一种高效提高新生仔猪肠道免疫力的新机制

The intestinal immunity of piglet is closely related with the growth and development of intestinal mucosal and the mature of intestinal innate immune. Vitamin A greatly influences the growth and development of intestinal epithelial cells. Dendritic cells (DCs) located in small intestine not only play an important role in intestinal innate immunity, but also involve in the transformation of vitamin A. Bacillus subtilis have the ability of promoting the intestine development, improving the immunity of intestinal system and protecting the intestine against infection. Therefore, firstly we will construct recombinant bi-function Bacillus subtilis (synthesizing carotene in Bacillus subtilis and displaying the peptide which can target to the swine intestinal DCs on the surface of Bacillus subtilis). Then we will demonstrate the intake ability of swine DCs to swallow the recombinant Bacillus subtilis in vitro (cell co-cultural system). After oral immunization with the recombinant Bacillus subtilis in newborn piglets, we will try to investigate whether the expression of carotene in small intestine and the antigen-presenting ability of DCs are improved significantly, at the same time the local immunity of small intestine and the ability of resisting infection in small intestine are also promoted. The immunity of newborn piglet intestinal will be improved in the following points, 'vitamins, immune and antagonistic', just like killing three birds with one stone. The result of our study will provide an effective strategy to promote piglet intestinal immunity and develop a convenient way for safe, efficient and economy prevention of piglet diarrhea.

仔猪肠道免疫功能与小肠黏膜的生长发育和肠道先天免疫的成熟密切相关。肠道上皮生长发育受维生素A（VA）的影响较大。小肠黏膜下树突状细胞（DCs）不仅在肠道的先天免疫中发挥着重要作用，还参与了VA的转化。枯草芽孢杆菌具有促进小肠发育、提高肠道免疫力和拮抗病原微生物感染等多种功能。因此，本项目首先构建重组的双功能枯草芽孢杆菌（菌体内可以合成胡萝卜素，菌体表面展示靶向猪肠道DCs多肽），然后应用体外试验证实猪DCs摄取构建的重组枯草芽孢杆菌的能力（细胞共同培养体系）。最后应用重组枯草芽孢杆菌口服免疫新生仔猪（体内试验），证实小肠中胡萝卜素的表达量、DCs的摄取能力和抗原呈递能力、小肠局部免疫力以及抵抗病原微生物感染的能力是否得到显著改善。本研究从"维生素、免疫和拮抗"三个方面提高新生仔猪肠道免疫力，发挥"一箭三雕"的目的，以一种全新的机制和方便简易的途径，为绿色、安全、高效预防仔猪腹泻。

仔猪肠道黏膜免疫系统不健全是导致流行性腹泻等病毒感染的重要原因。促进肠道黏膜免疫系统的发育，提高黏膜免疫力有助于降低病原感染，提高生产性能。本研究构建类胡萝卜素合成酶表达质粒并电击转化枯草芽孢杆菌，获得能够产生类胡萝卜素的重组枯草芽孢杆菌。体外试验表明，重组枯草芽孢杆菌合成的类胡萝卜素能够通过CD36和NF-κB信号诱导树突状细胞成熟并促进Th1细胞分化。体内试验表明，饲喂重组枯草芽孢杆菌能够促进肠上皮细胞产生CCL20，进而提高小鼠肠道固有层中树突状细胞的数量。此外，固有层中Th17和Treg细胞数量也增加。攻毒试验表明，饲喂重组枯草芽孢杆菌能够提高小鼠对沙门氏菌的抵抗力。最后，通过仔猪试验我们发现，饲喂重组枯草芽孢杆菌能够显著促进仔猪肠道黏膜免疫系统的发育。