TLRs激动剂对小鼠不同生活时期旋毛虫生物学特性的影响

发布时间：2018-05-24 03:37

本文选题：旋毛虫 + Toll样受体　；参考：《吉林大学》2017年硕士论文

【摘要】：旋毛虫病对人和动物健康、社会、经济产生重大影响的食源性人兽共患寄生虫病,该病是由寄生于哺乳动物横纹肌细胞内寄生线虫旋毛虫引起的。在我国及全世界广泛分布,给动物养殖业造成严重的经济损失,对社会公共卫生产生严重威胁。旋毛虫在长期进化过程中形成了通过抑制宿主免疫反应而逃避宿主免疫系统攻击的生存机制,使其得以顺利侵袭与寄生,使宿主处于旋毛虫感染的危险境地。这种免疫逃避使旋毛虫免疫预防失效。免疫逃避已成为当前旋毛虫有效免疫预防制剂研制的一个难以逾越的瓶颈。Toll样受体(TLRs)在众多的宿主免疫防御系统扮演了重要角色,TLRs激动剂作为抗旋毛虫免疫逃避制剂具有无法估量的发展前景。本研究前期通过灌胃感染旋毛虫,利用实时荧光定量PCR对旋毛虫感染的不同阶段的TLRs表达量进行检测,同时ELISA法检测Th1/Th2相关细胞因子白细胞介素-2(IL-2)、白细胞介素-4(IL-4)、白细胞介素-10(IL-10)、干扰素-γ(INF-γ)表达。结果表明:旋毛虫感染期间,旋毛虫以及其分泌产物能够不同程度的调节宿主的免疫应答,在肠期阶段TH1反应占优势,随后新生幼虫在骨骼肌期间TH2反应占优势。在感染初期,TLR1、TLR3、TLR4、TLR7和TLR9表达上调,新生幼虫阶段,几乎所有的TLRs表达均下调,这表明新生幼虫能够抑制大多数TLRs表达和信号传导。对宿主免疫应答具有广泛的抑制作用。旋毛虫能够调节TLRs的表达以及其信号传导通路。不同感染阶段的TLRs表达对相关的信号传导通路调控也不同,说明Toll样受体在旋毛虫感染期间发挥重要作用,旋毛虫感染能够诱导条件性Th1、Th2反应,调节T淋巴细胞和细胞因子产生,通过调节Toll样受体表达来调控宿主对寄生虫的免疫反应。基于上述研究,本研究用已报道的TLRs激动剂poly(I:C)、LPS-EB Biotin、TL8-506和OND M362对小鼠进行尾静脉注射,干预旋毛虫感染免疫逃避。感染后0、3、6、9、12、15、18、21、28、35天后剖杀小鼠,以人工胃液消化法收集肌幼虫并计数,计算减虫率。检测TLRs激动剂对旋毛虫感染阶段脾TLRs表达情况,分析血液中Th1/Th2细胞因子,综合评价4种TLRs激动剂对旋毛虫感染的干预效果。结果表明:感染旋毛虫的小鼠注射TLRs激动剂后,TLRs激动剂能够激活TLRs,在通过激动剂激活后Th1相关的细胞因子IL-2、IFN-γ水平升高,而Th2相关的细胞因子IL-4、IL-10降低,在感染的35d剖杀小鼠测旋毛虫的数量,计算减虫率,注射了TLRs激动剂后,肌幼虫的数量均有所下降,说明poly(I:C)、LPS-EB Biotin、TL8-506和OND M362均能够干预旋毛虫的感染,其中TLR3激动剂poly(I:C)的小鼠减虫率达到最高,效果最佳。本实验综合评价4种TLRs激动剂对旋毛虫感染的干预效果,筛选出TLR3激动剂poly(I:C)阻碍旋毛虫免疫逃避的最佳TLRs激动剂,用于旋毛虫免疫逃避干预治疗,为TLRs激动剂应用于旋毛病的防控奠定基础。本研究不仅对动物和人旋毛虫病防治疫苗研发意义重大,对其他寄生虫疫苗的研发将有不可估量的推动作用。
[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.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S852.7

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