Effects Of Trichinella Spiralis And Its Serine Protease Inhibitors On Intestinal Mucosal Barrier Function

Trichinosis is an important food-borne zoonotic disease,which is prevalent in many parts of the world and affects public health security.Many studies have shown that trichinella spiralis can damage the intestinal mucosal barrier function during invasion,thus facilitating invasion.However,the mechanism related to the damage of intestinal mucosal barrier during insect invasion remains to be further explored.In this study,the Kazal Serine protease inhibitor Ts Ka SPI and Serpin Serine protease inhibitor Ts Ad SPI in Trichinella spiralis and its excreted and secreted antigens were used as research objects to explore the effect of Trichinella spiralis,Ts Ka SPI and Ts Ad SPI on the intestinal mucosal physical barrier during the intestinal invasion.The influence of transcription levels of genes related to chemical barrier and immune barrier lays a foundation for further exploration of the related mechanism of intestinal invasion after trichinosis infection,and also provides a relevant basis for prevention and control of trichinosis.Firstly,in order to study the influence of trichinella spiralis on intestinal mucosal barrier during host invasion,the mechanical barrier,chemical barrier and immune barrier related proteins of duodenum,jejunum and ileum tissues were detected by carrying out infection tests of trichinella spiralis on different days,and the mechanical barrier of jejunum tissues was found in the process of trichinella spiralis infection.The chemical barrier and immune barrier were most significantly affected,so jejunum tissue was identified as the gut of choice to investigate the effect of trichinella spiralis on the intestinal mucosal barrier during host invasion.Then the damage of jejunum tissue was analyzed by hematoxylin-eosin(H&E)staining.The results showed that trichinella spiralis had intestinal villi loss,inflammatory cell infiltration in jejunum,and jejunum tissue lesions to a certain extent.real-time fluorescent quantitative PCR(RT-q PCR)was used to detect changes in transcription levels of genes associated with mechanical,chemical and immune barriers in intestinal mucosal barriers.ZO-1,Occludin,Claudin-3 in Tight junction protein(TJ)at the mechanical barrier level were detected by Western blot(WB).MUC1 and MUC2 in mucin(MUC)at the chemical barrier level;tumor necrosis factor-α(TNF-α)in pro-inflammatory cytokines at the immune barrier level,interleukin-1β(IL-1β)and transforming growth factor-β(TGF-β)in anti-inflammatory cytokines,The expression of interleukin-10(IL-10)and barrier related proteins such as TLR1,TLR2 and TLR4 in Toll-like receptor(TLR).The results showed that the intestinal mucosal barriers,including mechanical barriers,chemical barriers and immune barriers,were damaged during the intestinal stage of trichinella spiralis infection.We then further investigated the effects of inhibition in Trichinella trichinella’s secretion antigens,Ts Ka SPI and Ts Ad SPI,on the intestinal mucosal barrier.In this part of the experiment,the in vitro culture model of trichinella spiralis was established by intestinal epithelial cells(IPECs).Ts Ka SPI and Ts Ad SPI were selected for in vitro intervention.The expression of related proteins was detected by q PCR and Western blot,and the expressions of Claudin-3 and MUC2 were detected by Immunological Fluorescence Assa(IFA).enzyme-linked immunosorbent assay(ELISA)was used to detect the secretion and expression of inflammatory cytokines.The results showed that Ts Ka SPI and Ts Ad SPI caused a decrease in the expression of tight-junction proteins in the mechanical barrier and mucin in the chemical barrier,as well as an increase in the expression of pro-inflammatory factors and anti-inflammatory factors in the immune barrier,and an increase in the expression of TLR4 in the toll-like receptor.These results indicate that Ts Ka SPI and Ts Ad SPI can damage the mechanical,chemical and immune barriers in the intestinal mucosal barrier.Finally,we established a mouse model of intraperitoneal injection of Ts Ka SPI and Ts Ad SPI.immunohistochemistry changes of intestinal mucosal barrier related genes and proteins were assessed by hematoxylin-eosin(H&E)staining,q PCR,Western blot,and immunohistochemistry(IHC)and ELISA.The results also confirmed that Ts Ka SPI and Ts Ad SPI could damage intestinal mucosal barrier function.In summary,we can find that trichinella spiralis destroys intestinal mucosal barrier function during invasion,and Ts Ka SPI and Ts Ad SPI secreted by trichinella spiralis also play a positive role in this process.Our results will provide theoretical basis and experimental support for the pathogenesis of trichinosis.It lays a foundation for further exploring the relationship between trichinella spiralis and intestinal mucosal barrier function of the host,as well as exploring the related mechanism of trichinella spiralis invasion in the intestinal tract after infection,and also provides some experimental basis and relevant theoretical basis for the prevention and treatment of trichinosis.