The Role And Mechanism Of TLR4/MyD88/Nf-?B Signaling Pathway In Diarrheapredominant Irritable Bowel Syndrome

Objective Observe the acute and chronic stress method to construct the IBS-D rat model,analyze the changes of inflammatory factors,and clarify the role of TLR4/MyD88/NF-?B signaling pathway in IBS-D,and provide a new idea of the diagnosis and treatment for IBS-D.Methods Thirty 28-day-old SPF-class Wistar rats,females,averaged 70-90 g,were conditioned for one week after purchase.They were divided into 3 groups by random number method,10 rats were used as the blank group,10 rats were used as the IBS-D group,and 10 rats were used as the TAK-242 group.The IBS-D model was constructed by the acute and chronic stress method.The TAK-242 group was intraperitoneally injected with TAK-242(3 mg/kg)before the acute and chronic stress method,and the IBS-D group was given the same amount of normal saline.Do not do anything for the blank group.Observe the daily situation of the three groups of rats during the modeling process,three groups of rats were weighed and the fecal traits were evaluated by Bristol classification at 0,7,14,21,28 days.Assessment of intestinal pain sensitivity using AWR score,and observe the intestinal mucosa has no edema,erosion,ulcer,bleeding.The levels of MyD88,IL-1? and IL-6 in the serum of the three groups were detected by Elisa method.The expression levels of TLR4 and NF-?B in the colon tissues of the three groups were detected by Western blot and do HE staining to observe inflammatory infiltration on the 28 th day.Results(1)Before the experiment,each rat was active,with normal coat color,normaldiet and drinking water,and an elliptical bowel movement,which was consistent with Types 3,4,and 5 in Bristol stool type.With the increase of experimental stimulation,rats in IBS-D group and TAK-242 group were prone to irritability,low spirits,yellowish hair color,and the diet and drinking water were lower than those in the blank group.Excretion increased and did not form during stimulation,in line with Types 6 and 7 of Bristol's stool type,indicating successful modeling.In this process,no deaths occurred in each group of animals;(2)With the progress of modeling,the body weight of the three groups of rats gradually increased.However,compared with the blank group,the weight gain of the IBS-D group and the TAK-242 group was slow.At the same time,the weight of the IBS-D group was slower than that of the TAK-242 group.On the 7th,14 th,21st and 28 th days after the model was started,the difference in body weight between the three groups was statistically significant(P < 0.05);(3)With the progress of modeling,the Bristol stool typing scores of the IBS-D group and the Bristol stool stool typing of the TAK-242 group gradually increased.On the 0th day after the model was started,there was no significant difference in the scores of Bristol stools between the three groups(P>0.05).On the 7th,14 th,21st and 28 th days,there was a statistically significant difference in the scores of Bristol stools between the three groups(P<0.05);(4)With the progress of modeling,the pain threshold of rats in IBS-D group and the pain threshold of rats in TAK-242 group gradually decreased.On the day 0 after the model was started,there was no significant difference in pain threshold between the three groups(P>0.05).On the 7th,14 th,21st and 28 th days,the pain threshold of the three groups of rats was statistically significant(P<0.05);(5)With the progress of modeling,the maximum tolerated pressure threshold of rats in IBS-D group and the maximum tolerated pressure threshold of rats in TAK-242 group gradually decreased.On the 0th and 7th days after the model was started,there was no significant difference in the maximum tolerated pressure threshold between the three groups(P>0.05).On the14 th,21st,and 28 th days,the maximum tolerated pressure threshold of the three groupsof rats was statistically significant(P<0.05);(6)With the progress of modeling,the serum levels of MyD88,IL-1?and IL-6 in the IBS-D group and the serum levels of MyB88,IL-1?,IL-6 in the TAK-242 groups gradually increased.On the day 0 after the model was started,there was no significant difference in the expression levels of serum MyD88,IL-1?and IL-6 between the three groups(P>0.05).On the 7th,14 th,21st and28 th days,the expression levels of serum MyD88,IL-1?and IL-6 in the three groups of rats were significantly different(P<0.05);(7)Increased expression of TLR4 and NF-?B in colon tissue of rats in IBS-D group,and decreased NF-?B expression level after TLR4 inhibition by TAK-242.There was a statistically significant difference in the ratio of TLR4 and NF-?B gray values between the three groups(P<0.05);(8)The colonic mucosa of the three groups of rats was observed without erosion,ulcer and bleeding.The levels of colon tissue in the three groups were observed clearly,and there are mucosal layer,mucosal muscle layer,submucosa and muscle layer were observed.A small amount of inflammatory cells infiltrated in the mucosa of rats in the IBS-D group and the TAK-242 group,and the submucosal edema and the gap increased.Conclusions IBS-D may lead to the imbalance of inflammatory factors through the TLR4/MyD88/NF-? B signaling pathway,which may promote the development of IBS-D.