基于TLR3/NF-κB/MLCK通路研究蒲公英多糖改善虎龙斑肠道紧密连接损伤的分子机制

肠道紧密连接损伤是诱发鱼类肠道疾病的重要原因。研究表明TLRs/NF-κB/MLCK信号通路在肠道紧密连接调控中发挥重要作用。我们前期研究发现，投喂高脂饲料导致虎龙斑肠道的TLR3、NF-κB和MLCK基因表达量显著增加，肠道紧密连接损伤严重；而在高脂饲料中添加蒲公英多糖能显著降低上述基因的表达水平，改善肠道紧密连接损伤，但是其相关的作用机制尚不清楚。因此，本项目拟采用肠道上皮细胞培养、荧光定量PCR、Western blot和高通量测序等技术，探讨TLR3/NF-κB/MLCK通路活化对虎龙斑肠道上皮细胞紧密连接的影响，解析蒲公英多糖调控TLR3/NF-κB/MLCK通路影响肠道紧密连接的分子网络路径，阐明蒲公英多糖改善虎龙斑肠道紧密连接损伤的作用机制。本研究不仅可为防治鱼类肠道屏障损伤提供参考资料，还可为蒲公英多糖作为功能性饲料添加剂在水产养殖中的应用提供重要的理论依据。

The injury of intestinal tight junction is one of a important cause of intestinal diseases in fish. Studys have found TLRs/NF-κB/MLCK signaling pathway plays an important role in the regulation of intestinal tight junction. Our previous study found that the expressions of TLR3, NF-κB and MLCK genes significantly increased and the intestinal tight junctions were seriously damaged in the intestinal tract of hybrid grouper (Epinephelus lanceolatus♂ × Epinephelus fuscoguttatus♀) which feeding high fat diet. However, dandelion polysaccharide supplementation in high fat diets can significantly reduce the expression levels of these genes and improve intestinal tight junction damage. Nonetheless the relevant mechanism is still unclear. Therefore, this project intends to explore TLR3/NF-κB/MLCK pathway activation of dragon tiger spot tied to the intestinal epithelial cells and the effect of parsing the dandelion polysaccharide regulation TLR3/NF-κB/MLCK pathways affect intestinal tightly coupled molecular network path, clarify the dandelion polysaccharide improve fish bowel is closely connected to the action mechanism of the injury through the intestinal epithelial cell culture, fluorescent quantitative PCR, Western blot and high-throughput sequencing technologies. This study can not only provide reference for the prevention and control of intestinal barrier damage in fish, but also provide important theoretical basis for the application of dandelion polysaccharide as a functional feed additive in aquaculture.