅芯�,本研究用已報道的TLRs激動劑poly(I:C)、LPS-EB Biotin、TL8-506和OND M362對小鼠進行尾靜脈注射,干預旋毛蟲感染免疫逃避。感染后0、3、6、9、12、15、18、21、28、35天后剖殺小鼠,以人工胃液消化法收集肌幼蟲并計數,計算減蟲率。檢測TLRs激動劑對旋毛蟲感染階段脾TLRs表達情況,分析血液中Th1/Th2細胞因子,綜合評價4種TLRs激動劑對旋毛蟲感染的干預效果。結果表明:感染旋毛蟲的小鼠注射TLRs激動劑后,TLRs激動劑能夠激活TLRs,在通過激動劑激活后Th1相關的細胞因子IL-2、IFN-γ水平升高,而Th2相關的細胞因子IL-4、IL-10降低,在感染的35d剖殺小鼠測旋毛蟲的數量,計算減蟲率,注射了TLRs激動劑后,肌幼蟲的數量均有所下降,說明poly(I:C)、LPS-EB Biotin、TL8-506和OND M362均能夠干預旋毛蟲的感染,其中TLR3激動劑poly(I:C)的小鼠減蟲率達到最高,效果最佳。本實驗綜合評價4種TLRs激動劑對旋毛蟲感染的干預效果,篩選出TLR3激動劑poly(I:C)阻礙旋毛蟲免疫逃避的最佳TLRs激動劑,用于旋毛蟲免疫逃避干預治療,為TLRs激動劑應用于旋毛病的防控奠定基礎。本研究不僅對動物和人旋毛蟲病防治疫苗研發(fā)意義重大,對其他寄生蟲疫苗的研發(fā)將有不可估量的推動作用。
[Abstract]:Trichinosis has a great influence on human and animal health, society and economy. It is caused by the parasitic parasitic Trichinella parasitized in the mammalian rhabdomyid cells. It is widely distributed in China and the world, causing severe economic losses to the animal breeding industry and serious social public health. In the course of long-term evolution, Trichinella spiralis has formed a survival mechanism by inhibiting host immune response and escaping the host immune system attack, making it successful in invasion and parasitism, causing the host to be in the danger of Trichinella infection. This immune escape makes Trichinella Trichinella immune prevention failure. Immune evasion has become a current Trichinella. An insurmountable bottleneck.Toll like receptor (TLRs) has played an important role in many host immune defense systems. The TLRs agonist as an anti Trichinella immune escape agent has an immeasurable prospect. In this study, Trichinella spiralis was infected by gavage of Trichinella spiralis and real time fluorescence quantitative PCR was used for Trichinella spiralis TLRs expression at different stages of infection was detected, while ELISA assay was used to detect interleukin -2 (IL-2), interleukin -4 (IL-4), interleukin -10 (IL-10), and interferon - gamma (INF- gamma). The results showed that Trichinella spiralis and its secretory products could regulate the host in varying degrees during the period of Trichinella infection. The immune response, TH1 reaction predominant during the intestinal stage, and then the TH2 reaction of the newborn larvae during the skeletal muscle. In the early stage of infection, the expression of TLR1, TLR3, TLR4, TLR7 and TLR9 is up regulated, and almost all TLRs expressions are down regulated in the newborn larvae stage, which indicates that the newborn larvae can inhibit most TLRs expression and signal transduction. The answer has extensive inhibition. Trichinella can regulate the expression of TLRs and its signal transduction pathway. The regulation of TLRs expression in different stages of infection is also different, indicating that Toll like receptors play an important role during Trichinella infection. Trichinella infection can induce conditioned Th1, Th2 reaction, and T lymphatic regulation. Cells and cytokines produced by regulating the expression of Toll like receptors to regulate the host's immune response to parasites. Based on the above study, the present study used the reported TLRs agonists, poly (I:C), LPS-EB Biotin, TL8-506 and OND M362 to injecting the tail vein in mice to interfere with the immune evasion of the worm infection. The 0,3,6,9,12,15,18,21,28,35 days after infection. The muscle larvae were collected and counted by artificial gastric juice digestion and the rate of worm reduction was counted. The expression of TLRs agonists on the spleen TLRs expression in the stage of Trichinella Trichinella infection, the analysis of Th1/Th2 cytokine in the blood and the intervention effect of 4 TLRs agonists on Trichinella Trichinella infection were evaluated. The results showed that the mice infected with Trichinella Trichinella were injected with TLRs agonist. After that, TLRs activator activates TLRs and increases the level of Th1 related cytokines IL-2 and IFN- gamma after activation of the agonist, while Th2 related cytokines IL-4 and IL-10 decrease. The number of Trichinella spiralis in infected 35d Caesarean mice is measured and the rate of worm reduction is calculated. After the TLRs agonist is injected, the number of muscle larvae decreases, indicating poly (I:C). B Biotin, TL8-506 and OND M362 can all interfere with Trichinella Trichinella infection, of which the TLR3 agonist poly (I:C) has the highest worm reduction rate and the best effect. This experiment comprehensively evaluated the effect of 4 TLRs agonists on Trichinella Trichinella infection, and screened the best agonist for TLR3 agonist poly (I:C) immune escape from Trichinella Trichinella, used in spin hair. Insect immune avoidance intervention is the basis for the application of TLRs agonists to the prevention and control of circumflex disease. This study is not only of great significance to the research and development of animal and human Trichinella vaccine, but also has an inestimable role in the research and development of other parasitic vaccines.
【學位授予單位】：吉林大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：S852.7

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