| Objectives 1.To research the effect of experimental colitis induced by different concentrations of trinitrobenzene sulfonic acid(TNBS)in rats?According to the number of surviving rat,DAI sores,CMDI sores and TDI sores,we can choose the optimum concentration of TNBS and appropriate time of drug intervention?2.To investigate the therapeutic effect of thalidomide used in the early phase of the disease on TNBS-induced experimental colitis in rats,and compare the therapeutic effect with infliximab and 5-aminosalicylic acid,then to explore its possible mechanisms.Methods 1.The twenty-four virgin male Sprague-Dawley rats were randomly divided into three groups:normal control group(group?,4),TNBS 100mg/kg group(group?,10)and TNBS 150mg/kg group(group?,10)?Then rats in group?were processed by rectal administration of 0.9ml normal saline?And experimental colitis was induced respectively in rats of group ?and group ? by rectal administration of 100mg/kg and 150mg/kg TNBS dissolved in anhydrous ethanol?In group?,a rat was sacrificed at the days of 1,3,7 and 14 after induction of colitis?In group?and group?,two rats were sacrificed at the days of 1,3 and 7 after induction of colitis,and the rest of rats were sacrificed at the days of 14 after induction of colitis?Then the natural death number of rats was recorded?The DAI sores,CMDI sores and TDI sores were evaluated?According to the consequence,we can choose the optimum concentration of TNBS and appropriate time of drug intervention?2.The fifty virgin male Sprague-Dawley rats were randomly divided into five groups:normal control group,model group,thalidomide group,infliximab group,5-aminosalicylic acid group?Then rats in normal control group were processed by rectal administration of 0.9ml normal saline?And experimental colitis was induced in rats of the last four groups by rectal administration of 5%TNBS(100mg/kg)dissolved in anhydrous ethanol(V:V=2:1)?From the first day after induction of colitis,rats in normal control group and model group were given normal saline(4ml/kg/d)by gavage,and rats in thalidomide group,infliximab group,5-aminosalicylic acid group were given thalidomide(200ml/kg/d)by gavage and infliximab(5ml/kg/d)by intraperitoneal injection and 5-aminosalicylic acid(100ml/kg/d/)by gavage?After fourteen days,all the rats were sacrificed?The DAI sores?CMDI sores and TDI sores were evaluated?The expression of TNF-? and NF-?Bp65 in the colonic tissues were detected by immunohistochemistry?The content of TNF-? in the serum were measured by Enzyme linked immunosorbent analysis(ELISA)?Results 1.Both of 100mg/kg and 150mg/kg TNBS dissolved in anhydrous ethanol can induce the experimental colitis model in rats successfully?One and three rats in TNBS 100mg/kg group and TNBS 150mg/kg group had died of natural causes respectively?And compared with TNBS 150mg/kg group,the severity of symptoms,gross pathology and histopathology in TNBS 100mg/kg group were slighter,and the scores of DAI,CMDI,TDI were lower[3.00 VS 3.34,6.00 VS 7.50,5.50 VS 7.50(rats executed in the first day);3.17 VS 3.67,7.50 VS 8.50,7.50 VS 8.50(rats executed in the third day);2.67 VS 3.17,6.50 VS 7.50,6.00 VS 7.00(rats executed in the seventh day);2.44 VS 2.67,5.33 VS 6.00,5.67 VS 7.00(rats executed in the fourteenth day)]? 