基于p38 MAPK信号通路研究浒苔多糖对仔猪肠道炎症损伤的调控作用及机制

The intestinal mucosal barrier injury is one of the most important factors that affect growth rate and health status of weaned piglets. It has been demonstrated that Enteromorpha polysaccharide (EP) is related to scavenging free radicals, restraining bacteria growth and inhibiting Inflammation. Furthermore, our previous study also showed that dietary supplementation with EP could improve the antioxidant capacity and intestine development of weaned piglets, However, the molecular mechanism of EP in regulating the intestinal barrier dysfunction mediated by p38 MAPK pathway remains unclear. Based on the scientific hypothesis that, "EP regulates intestinal barrier dysfunction of piglets through p38 MAPK pathway", this research would be conducted with animal feeding study and enterocytes (IPEC-J2) cell culture model to clarify the molecular mechanism that EP regulates the intestinal epithelial cell injury in piglets mediated by p38 MAPK signaling pathway through determining the integrity of intestinal mucosal barrier, secretion of inflammatory cytokines and the variation of p38 MAPK signaling pathway in the intestine by cellular respiratory capacity monitoring, flow cytometry, immunofluorescence, TUNEL and western-blot when piglets or IPEC-J2 challenge with LPS. Taken together, this project is supposed to reveal the mechanism of EP in regulating the intestinal injury mediated by p38 MAPK signaling pathway, then provides theoretical basis for the supplementation with EP as feed additive in the livestock husbandry.

肠道炎症导致的黏膜屏障功能损伤是影响断奶仔猪生长及群体健康水平的重要因素之一。浒苔多糖有抗菌、清除自由基、抑制炎症反应等多种生理功能。我们的研究也发现，浒苔多糖替代饲用抗生素可缓解断奶仔猪氧化应激，促进小肠发育等。但是，目前关于浒苔多糖对仔猪肠道健康的影响还未见公开报道；应激/炎症关键通路p38 MAPK在其调控仔猪肠道屏障功能中扮演什么角色尚不清楚，迫切需要进一步研究。因此，基于“浒苔多糖通过p38 MAPK信号通路调节仔猪肠道黏膜屏障损伤”的科学假设，本项目拟以LPS诱导的断奶仔猪和猪肠上皮细胞为研究模型，通过免疫荧光、细胞呼吸代谢、流式细胞术、TUNEL、Western blot等方法研究浒苔多糖调控仔猪肠上皮细胞损伤的作用，阐明浒苔多糖通过p38 MAPK通路调控仔猪肠道黏膜屏障损伤的作用机制。拟开展工作将为浒苔多糖在生猪养殖中的应用提供科学依据。

仔猪在养殖过程中，容易受到环境、饲料等因素的影响，使仔猪肠道屏障受损，引起炎症，导致仔猪生长缓慢，甚至死亡。本研究中通过体内试验：LPS仔猪炎症造模、DSS和LPS小鼠炎症造模，以及体外实验：敌草快诱导IPEC-J2细胞损伤试验，应用ELISA、透射电镜、免疫荧光、免疫组化、蛋白印迹、RT-qPCR等实验技术，考察仔猪及小鼠血清的炎症相关指标、肠道形态以及细胞凋亡情况，并分析肠道紧密连接蛋白的表达与分布、肠道通透性以及IPEC-J2细胞的抗氧化相关指标，Western blot和RT-qPCR评价p38 MAPK及其相关因子的表达，考察浒苔多糖浒苔多糖对肠道屏障损伤的保护作用及可能的机理，为浒苔多糖及其衍生物在生猪养殖中的应用提供参考。