Pathological Mechanism Of ES2A In Ulcerative Colitis

The health of intestinal have a very important impact on our daily life,it is because intestinal is the largest digestive organ and the largest immune organ.Inflammatory bowel disease(IBD)are chronic inflammation which develop in the gut,and IBD include two major types:ulcerative colitis(UC)and crohn's disease(CD).The specific pathogenesis of UC pathogenesis is not clear at present.It is believed that UC is caused by multiple factors,mainly including microbial,environmental,genetic and immune factors.Our study mainly used mice model of colitis to explore the mechanism of ES2A.We collected the plasma and intestinal mucosal tissues of 20 UC patients and 20 normal people from the hospital,the plasma ES2A level were determined by ELISA and the colonic ES2A level were determined by western blot,the results showed that the expression level of ES2A of UC patients was decreased by 26.5%and 55.3%.in order to explore the mechanism of ES2A in UC,we used mice colitis model induced by Dextran sodium sulfate(DSS),we measured indexes such as weight change,disease activity index,histopathological score and colonic shortening of overexpressed ES2A group,the knockdown ES2A group,and the control group.The results showed that during the induction of DSS,compared to DSS control group,the ES2A overexpressed mice suffered from less body weight in day 10(8.5±3.4%vs 13.4±2.8%),got lower disease activity index(5.8±1.7 vs 7.3±1.8),got lower histopathological score(3.2±0.7 vs 5.2±1.7)and suffered from less shortening of the colon(colon lenth:7.1±0.4 vs 6.3±0.5 cm),while the ES2A knockdown mice suffered from greater body weight in day 10(21.8±4.1%vs 13.4±2.8%),got higher disease activity index(10.1±1.5 vs 7.3±1.8),higher histopathological score(7.0±1.0 vs 5.2±1.7)and suffered from less shortening of the colon(colon lenth:5.8±0.3 vs 6.3 ± 0.5 cm).We isolated the serum and colon tissues and detected the expression level and transcription level of inflammatory cytokines by ELISA and qPCR.The results showed that overexpression of ES2A reduced the level of inflammatory cytokines on DSS-fed mice,while knockdown of ES2A increased the level of inflammatory cytokines on DSS-fed mice.The mesenteric lymph nodes and colon tissue were collected from each group and digested into single cell,the proportion of Treg cells and Th17 cells in lamina propria lymphocytes(LPL)and mesenteric lymph nodes(MLN)were determined by flow cytometry.The results showed that overexpression of ES2A increased the proportion of Treg cells(LPL:3.0±0.3%vs 2.5±0.3%,MLN:22.8±4.7%vs 16.7±2.3%)while the knockdown of ES2A decreased the proportion of Treg cells(LPL:2.1 ±0.4 vs 2.5 ± 0.3%,MLN:13.14±2.7 vs 16.7±2.3%),and the overexpression of ES2A decreased the proportion of Th17 cells(LPL:11.1±2.8%vs 17.9±3.1%,MLN:6.3±1.6 vs 9.5±0.3%)while knockdown of ES2A increased the proportion of Th17 cells(LPL:23.9±2.8%vs 17.9±3.1%,MLN:13.3±1.6%vs 9.5±0.3%).These results suggested that ES2A increased the proportion of Treg cells and decrease Th17 cells in the inflammatory process to inhibit inflammation.We also explored the role of ES2A in the model of TCR ?-/-mice,the TCR ?-/-mice developed the spontaneous colitis at the age of 20th week.At 20th week we collected the colon tissues from the group of wild type TCR ?-/-mice,ES2A overexpressed mice and ES2A knockdown mice.By making paraffin sections and HE stanning,we found that ES2A overexpressed TCR ?-/-mice got lower histopathological score(3.5 ± 0.4 vs 4.2±0.3)while ES2A knockdown TCR ?-/-mice got higher histopathological score(5.7±0.6 vs 4.2±0.3)compared to wild type TCR?-/-mice.After the BNLCL.2 cells and Hep G2 cells were stimulated by IL-6 for 12 h,the expression level of ES2A in cellular supernatant was determined by ELISA,the results showed that ES2A decreased by 13.5%and 28.4%.The ES2A level of serum and hepatic mRNA decreased by 22.3%and 52.3%after the mice stimulated by IL-6.Therefore,it is concluded that ES2A inhibits inflammation by increasing the proportion of Treg cells and decreasing the proportion of Th17 cells,and IL-6 contributes to pathogenesis of ulcerative colitis by down-regulating the expression of ES2A.In another study,we discovered a novel elastase inhibitor from Hirudinaria manillensis and named it HMEI-A.In addition,HMEI-A has the ability to inhibit NETs formation and may plays a crucial role in blood-sucking by inhibiting the immune response of hosts.