The Clinical Meaningfulness And Expression Of MMP-7 And Tight Junction Proteins In Inflammatory Bowel Disease

BackgroundInflammatory bowel disease(IBD)is a chronic recurrent inflammatory disease which mainly involves the gastrointestinal tract.It includes these two types,ulcerative colitis(UC)and Crohn’s disease(CD).Current researchs on the etiology of IBD shows that inappropriate inflammation is tightly related to genetic susceptibility,environmental factors,and the intestinal mucosal-immune system.The intestinal barrier is an important mechanical and immune defense against toxins in vitro and vivo,extraneous substances such as viruses,bacteria and antigens from the food in the body.The loss of the function of intestinal epithelial barrier is an important factor in the development and Prolongation of the inflammation of IBD.When it is destroyed,innate immune cells will Powerfully response to suppress and clear extraneous substances.The intestinal tight junction(TJ)is one of the most crucial structures to maintain the integrity of the intestinal epithelial barrier,it includes zonula occludens(ZO),Claudin,occludin,etc.The interaction between tight junction proteins ensure the tight junctions more stable.Matrix metalloproteinase 7(MMP-7)is one of the most important enzymes of the family of matrix metalloproteinases.It plays a vital role in the wound healing and tissue remodeling,as an important mediator of tissue injury during inflammation and a potential target for treatment of IBD.Many studies have proved that the level of MMP-7 expression is up-regulated in IBD,but its relationship with the severity of IBD has not been studied clearly.ObjectiveEvaluate the locations and levels of MMP-7 and tight junctions proteins such as ZO-1,Claudin-1,occludin,as well as the relationship between MMP-7 and IBD,in the colonic tissues of the active IBD(mild,moderate,and severe),clinical remission IBD,and the controls in this experiment.The correlation between MMP-7 and clinical activity evaluation indexes like C-reactive protein and white blood cells,the above Tight junction proteins was analyzed,so as to understand whether MMP-7 can be used as an indicator of IBD inflammatory activity.Methods1.Collection of patients and biopsy specimens: A total of 38 cases of colon biopsy samples were collected from patients with IBD who were hospitalized in the Second Affiliated Hospital of Zhengzhou University from January 2019 to January2021,including 20 cases of ulcerative colitis and 18 cases of Crohn’s disease.The paracancerous tissues(more than 5 cm away from the carcinoma)of 5 patients with colon cancer were collected as the control group.The results of CRP and WBC from the first fasting blood sampling after admission were collected.The study was approved by the Ethics Committee of the Second Affiliated Hospital of Zhengzhou University,and informed consent was signed.2.Expression of immunohistochemical protein: All tissues were fixed in fixative,dehydrated,removed,paraffin embedding and pathological sectioning after electronic colonoscopy.Immunohistochemical staining was used to analyze the expression of MMP-7,ZO-1,Claudin-1 and occludin in colon tissues of patients with IBD and the control group.The images were observed,scanned,and captured on a biological microscope and scanner.All of the images were analyzed with Image-J and Image-Pro Plus 6.0.3.Statistical analysis: Graph Pad Prism 8.0.2 was used for statistical analysis,data processing and graphics making.Counting analysis data is described by mean ±standard deviation.Chi-square test was used for comparison between groups.After the normal test and homogeneity test,the comparison between two groups was performed by t test or the Mann-Whitney U test,and the comparison across multiple groups were performed with one-way ANOVA or Kruskal Wallis test.Pearson correlation test was used to analyze the correlation.Correlation analysis was performed using Pearson or Spearman test to analyze correlations between variables.For the above,The test standard α=0.05,the difference was regarded to be statistically significant when P<0.05.Results1.The relationship between the expression of MMP-7,tight junction proteins and the general situation of Patients.Among Patients with IBD,there were no statistically significant differences in the expression of MMP-7 and tight junction proteins like ZO-1,Claudin-1,and occludin among different genders,ages,and whether treated with anti-TNF-alpha biologic agents within 3 months.2.The Localization and expression of MMP-7 in colon tissuesIn paracancerous tissues and the tissues of patients with IBD in clinical remission,MMP-7 is only slightly expressed in the cytoplasm,and was less intensively stained,while in patients with active IBD,a large number of positive areas with light brown or dark brown staining can be seen,mainly concentrated in the epithelial cells surrounding the active inflammation,and a few inflammatory cells.Quantitative analysis showed that the expression of MMP-7 was up-regulated in both the active UC group and the severe CD group,compared with the control group,and the difference was statistically significant(P<0.05).As the severity increases,the expression of MMP-7 was gradually up-regulated.Compared with the respective remission group,the expression of severe UC group and CD group was significantly up-regulated(P<0.05).There was no significant difference between UC and CD patients of the same severity(P>0.05).3.The Localization and expression of ZO-1,Claudin-1 and occludin in colon tissuesZO-1,Claudin-1 and occludin are all mainly positively expressed on the cell membrane and a small amount of staining can also be seen in the cytoplasm of these cells.There were more brownish yellow staining areas in the control group and remission IBD,and the staining intensity was stronger,while the staining was lighter in the tissues of active IBD,among which severe UC and CD had the weakest staining intensity.Quantitative analysis showed that the expression of ZO-1,Claudin-1 and occludin in the active IBD showed a decreasing trend compared with the control group and remission IBD,and as the severity increases,the expression of these tight junction proteins was gradually down-regulated,especially in severe UC and CD(P<0.01).There was no significant difference in the expression of tight junction proteins between UC and CD of the same severity(P>0.05).4.Correlation analysis of MMP-7 and tight junction proteinsThe expression of MMP-7 in the tissues of patients with active IBD is negatively correlated with the expression of tight junction proteins such as ZO-1,Claudin-1,and occludin(P<0.001).5.Correlation analysis of MMP-7,Claudin-1,ZO-1,occludin and clinical dataThe expression of MMP-7 in the tissues of patients with active IBD is significantly positively correlated with clinical data such as CRP and WBC(P<0.001),while tight junction proteins such as ZO-1,Claudin-1,and occludin are significantly negatively correlated with the above indexes(P<0.001).Conclusions1.The expression of MMP-7,ZO-1,Claudin-1 and occludin was all correlated with the activity and severity of IBD to some extent,and was not correlated with ages,genders,and whether treated with anti-TNF-alpha agents.2.The expression of MMP-7 is significantly positively correlated with common activity indexes CRP and WBC,while the expression of MMP-7 was negatively correlated with these tight junction proteins.3.To some extent,MMP-7 can indicate the severity of intestinal intestinal inflammation,and may be an important indicator of inflammatory activity in IBD.