The Effects And Mechanisms Of ?-arrestin2 On Mouse Colitis And Epithelial Barrier Dysfunction

Background and ObjectionUlcerative colitis(UC)is a long-term condition that results in inflammation and ulcers of the co lon and rectum.The primary symptom of active disease is abdominal pain and diarrhea mixed with blood.Weight loss,fever and anemia may also occur.Symptoms typically occur intermittently with periods of no symptoms between flares.Complications may include megacolon,colon cancer,inflammation of the eyes and joints.The causes and mechanisms of UC are still unknown.Upon their discovery,?-arrestinl and 2 were named for their capacity to sterically hinder the G protein coupled receptors(GPCRs).Recent studies show that ?-arrestins can also serve as multiprotein scaffolds and active signaling cascades independently of G protein activation.They participate in various processes including proliferation,differentiation and apoptosis.?-arrestins modulate a plethora of cell signaling pathways in response to a variety of ligands in both immune and non-immune cells.Therefore,it is no surprising that ?-arrestins play important role in pathogenesis of autoimmune diseases such as inflammatory bowel diseases.Intestinal mucosal barrier dysfunction has been implicated in inflammatory bowel diseases.Genome-wide association studies(GWAS)have been successful in IBD and analyses of the genes and genetic loci implicated in IBD show several pathways that are crucial for intestinal homeostasis include barrier function and epithelial restitution.Neverthless,there is a growing body of evidence that implicates increased intestinal permeability as an etiologic factor of IBD pathogenesis.The intestinal epithelium is the foremost component of the intestinal mucosal barrier.It consists of the layer of epithelial cells lining the intestine.Sealing of the space between adjacent cells is mediated by junctional complexes formed by protein connections elaborated by each individual cell.And tight junction is the most important part in cell junctions.Although more proteins are present,the major types of tight junctions are the claudins and the occludins.These proteins associate with different peripheral membrane proteins such as zona occluden-1(ZO-1)located on the intracellular side of plasma membrane,which anchor the strands to the actin component of the cytoskeleton.Recently attention of ?-arrestin has focused on its role in the immune mechanisms,epithelial cell apoptosis and repair in IBD.However,there is little research on intestinal permeability and tight junction proteins.Moreover,there are arguments about wether ?-arrestins promote or inhibit the colitis.In this case,we establish DSS-induced acute colitis in mice firstly and investigate the influence of?-arrestin2 on colits.For further exploration,we illustrate the mechanisms from the perspective of intestinal permeability and tight junction proteins.MethodsDextran sodium sulfate(DSS)is used to induce intestinal colitis in mice.The experiment groups are designed as wild type control(WT),wild type colitis(WT+DSS)and knockout colitis(KO+DSS).The expression of ?-arrestin2 gene at mRNA and protein level is compared between WT group and WT+DSS group.The difference of weight loss,Disease Activity Index(DAI),spleen weight,colon length,histological score,intestinal permeability and important tight junction proteins(occludin,claudinl and ZO-1)are detected in WT+DSS and KO+DSS group.All data are expressed as the x±S x The differences in values between two groups were determined using Student's t-test.To compare multiple groups,one-way ANOVA was used.A difference was considered statistically significant when P<0.05.Results1.?-arrestin2 is involved in mouse colitis.Compared with WT group,the mRNA expression of ?-arrestin2 is significantly higher in the colon of WT+DSS group(1.006±0.066 VS 2.103±0.438,P=0.048).The protein level of ?-arrestin2 is elevated,too.2.?-arrestin2 deficiency protects mice from DSS-induced colitis.Compared with WT+DSS group,KO+DSS group has a slower weight loss in Day5,Day6 and Day7(98.262±0.870 VS 94.524±1.253,P=0.045;94.167±0.766 VS 89.458±1.462,P=0.029;89.523±2.301 VS 81.094±1.402,P=0.020),lower DAI in Day5,Day6 and Day7(1.585±0.210 VS 2.468±0.200,P=0.019;2.168±0.397 VS 3.20±0.200,P=0.042;2.918±0.158 VS 3.934±0.066,P<0.001),smaller spleen(74.950±2.430 VS 127.180±9.801,P=0.002),longer colon length(6.425±0.217 VS 5.320±0.227,P=0.011)and lower histological score(2.333±0.211 VS 3.667±0.211,P=0.002).3.p-arrestin2 results in weak epithelial barrier dysfunction.Lower permeability(7.835±0.520 VS 39.696±8.357,P=0.009)and higher protein level of claudinl and occluding are found in KO+DSS group.There is not significant difference in ZO-1 between the two groups.Conclusions?-arrestin2 may promote mouse colitis through the impairment of epithelial tight junctions and intestinal barrier function.