德氏乳杆菌对哺乳仔猪肠道树突状细胞成熟影响及其TLR4信号转导通路研究

The major function of the Lactobacillus on the animals is to enhance the immune response in the gastrointestinal tract of the animals, thus the investigation of the underlying mechanisms on the effects of Lactobacillus on the muscoal immune function may shed light on the development of Lactobacillus-based probiotic and its good application. This study aimed at evaluating the effect of oral administration of Lactobacillus delbrueckii on the distribution, production and maturation of the dendritic cell(DC), the levels of mRNA expression and protein contents of some cytokines in the intestinal of the suckling piglets using immunohistochemical techniques and real time PCR techniques，analyzing the immune function of intestinal DC co-culture with CD4 + T cells in vitro. Therefore reveals the regulatory role of the Lact. delbrueckii on the suckling piglets intestinal DC development. By investigating the Lactobacillus delbrueckii and cellular components to regulate the intestinal DC maturation differences in vitro, reveal the material basis of Lactobacillus delbrueckii which play its immunomodulatory roles. By evaluating the mRNA expression and protein contents of DC surface receptor TLR4 and their major downstream signal transtraction pathway components, the signal transtraction pathways by which Lact. delbrueckii regulate DC maturation are revealed. Therefore, the main objective of this study is to reveal the immnuomodulatory mechanisms of the Lactobacillus delbrueckii through cell and molecular level and form the theoretical basis for the development of Lact. delbrueckii-based probiotic and its application in the livestock production.

乳酸菌具有增强动物黏膜免疫反应的重要作用，探明乳酸菌促进黏膜免疫反应的机理是开发理想乳酸菌制剂及合理应用的关键。本研究应用免疫组织化学技术、实时荧光定量PCR技术考察哺乳仔猪灌服德氏乳杆菌菌液对哺乳仔猪肠道树突状细胞的成熟以及肠道粘膜组织中相关细胞因子mRNA表达影响，分析肠道DC与CD4+T细胞体外共培后CD4+T细胞的免疫功能变化，揭示德氏乳杆菌对哺乳仔猪肠道DC免疫功能的调控作用。通过德氏乳杆菌及细胞组分调节体外培养仔猪肠道DC成熟的差异性，考察DC TLR4受体的mRNA与蛋白的表达，分析TLRs下游信号通路重要元件的TRIF、NF-κB、IκB、IRF-3mRNA的表达量及蛋白水平，揭示德氏乳杆菌及活性组分调控DC成熟的信号转导通路。本研究旨在细胞水平和分子水平上探讨德氏乳杆菌免疫调控机理，为乳酸菌制剂的开发和在畜牧生产中应用提供理论依据。

乳酸菌具有增强动物黏膜免疫反应的重要作用，探明乳酸菌促进黏膜免疫反应的机理是开发理想乳酸菌制剂及合理应用的关键。本研究旨在细胞水平和分子水平上探讨德氏乳杆菌肠道免疫调控机理，为乳酸菌制剂的开发和在仔猪生产中应用提供理论依据。.试验采用德氏乳杆菌活菌、热灭活菌体及耗尽培养上清液灌服小鼠，分析血清免疫指标、空肠粘膜细胞因子和防御素等基因的表达，探讨德氏乳杆菌对小鼠肠道先天性免疫的调节作用；采用德氏乳杆菌菌液灌服新生哺乳仔猪，分析仔猪血清、肠道黏膜中细胞因子水平、肠道组织中免疫细胞的数量、树突状细胞（DC）相关趋化因子及其趋化因子受体、成熟度表面分子、TLRs及其下游信号分子表达量，探讨德氏乳杆菌对哺乳仔猪免疫调节作用的影响，研究德氏乳杆菌调控DC成熟及其信号转导通路；应用德氏乳杆菌活菌和热灭活德氏乳杆菌刺激体外培养骨髓源树突状细胞，检测DC细胞表型分子、上清培养液中细胞因子和趋化因子，研究德氏乳杆菌活菌体及热灭活菌体调控免疫调节作用及差异性；应用Western blotting对树突状细胞表面识别受体TLRs、胞内下游信号传导分子及核因子蛋白表达量进行分析，探讨德氏乳杆菌对DC表面受体TLRs及其下游信号分子的调控作用，揭示德氏乳杆菌调控DC成熟的信号转导通路。. 研究表明，德氏乳杆菌可以通过调节哺乳仔猪肠道DC来介导肠道粘膜免疫调节作用，改善哺乳仔猪肠道黏膜免疫功能，提高哺乳仔猪的生产性能。德氏乳杆菌能调节树突状细胞的成熟和免疫功能，可通过TLRs介导信号通路调节DC的成熟，信号传导途径为MyD88依赖性途径，活菌和热灭活菌体对树突状细胞的成熟、功能及信号转导途径存在一定差异。