Molecular Mechanism Of Gallic Acid Regulating The Intestinal Inflammation In Weaned Piglets

Piglets are susceptible to immune stress caused by external influences such as weaning,which induces to some health problems such as intestinal inflammation as well as diarrhea.Previous studies observed that plant-derived bioactive substances could improve the intestinal health of livestock and poultry by regulating immune signaling pathways.However,there is limited information concerning the protective effects of gallic acid（GA）on the intestinal inflammatory response in weaned piglets.Thus,this study aims to investigate the effects of gallic acid on intestinal immune function in piglets by both in vivo and in vitro experiments.Our study includes the following four parts:part 1,a total of 96 male weaned piglets（25±1 days of age）with an initial body weight of 8.40 kg were allocated into 4experimental treatments with 6 replicated pens（4 piglets per pen）each,which focused on the effects of dietary gallic acid（GA）at 0,100,200,or 400 mg/kg on growth performance and diarrhea incidence;part 2,a total of 24 piglets（1 piglet/pen）received an intraperitoneal lipopolysaccharide（LPS）injection on day 21,and the piglets were slaughtered at 4 h postchallenge to evaluate the effects of different level of dietary gallic acid supplementation on intestinal morphology,plasma antioxidant indices and immune response in weaned piglets;part 3,the IPEC-J2 cells were exposed to H₂O₂ or LPS in the presence of gallic acid to explore the protective effect of gallic acid on injured cells;part 4,the specific pathway inhibitor of NF-κB signaling bay11-7082 was used to inhibit NF-κB signaling pathway,in order to demonstrate the key target of regulating the immune function and barrier function of IPEC-J2 cells by gallic acid.The results showed that dietary supplementation of gallic acid had no significant effect on the growth performance of piglets as well as the antioxidant status before and after LPS challenge（P>0.05）.However,dietary GA with 400 mg/kg significantly reduced the diarrhea incidence of piglets,and dietary supplementation with GA improved the intestinal morphology of piglets by increasing villus height,reducing crypt depth and increasing the ratio of villus height to crypt depth（P<0.05）,in which the effect on the jejunum was better than on the ileum.In addition,dietary GA can also regulate the immune function of challenged piglets by increasing the secretion of sIgA in the intestinal mucosa and reducing the expression of pro-inflammatory cytokines（P<0.05）.In the in vitro experiments,gallic acid can significantly reduce the expression of pro-inflammatory mediators and the level of inflammatory enzyme COX-2 by inhibition the NF-κB-IκBα-IκKα/βactivation（P<0.05）,and there was a tendency to increase the activity of catalase and upregulate its mRNA level in the cells.At the same time,GA increased the barrier function of IPEC-J2 cells by up-regulating the expression of tight junction proteins ZO-1 and Occludin.Our study demonstrated the effect of gallic acid on reducing the diarrhea incidence by regulating the intestinal immune function of weaned piglets,and uncovered the molecular mechanism of GA regularing the intestinal inflammatory response via inhibiting the NF-κB signaling pathway,which could provide a new strategy for the application of gallic acid in weaned piglets.