Base On The TGF-β1/P38 Pathway To Explore The Pathogenesis Of Ileum Injury Induced By Chronic Restraint Stress In Rat

Chronic stress interferes with homeostasis,intake,weight and production performance of animals.Chronic stress can induce a variety of digestive tract diseases such as functional abdominal pain syndrome and peptic ulcer.Chronic stress has the most significant impact on the intestinal tract,such as inflammatory bowel disease,which not only has a high fatality rate,but also greatly affects the economic value of livestock.But,the specific mechanism of chronic stress-induced intestinal injury is still unknown.In order to explore the mechanism of TGF-β1 in chronic stress-damaged intestinal barrier,the chronic stress model was established to examining the ileum tissue pathology,intestinal structure and the changes of the ultrastructure of intestinal epithelial cells.preliminary validation chronic stress damage of the ileum,and detected TGF-β1 signaling pathways related protein expression.40 healthy male Wistar rats were randomly divided into control group（group C）of 8 rats and a chronic stress group（group CS）of 32 rats.The CS group was further divided into the group with no immediate detection for 14 days(S₁₄),the group with immediate detection for 14 days(S_14L),the group with no immediate detection for 21 days(S₂₁)and the group with immediate detection for 21days(S_21L).After 14 days of modeling in the S₁₄and S_14L groups,21 days after modeling in the S₂₁and S_21L groups(the S₁₄ and S₂₁ groups were tested on the second day after the end of modeling,open field experiments were conducted and blood were collected.Then the rats were sacrificed and the ileum tissues were taken for reserve.The success of the establishment of chronic stress model was verified by detecting the content of related hormones in the behavior and serum of rats.By detected the ileum tissue pathology,ileum ultrastructural organization structure,the expression of inflammation related protein,mitochondrial apoptosis related proteins expression,and the m RNA level and expression of TGF-β1,we explored the mechanism of TGF-β1 in chronic stress-induced intestinal barrier damage.The results of open field experiments show that compared with C group,the S₂₁ group has reduced trajectory and exercise level（p<0.01）,and the hormone content results show that compared with C group,the content of CORT and CRH in S₁₄ group and S₂₁ group were significantly reduced（p<0.01）.Based on the above results,it was proved that 21 days of modeling can cause chronic stress to rats,and in the following experiments,rats of 21 days of stress were set as the CS group.Histopathological results showed that the structure of the ileum in group C was clear,and no inflammatory edema were found.Mild edema of the ileal mucosal epithelial cells in CS group was seen with localized infiltration of small focal inflammatory cells.Intestinal mucosa villi were reduced and mild.Malformation,shedding of ileum villi and shortening.The ultrastructural observation of the ileum showed that the intestinal epithelial cells of C group were tightly arranged and the tight junction structure between the cells was clear;the surface of intestinal epithelial cells of CS group was loose with microvilli,the mitochondrial matrix was swollen,and the continuity of the tight junction structure between integrity was disrupted,and immunohistochemical and real-time quantitative PCR results showed that the expression of tight junction structural proteins and m RNA levels of Claudin3,ZO-1,and Occludin in the CS group were significantly reduced.Western blot results showed that the expression of IL-1β,IL-6,TNF-α,CLIC4,cleaved caspase-3,cleaved caspase-9,Cyt C,and Bax/Bcl-2 in the CS group increased significantly（p<0.01）.It shows that chronic stress-induced intestinal barrier damage is related to inflammation and apoptosis of mitochondrial pathway.Western blot results showed that the phosphorylation level of P38 in the CS group was significantly increased（p<0.01）,while the phosphorylation levels of smad2/3,ERK,and JNK were not significantly changed compared to the C group p>0.05),proving inflammatory factors Activation and activation of mitochondrial apoptotic pathway are related to the TGF-β1/P38pathway.The results of this experiment confirmed that chronic restraint for 21 days can cause chronic stress in rats.The TGF-β1/P38 pathway related with the damage of tight junction structure of intestinal epithelium induced by chronic stress through increasing inflammatory cytokines and epithelial cell apoptosis.The results of this test indicate that the TGF-β1/P38 pathway may be a key target for preventing and treating chronic ileal injury induced by chronic stress.