2.Compared with normal control group,the DAI sores?CMDI sores and TDI sores in model group were increased significantly[2.46±0.87 VS 0.07±0.14,5.38±1.19 VS 0.20±0.42,5.88±1.25 VS 0.10±0.32,P<0.05];Then compared with model group,the DAI scores,CMDI scores and TDI scores in thalidomide group,infliximab group and 5-aminosalicylic acid group were reduced significantly[0.83±0.72?0.67±0.78?1.22±0.78 VS 2.46±0.87,2.30±1.16?2.11±1.27?3.56±1.33 VS 5.38±1.19,2.70±0.95?2.44±0.88?3.89±1.05 VS 5.88±1.25,P<0.05]?From the therapeutic effect,the DAI scores,CMDI scores and TDI scores in 5-aminosalicylic acid group were higher than those in thalidomide group and infliximab group?Among these scores,CMDI scores and TDI scores were different significantly[0.83±0.72?0.67±0.78 VS 1.22±0.78,P>0.05;2.30±1.16?2.11±1.27 VS 3.56±1.33,2.70±0.95?2.44±0.88 VS 3.89±1.05,P<0.05];And compared with infliximab group,the DAI scores,CMDI scores and TDI scores in thalidomide group were higher?But there were not significant difference between two groups[0.83±0.72 VS 0.67±0.78,2.30±1.16 VS 2.11±1.27,2.70±0.95 VS 2.44±0.88,P>0.05]?It's indicated that the effect of thalidomide and infliximab are better than 5-aminosalicylic acid?And the effect of thalidomide is similar to infliximab?Compared with normal control group,the expressions of TNF-? and NF-?Bp65 in colonic mucosa in model group were increased significantly[9.75±1.28 VS 0.80±0.92,8.63±1.41 VS 0.50±0.71,P<0.05];Then compared with model group,the expressions of TNF-? and NF-?Bp65 in colonic mucosa in thalidomide group,infliximab group and 5-aminosalicylic acid group were decreased significantly[3.80±1.23?3.22±1.09?5.22±1.39 VS 9.75±1.28,2.60±1.27?2.33±1.23?4.22±1.64 VS 8.63±1.41,P<0.05];Compared with 5-aminosalicylic acid group,those in thalidomide group and infliximab group were decreased significantly[3.80±1.23?3.22±1.09 VS 5.22±1.39,2.60±1.27?2.33±1.23 VS 4.22±1.64 P<0.05];Compared with infliximab group,the expressions of TNF-? and NF-?Bp65 in colonic mucosa in thalidomide group were higher,but there were not significant difference between two groups [ 3.80±1.23 VS 3.22±1.09,2.60±1.27 VS 2.33±1.23,P>0.05]?In the tissue sections,NF-?Bp65 was positive mainly in the nucleus in the model group,NF-?Bp65 was positive mainly in the cytoplasm in the other four groups?Compared with normal control group,the concentration of TNF-? in the serum in model group were increased significantly[291.65±42.71 VS 6.08±0.47,P<0.05];Then the concentration of TNF-? in the serum in thalidomide group,infliximab group and 5-aminosalicylic acid group were decreased significantly than those in model group[34.56±3.07?22.19±2.78?61.60±5.10 VS 291.65±42.71,P<0.05];Compared with 5-aminosalicylic acid group,the concentration of TNF-? in the serum in thalidomide group and infliximab group were decreased significantly[34.56±3.07?22.19±2.78 VS 61.60±5.10,P<0.05];Compared with infliximab group,the concentration of TNF-? in the serum in thalidomide group were higher,but there were not significant difference between two groups[34.56±3.07 VS 22.19±2.78,P>0.05]?The expressions of NF-?Bp65 in colonic mucosa was positively correlated with the expressions of TNF-? in colonic mucosa(r=0.876,P<0.05)?And it also was positively correlated with the concentration of TNF-? in the serum(r=0.848,P<0.05)?Conclusions 1.Both of the experimental colitis model in rats induced by 100mg/kg TNBS and 150mg/kg TNBS dissolved in anhydrous ethanol are similar with Crohn's disease?But according to the number of surviving rat,DAI sores,CMDI sores and TDI sores,the optimum concentration of TNBS is 100mg/kg?It conform to the requirements of the experiment?And the fastigium of DAI sores,CMDI sores and TDI sores is the third day after induction of colitis,so the first day after induction of colitis is the appropriate time of drug intervention?2.Thalidomide has the therapeutic effect on TNBS-induced colitis in rats?Blocking the path of NF-?B and decreasing the level of TNF-? may be one of its main mechanisms?... |
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