Emodin Improves Intestinal Dysfunction In Septic Mice By Regulating The Intestinal Barrier, Intestinal Flora And Immune Homeostasis

BackgroundSepsis is an uncontrolled reaction of the host to infection and leads to life-threatening organ failure.It is a common disease in ICU and has a high mortality rate.The pathological mechanism of sepsis and its complexity are currently not very clear.The treatment of sepsis is generally supportive,including fluid resuscitation,vasoactive substances and antibiotic treatment to eliminate the pathogen.Patients with sepsis are often accompanied by gastrointestinal dysfunction,causing intestinal microflora disorder and increased intestinal permeability.The bacterial translocation further exacerbates the disease,aggravates the patient's condition,and seriously affects its prognosis.The traditional Chinese medicine Rhubarb is a commonly used Chinese medicine for diarrhea.It has the effects of diarrhea attack,detoxification,stasis and menstruation.It is widely used in shock,acute pancreatitis,multiple organ dysfuction syndrome,and sepsis,abdominal infections,etc.Emodin is an active ingredient that can improve intestinal barrier dysfunction.However,emodin is not clear on the integrity of intestinal mucosal barrier,intestinal microecology and immune homeostasis in sepsis.In this study,emodin was used to explain the mechanism of intestinal dysfunction in sepsis mice from the aspects of regulating the integrity of intestinal mucosal barrier,intestinal flora and immune homeostasis,providing experimental support for clinical application.ObjectiveObserve the effects of emodin on the general life status,survival rate,cellular inflammatory factors,mesenteric lymph node bacterial translocation,intestinal pathological changes,etc.in sepsis mice;Explore the effect of emodin on the intestinal microecology of sepsis,and analyze the characteristics of the intestinal microecological structure changes in sepsis mice to provide a theoretical basis for using the intestinal microecology as a biomarker of sepsis;The sensitive molecular markers of sepsis were screened at the level of peripheral blood protein,and the effect of emodin on the inflammatory immune response of sepsis was analyzed.MethodExperiment 1:Cecal ligation and punctual(CLP)was used to establish a sepsis mouse model.The experiment was divided into:sham operation group,model group and emodin group.The emodin suspension solution(high,medium and low dose group)and normal saline of the model group were injected intraperitoneally for 5 days before observation.The general life status and 7-day survival rate of mice in each group were observed.Experiment 2:The dose with the most significant increase in survival rate was used as the follow-up experimental dose.The sham operation group,model group and emodin group were sampled and indexed at 24h and 48h after surgery.HE staining of mouse ileal tissue to observe the pathological state of intestinal wall;Detection of serum inflammatory factors IL-13,IL-6 and TNF-? in mice by ELISA;Western blot detection of Occludin protein expression in ileal tissue;The traditional bacterial culture method was used to detect the shift of E.coli in mesenteric lymph nodes in each group at 48 hours after operation.Experiment 3:The feces of the sham operation group,model group and emodin group were collected at 24h and 48h after operation,and the intestinal microecological diversity was analyzed by 16SrDNA high-throughput sequencing technology,and the main components of the mouse intestinal flora were compared between different groups and colony expression differences.Experiment 4:Sera from the sham-operated group,model group and emodin group were collected at 24h and 48h after the operation.Using liquid chip technology,biomarkers with diagnostic value for sepsis were screened to establish immune homeostasis.ResultsExperiment 1:Rhubarb medium dose group significantly improved the survival rate and general life status of mice with sepsisExperiment 2:(1)The emodin group reduced ileal epithelial cell infiltration,significantly improved ileal epithelial cell morphology,and enhanced the integrity of epithelial cells;(2)The emodin group significantly reduced the expression of serum inflammatory factors IL-1?,IL-6 and TNF-? in sepsis mice;(3)The emodin group reduced the bacterial translocation of mesenteric lymph nodes and increased the expression of Tight junction-related proteins in ileal epithelial cells.Experiment 3:(1)Significant changes in the composition of intestinal flora in sepsis mice caused disorder of intestinal flora,reduction of thick-walled phylum and increase of proteobacteria.The abundance of sex anaerobic bacteria is reduced;(2)The emodin group significantly increased the abundance and diversity of intestinal bacteria in mice with sepsis;(3)In the early stage of sepsis,emodin can effectively balance the composition of intestinal flora in mice with sepsis and inhibit the growth of pathogenic bacteria.Experiment 4:(1)The data were analyzed using Moderated t-statistics and Fisher's exact test.Compared with the model group,the levels of anti-inflammatory factors in mice in the emodin group such as IL-4 and IL-10 and proinflammatory factors IL-1?,IL-2,IL-6 and TNF-?;(2)For enrichment analysis of pathways,the differentially expressed genes in the model group and the emodin group are also mainly involved in signaling pathways related to inflammatory immunity,cytokine-cytokine-receptor interaction,chemokine signaling pathway,and inflammatory bowel disease IBD),IL-17 signaling pathway,T cell receptor signaling pathway,etc.,so as to establish immune homeostasis,has a good early warning diagnosis value for sepsis.Conclusion Emodin improves intestinal dysfunction in sepsis mice by regulating intestinal mucosal barrier integrity,intestinal flora and immune homeostasis.