磷壁酸在植物乳杆菌与银鲳肠粘膜细胞互作中的作用探索

Teichoic acid (LTA) is one of the major components of cell wall of Gram-positive bacteria, which plays important roles in adhesion, colonization and immune regulation of probiotics. However, many reports indicate that depletion of LTA in Lactobacillus plantarum results in enhanced probiotic efficacy. It is speculated that the teichoic acid of Lactobacillus plantarum has unknown characteristics. Our previous studies have revealed that Lactobacillus plantarum can improve the growth performance and immunologic function of silvery pomfret juveniles. Based on previous studies, this project will study on the effects of LTA on the interactions between Lactobacillus plantarum and silvery pomfret intestinal epithelial cells. A GFP (green fluorescent protein)-tagged mutant of Lactobacillus plantarum deficient in the LTA will be constructed by homologous recombination, and the LTA will be isolated and purified by DEAE-52 chromatography in this study. The capacity of adhesion, colonization and immune regulation of mutant of Lactobacillus plantarum deficient in the LTA will be investigated, and the effects of LTA on the adhesion of Lactobacillus plantarum will be determined, and the immunological characteristics of LTA will also be identified through a series of experiments in vitro and in vivo. This project can not only explore the functional properties of LTA in depth, but also open up a new perspective for the study of mechanism of probiotics.

LTA（磷壁酸）是革兰氏阳性菌细胞壁的重要组分，在益生菌的粘附定植、免疫调控过程中发挥着重要作用。然而许多文献指出，敲除LTA基因的植物乳杆菌具有了更有效的益生作用。我们推测植物乳杆菌的LTA具有未探明的特性。前期研究发现，植物乳杆菌能够显著改善银鲳幼鱼的生长性能和免疫功能。本项拟在此基础上，重点针对LTA在植物乳杆菌与银鲳肠道粘膜互作中的作用展开探索，采用基因重组技术构建GFP（绿色荧光蛋白）标记的植物乳杆菌LTA突变株，运用DEAE-52层析法分离、纯化植物乳杆菌的LTA，通过开展体内、体外实验，分析植物乳杆菌LTA突变株的粘附定植、免疫调控能力，测定LTA对植物乳杆菌粘附定植的影响，进而对LTA的免疫特性进行鉴定。本项不仅可以深入揭示植物乳杆菌LTA的功能特性，同时也为益生菌的机理研究开辟了新的视角。

本项目将银鲳肠道黏膜细胞作为体外模型系统分析了植物乳杆菌LTA的免疫学特性，主要研究内容包括：植物乳杆菌LTA可以抑制鳗弧菌（致病菌）的黏附，并能够缓解由鳗弧菌诱导的炎症反应和细胞凋亡；克隆了银鲳TLR2、TLR5和TLR9的cDNA序列，并发现植物乳杆菌LTA可以特异性的刺激TLR2的表达；与鳗弧菌鞭毛蛋白和嗜水气单胞菌脂多糖相比，植物乳杆菌LTA诱导了最轻微的细胞凋亡，这可能与植物乳杆菌的益生特性有关。研究结果初步阐释了植物乳杆菌LTA的益生机理，并可为益生菌的机理探讨和推广应用提供了数据参考和理论指导。