Study On The Regulation Mechanism Of Sanbai Tang On TLR4/NF-κB Signaling Pathway In Ulcerative Colitis Rats

Objective: To study the mechanism of clinically effective prescription and empirical prescription Sanbai Tang through the TLR4/NF-κB signaling pathway to regulate the intestinal inflammation of ulcerative colitis rats induced by acetic acid.Methods:1.Randomly divide 60 SD rats into normal group,model group,Sanbaitang low,medium and high dose groups and positive control group,10 rats in each group.In addition to the normal group,the remaining 5 groups used 8% acetic acid to stimulate the colon to establish a rat model of ulcerative colitis.Three days after modeling,drug intervention was given,and the normal group and the model group were given double-distilled water;the Sanbaitang low,medium,and high-dose groups were given Sanbaitang 9.4g crude drug/kg/day,18.8g crude drug/kg/day,37.6g Crude drug/kg/day;the positive control group was given sulfasalazine 0.18g/kg/dayfor 14 days.2.Calculate the DAI score of the rat based on the rat’s weight loss percentage score,stool trait score,gross blood stool or occult blood test score.3.Using HE staining method to prepare pathological sections of rat colon tissue and observe the pathological changes.4.Using enzyme-linked immunosorbent assay(ELISA)to determine the expression of IL-1β,IL-6,IL-8 and other inflammatory cytokines and the content of anti-inflammatory cytokines IL-4 and IL-10 in the serum of rats in each group.5.Western blotting(WB)was used to detect the expression of TLR2,TLR4 and NF-κBp65 related proteins in TLR4/NF-κB signaling pathway in the colon tissues of various groups of rats.Results:1.Compared with the normal group,the DAI score and spleen coefficient percentage of the model group were significantly increased(P<0.01),body weight was significantly reduced(P<0.01),colon length was significantly shortened(P<0.01),and colonic tissue pathological sections There was erosion,intestinal wall adhesion,intestinal thickening,disappearance of fold texture,submucosal tissue necrosis or loss of mucosa,and a large number of inflammatory cell infiltrations were seen,including neutropenia and loss of glands;compared with the model group,The DAI scores of the low,medium and high dose groups of Sanbai Tang and the positive control group decreased significantly(P<0.01),the colon length of the high dose group of Sanbai Tang and the positive control group increased significantly(P<0.01),and the weight of each treatment group increased significantly(P<0.05 or P<0.01),the spleen coefficient of Sanbai Tang’s low and high dose groups and the positive control group was significantly reduced(P<0.05 or P<0.01),rats in each treatment group showed varying degrees of ulcer healing,erosion and bowel Decreased wall adhesions,submucosal tissue necrosis and mucosal loss were reduced,and the degree of inflammatory cell infiltration was reduced.Among them,the high-dose group of Sanbai Tang and the positive control group showed the most obvious improvement,followed by the medium-dose group of Sanbai Tang,and finally Decoction low dose group.2.Compared with the normal group,inflammatory cytokines such as IL-1β,IL-6,IL-8 and anti-inflammatory cytokines such as IL-4 and IL-10 in the model group increased significantly(P<0.01);Compared with the model group,the inflammatorycytokines IL-1β,IL-6 and IL-8 in the serum of Sanbai Tang,high-dose group and positive control group were significantly reduced(P<0.05 or P<0.01),while Sanbai Tang was low and medium,High-dose group and positive control group anti-inflammatory cytokines IL-4 and IL-10 were significantly reduced(P<0.01);inflammatory cytokines IL-1β,IL-6,IL-8 and anti-inflammatory cells in rat serum The content of factors IL-4 and IL-10 is inversely proportional to the dose of Sanbai Tang.3.Compared with the normal group,the TLR2,TLR4 and NF-κBp65 protein in the colon tissue of the model group increased significantly(P<0.01);compared with the model group,the TLR2 in the colon tissue of the Sanbai Tang,high-dose group and positive control group Significantly reduced protein expression(P<0.01);TLR4protein expression in colon tissue of Sanbai Tang high-dose group and positive control group was significantly reduced(P<0.05 or P<0.01);Sanbai Tang low-,medium-,and high-dose group and positive control group The expression of NF-κBp65 protein in colon tissue was significantly decreased(P<0.01);the expression of TLR2,TLR4 and NF-κBp65 protein in rat colon tissue was inversely proportional to the dose of Sanbai Tang.Conclusion: 1.Sanbai Tang has a certain therapeutic effect on acetic acid-induced UC rats,which can significantly improve the symptoms of loose stools and blood in the UC rats,and reduce or even heal ulcers.2.The anti-ulcerative colitis mechanism of Sanbai Tang may be related to inhibiting the over-activation of TLR4/NF-κB signaling pathway,thereby down-regulating the expression of TLR2,TLR4 and NF-κBP65 protein in the colon tissue of UC rats and reducing the phosphorylation of NF-κBp65 Inflammation reduces the release of inflammatory cytokines such as IL-1β,IL-6 and IL-8,thereby reducing the intestinal inflammatory infiltration and inflammatory response.3.The degree of Sanbai Tang inhibiting the TLR4/NF-κB signaling pathway,reducing the release of intestinal inflammatory mediators and reducing the inflammatory response are related to the dosage of Sanbai Tang,with the highest dose of Sanbai Tang being the best.