Effects And Mechanisms Of Tauroursodeoxycholic Acid Combined With Salicylazosulfapyridine Against Colitis Model Of Mice

Background and ObjectiveUlcerative colitis(UC)as a kind of inflammatory bowel diseases,the lesion is mainly limited to colonic mucosa and submucosa,showing many immunological disease characteristics.The main clinical manifestations of ulcerative colitis include abdominal pain,diarrhea and purulent stools,recurrent attacks and relapses,Patients with UC do have an increased risk of colorectal cancers.The pathogenesis of UC have not yet been elucidated,but most researcher think that it is related to genetic,inflammation,immune,and environment factors.The treatment drugs include aminosalicylic acid,glucocorticoid,immunosuppressants agents and biological agents.However,due to these drugs adverse reactions,the clinical application is limited.Tauroursodeoxycholic acid(TUDCA)is a natural combination of bile acid,he main component is ursodeoxycholic acid,it can be used in treatment of primary biliary cirrhosis and primary sclerosing cholangitis.In recent years,tauroursodeoxycholic acid have showed stronger cytoprotection and anti-inflammatory effects to ursodeoxycholic acid,whether in vivo or in vitro.Many scholars recommended it can be a new treatment choice for UC.Caudal-related homeobox transcription factor 2(Cdx2)is a basic nuclear transcription factor that regulates the differentiation and proliferation of intestinal epithelial cells.Cdx2 mainly exists in the crypt villi of the small intestine and colon,especially in the differentiated intestinal epithelial cells.Notch signaling pathway is a highly conservative conduction system affecting the cell fate through the interaction between adjacent cells,is mainly composed of Notch receptors、corresponding ligands(DSA protein)and CSL DNA binding protein,it almost exists in all kinds of animals,playing a important role in the regulation of cell differentiation,proliferation,apoptosis and a series of pathological and physiological process,Hairy enhancerof split 1(Hes1)is the key target gene of Notch signaling pathway downstream.The abnormal expression of key genes in Notch signal transduction pathway are closely related with liver disease,cancer,neurological diseases,congenital dysplasia and inflammatory bowel disease diseases.Dextran sulfate sodiam(DSS)is a kind of polysaccharide sulfate synthesized from sucrose,it has been matured application in the preparation of colitis model in mice.The Study found that DSS induced colitis is particularly similar to human UC in both clinical and histopathological manifestations,and has been widely recognized as a model in the study of UC.The main purpose of this study is to compare the efficacy of TUDCA and SASP intervention in the treatment of colitis model,to understand whether the combined drug has a synergistic effect,and to explore the possible mechanism of the TUDCA intervention model.Methods1、 To establish the experimental colitis on mice:50 male Balb/c mice were randomly divided into five groups,10 mice in each group:normal group,model group,SASP(500mg/kg)group,TUDCA(100mg/kg)group,TUDCA/SASP combination group.Normal control group was fed with distilled water,the rest groups were fed with 5%DSS for 7 days.Each treatment group was orally given certain agent one day before DSS was given.2、The general condition and the DAI score in mice:observed the general state,recorded changes of body weight,loose stools,blood in the stool everyday to assess disease activity index(DAI).3、Data detection of inflammatory cytokine:The concentration of TNF-a and IL-6 was measured by ELISA kit After collecting blood samples through eye socket venous,absorbance value was measured in the enzyme standard instrument,the concentration values calculated by the absorption value substituted into standard curve equation.4、Histopathological observation:Measured colon length of mice after were sacrificed,cut off part of the colon were fixed,sectioned and stained with hematoxylin and eosin to mirror check,according to the inflammation,lesion depth,crypt damage and the extent of the lesion to evaluate the degree of histological damage.5、Data detection of colon tissue associated protein:Hes1,Atoh1 and Cdx2 expression were detected by immunohistochemistry SP,and the computer software image analysis system was used to analyze the associated protein expression.Results1 、 TUDCA 、 SASP 、 TUDCA/SASP group can significantly alleviate the clinical symptoms of experimental colitis in mice,reduce the score of DAI,ease the shortening of colon length,Among them,the TUDCA/SASP group works best,the TUDCA group and SASP group had a quite curative effect;Histopathology showed that the drug intervention group can obviously reduce inflammatory cell infiltration,glandular and crypt destruction,goblet cells decreased on the colitis model,the TUDCA/SASP group was the least inflammatory damage,There were no statistical difference on the pathological damage score between TUDCA and SASP group.2 、 The ELISA detection showed that TUDCA,SASP and TUDCA/SASP group could inhibit the protein expression of serum TNF-α and IL-6 on colitis mice,and the TUDCA/SASP group had the strongest inhibitory effect.Immunohistochemistry detection of Hes1,Atoh1 and Cdx2 in colonic tissues showed that TUDCA could increase the expression of Atoh1 and Cdx2,and inhibit the expression of Hes1 in colonic tissues.ConclusionTUDCA、SASP、TUDCA/SASP group could reduce DSS induced mice colitis severity significantly,TUDCA/SASP preventive effect may be better than TUDCA and SASP,TUDCA and SASP showed similar effects in the course of intervention mice colitis from either clinical or histopathologic score,the two drugs efficacy without obvious difference.The mechanism ofTUDCA intervention in mice with colitis may be related to inhibiting the expression of inflammatory cytokines,reducing the activation of Hes1 protein and increasing the expression of Cdx2 protein in colonic tissues.