| Objective: Microsurgical technique is used to establish the rat total colorectomy ileum pouch anal anastomosis(IPAA)model,respectively by trinitrobenzene sulfonic acid(TNBS),and dextran sodium sulfate(DSS)induced rats ileum pouchitis,compare two methods of inducing the rat pouchitis model,the characteristics of the two models can be evaluated.Methods: Microsurgical technique is used to establish IPAA male SD rats model.The model of rats were randomly divided into 5 groups: namely IPAA +DSS group,IPAA postoperative day 31,give the rats 4% DSS solution free drink for 4 days;IPAA + TNBS group,postoperative day 31,TNBS is instilled by a suitable medical-grade polyurethane catheter for enteral feeding approximately 3.5 cm proximal to anal verge,established the model used a dose of 200 mg/kg,TNBS dissolved in 50% ethanol per time;IPAA + NS group,postoperative day 31,IPAA for saline enema per time;IPAA for no intervention group,namely IPAA group,postoperative without any intervention;Sham group were as control group,only the abdominal operation.During the experiment,after the enema of pouch angiography examination,rats were observed the change of the contrast agents in the pouch and explore the position of enema in rats and the enema tube insertion length.During the experiment,we observed and recorded each group general situation of rats,mental state,weight change,bloody situation,fecal character;Feces direct smear method was used to detect the distribution of the flora;On the day 35 th,the pouch tissue of all the groups except Sham group were taken,resreachers took the rat proximal small intestine,middle of small intestine and distal small intestine and the pouch,only in the Sham group,the proximal small intestine of rats were taken,small intestine middle and distal small intestine tissue,using HE staining method for detection and histopathological grading;Take immune histochemical method testing each group intestinal mucosa which is closely connected to the expression of the protein occuldin level.Results: 1.The ileal pouch imaging results showed that in order to be fully contact with ileal pouch mucosa by the contrast agents,left lateral position to give enema was better,and determined tube inserted into the anus depth of 3.5 cm.3.In general,the IPAA + DSS and IPAA + TNBS group of rats showed different degrees of lazy,tired,hair dry,dull,but the rats IPAA + NS group,the IPAA group and Sham group activities agile,bright hair;the weight of IPAA group rats gradually decreased after the operation,the weight of the Sham group was steady growth;we began to intervene at 31 th,after the intervention,the weight of IPAA + DSS group appeared to decline at(58.83±5.88)g,IPAA + TNBS group also continued to decline at(28.67 ± 5.57)g,however,the IPAA + TNBS group rats weight drop less than IPAA + DSS group,the difference was statistically significant(P < 0.05),IPAA + NS group,IPAA group and Sham group rats weight were steady increase.4.The situation of blood and feces,IPAA + DSS group of rats were bloody stool around the anal(6/6),IPAA + TNBS group of rats are loose,some of the rats in mucus purulent blood(3/6),normal stool were observed in IPAA + NS,IPAA + NI and Sham group.Before the intervention,fecal score comparison among the groups were no statistically significant difference(P > 0.05),IPAA + DSS group and IPAA + TNBS group rats after the intervention,fecal score(1.50 ±0.55),(1.67±0.82)are lower than IPAA+NS group,the IPAA group and Sham group(4.01±0.89),(4.33±0.82),(4.03±0.78),the difference was statistically significant(F = 16.96,P < 0.01),compare the IPAA+DSS group with IPAA+TNBS in fecal score,there was no statistically significant difference(P > 0.05).5.The fecal flora distribution showed that the total number of bacteria in the IPAA+DSS group,was decreased slightly,gram positive bacteria was significantly reduced than the normal range and gram negative bacilli was increased,gram positive cocci and gram negative cocci were in the normal range;the total number of bacteria in IPAA+TNBS group was decreased slightly,gram positive bacilli decreased significantly compared with normal,leather the gram negative bacilli was significantly increased compared to the normal range,gram positive cocci and negative cocci were in the normal range;the fecal flora distribution of IPAA+NS,IPAA+NI group were in the normal range.6.The pouch and small intestine tissue pathology were taken,after the intervention,IPAA+DSS and IPAA+TNBS group pouch histologic grading were higher than IPAA,IPAA+NS group,Sham group(F =53.8,P < 0.05),and the IPAA+TNBS pouch score higher than the IPAA + DSS group,the difference was statistically significant(P < 0.05),the IPAA+NS group,the IPAA group and Sham group rats was no statistically significant difference(P > 0.05);proximal,middle and small intestine pathological and histological grading of distant IPAA+DSS group is higher than the IPAA+TNBS,IPAA +NS group,the IPAA group and Sham group(F = 76.156,P < 0.05),IPAA+TNBS and IPAA+NS group,the IPAA group and Sham group of rats was no statistically significant difference(P>0.05).7.The immunohistochemical results showed that the ileum pouch tissue was closely connected to the expression levels of the protein occludin,intestinal barrier IPAA+DSS and IPAA +TNBS group were lower than IPAA+NS group,the IPAA group and Sham group,the difference was statistically significant(F = 57.8,P < 0.05),and the IPAA + TNBS was lower than IPAA + DSS group(P < 0.05),IPAA + NS group,the IPAA group and Sham group rats,there was no statistically significant difference(P>0.05).Occludin protein expression levels in small intestine proximal,middle and distal were as followed,IPAA + DSS group is lower than the IPAA+TNBS,IPAA+NS group,the IPAA group and Sham group(F = 20.55,P<0.05),IPAA +TNBS and IPAA+NS group,the IPAA group and Sham group,there was no statistically significant difference(P>0.05).Conclusions: 1.Using microsurgical technique,IPAA model of rats were successfully established;2.The use of DSS and TNBS can induce pouchitis rats model;3.The mount of bacteria induced by TNBS was lower than that induced by DSS,and the change of bacillus aureus was more severe than that induced by DSS;the damage of pathological histology induced by TNBS was more severe than that induced by the DSS,but the damage of pathological histology in small intestine tissue induced by DSS was more severe than that induced by the TNBS;the intestinal barrier damage in pouch tissue in rats induced by TNBS was severe than DSS;4.On the basis of rats IPAA model,TNBS can establish more stable and reliable ileum pouchitis model in rats than DSS. |
PDF Full Text Request