Ageratina Adenophora Destroys The Intestinal Mechanical Barrier And Function Of Goats Through The MLCK/ROCK Signaling Pathway

Ageratina adenophora is a toxic invasive alien plant.A large number of studies have shown that animals ingesting Ageratina adenophora will have different degrees of toxic reactions,which are characterized by a series of pathological changes characterized by inflammatory damage to the liver,spleen,kidneys,and other parenchymal organs.The intestine is the organ of digestion and absorption of Ageratina adenophora after feeding,and is the first to be exposed to its toxic components,but the effects of Ageratina adenophora on the intestine of animals with compound stomach animals have not been reported.In order to explore whether and how Ageratina adenophora can cause intestinal injury in goats.In this experiment,Chengdu Ma Goat was used as the experimental animal,and 40% of the diet forage was replaced with Ageratina adenophora powder.After feeding for 90 days,samples of each intestinal segment were collected.We examined the intestinal histopathological damage,changes in the expression of each protein of tight junctions in the intestinal mechanical barrier,the expression of key signaling molecules of the tight junction regulatory pathway MLCK/ROCK signaling pathway,and changes in the expression of inflammatory factors.It provides a basic study to explore the toxicological mechanism of Ageratina adenophora.The main results of this study are as follows:(1)The damage to the small intestine was mainly characterized by loss of the apical structure of the intestinal villi,hemorrhage,edema,and inflammatory cell infiltration;the height of the intestinal villi decreased,the depth of the crypt increased and the ratio of villi to crypt increased in all intestinal segments of the small intestine;while the damage to all intestinal segments of the large intestine was less severe,mainly characterized by edema and a small amount of inflammatory cell infiltration(2)The intestinal damage markers DAO and D-LA were significantly increased(p <0.05)after 60 d of feeding Ageratina adenophora,and their concentrations increased gradually with the increase of feeding time in a time-dependent manner;the culture results of intestinal translocated bacteria were positive,i.e.,intestinal translocated bacteria were present in the liver,mesenteric lymph nodes and portal blood.The concentration was the highest in the mesenteric lymph nodes,followed by the liver,and the lowest in the portal vein blood.(3)By detecting the expression of inflammatory factors in each intestinal segment of the small intestine,it was found that Ageratina adenophora caused significant or highly significant up-regulation(p < 0.01 or p < 0.05)of pro-inflammatory factors(TNF-α,IFN-γ,IL-1β,IL-6)and significant or highly significant down-regulation of anti-inflammatory factors(IL-4,IL-10,IL-13)in each intestinal segment of the experimental group,and intestinal immune function disorders(4)By detecting changes in mRNA and protein expression of goat ileal tissue tight junction proteins(Occludin,Claudin-1,ZO-1)and signaling molecules(MLCK,MYPT-1,p-MYPT1,MLC2,p-MLC2,ROCK1,ROCK2)in the MLCK/ROCK signaling pathway.We found that Ageratina adenophora caused highly significant down-regulation(p < 0.01)in the expression of Occludin,Claudin-1,and ZO-1 at both gene and protein levels in goats;the MLCK/ROCK signaling pathway was activated in the experimental group,with significant or highly significant changes(p < 0.01 or p < 0.05)in mRNA and protein expression of each signaling molecule.In summary,when goats were poisoned with Ageratina adenophora,the intestinal structure was damaged,and the most serious damage was found in each intestinal segment of the small intestine;the intestinal barrier function was damaged and permeability was increased,the contents of intestinal damage markers DAO and D-LA were increased,and intestinal bacteria were translocated to other organs;the intestinal inflammatory damage occurred,the expression of pro-inflammatory factors increased and the expression of antiinflammatory factors decreased,and the intestinal immunity was disrupted;the intestinal mechanical barrier was destroyed.The MLCK/ ROCK signaling pathway was activated,resulting in decreased expression of the tight junction proteins Occludin,Claudin-1,and ZO-1.The intestinal mechanical barrier was disrupted.