The Partial Fountion Analysis And Mechanism Study Of Gut-liver Axis On Rats Suffered Severe Intra-abdominal Infection

Objective:To confirm the quantum of bacteria inducing a stable rats model of severe intra-abdominal infection.Dynamic observe organ injuries of lung,liver and ileum of rats suffered severe intra-abdominal infection. the changes of cell factors,the relationship of the different endotoxin level of portal vein with the degree damage of the liver. To explore the functional changes of gut-liver axis on rats suffered severe intra-abdominal infection.To observe the expression changes of interleuk in-1 receptor associated kinase-4 protein in liver injury during severe intra-abdominal infection in rats.To discuss the mechanism of interleuk in-1 receptor associated kinase-4 in liver injury during severe intra-abdominal infection. To investigate the effect of QingFeiChengQi Herbs on the lung and liver injury due to severe intra-abdominal infection.Methods:(1)Select 30 healthy adult male Wistar rats,They were divided intoA,B,Cgroups randomly.The rats of the model groups were injected with artificial gastric fluid and Escherichia coli mixed with adjuvant. Choose different level of bacteria as follows: 1ml/100g,0.9ml/100g、0.8ml/100g. The concentration of Escherichia coli Was 2x10 cfu/ml.To observe the mortality of rats in 48h, the pathology of lung,liver and Ileum. Choose the optimum dose of the E.coli.(2)When the rats model of severe intra-abdominal infection inducing multiple organ dysfunction syndrome was stable, Select 60 healthy adult male Wistar rats, divided into3 groups randomly:sham operation group,model group, the Chinese herb treated group. The model of the model group and the treatment group were injected with artificial gastric fluid and Escherichia coli mixed with adjuvant. The model of sham operation group was injected with the same dose of 10%BaS04 beef soup, QingFei ChengQi Herbs was poured into rat’s stomach of the treatment group through mouth in 2hours before molding and 10 hours after molding, the model group and the treatment group were poured into stomach through mouth with the same dose of distilled water, rats. At 3,6,12,24h after operation, Endotoxin content in portal vein,postcava and liver were measured using limulus test, TNF-α and IL-6 content were measured by ELISA,also ALT,AST were detected. ELISA. Pathological changes in lung,liver and ileum tissue were detected by light microscopy.(3) At 3,6,12,24h after operation, the expressions of interleuk in-1 receptor associated kinase-4 protein in liver tissue were assayed by western blot. Analysis the relationship of Endotoxin content and TNF-α with the expressions of interleuk in-1 receptor associated kinase-4 protein, the differences of all above date of 3groups are compared at same time.Results:(1) The mortality of model A,B,C was 50%、30%、10%. pathological damage of lung, liver and ileum were appeared in three groups.Model group A was the worst in three groups.(2) Endotoxin content was detected in 3h in three groups, the model group and the treatment group were higher than sham operation group,but there is no statistical significance (P<0.05),higher significance in 6h (P>0.05),slight lower in12h,riser in 24h.Compared the treatment group with the model group,there is no statistical Significance in Endotoxin content in 3h and 6h, lower significant than the model group in 12h and 24h (P<0.05); Compared the treatment group with the model group in endotoxin content of inferior vena cava, There were no significant differences in 12h (P>0.05). significantly reduced in 24h (P<0.05); Compared endotoxin content of portal vein with endotoxin content of inferior vena cava of the model group, There have obviously significant differences in 12h (P<0.05).no significant differences in 24h (P>0.05) Endotoxin content was detected in 3h in three groups,the model group and the treatment group were higher than sham operation group,,higher significance in 6h (P>0.05),slight lower in12h,riser in 24h. Compared the treatment group with the model group in endotoxin content of the liver, There were no significant differences in3h,6h,12h (P>0.05). significant higher than the sham group in every time period (P<0.05); significant lower than the model group in 24h (P<0.05); Compared the treatment group and the model group with the sham operation group, TNF-a content significantly rised in different time point (P<0.05). TNF-a content began to rise in 3h,came to peak inl2h,slightly decreased in 24h, Compared the treatment group with the model group, TNF-a content significantly decreased in12h and 24h(P<0.05); Compared the treatment group and the model group with the sham operation group, IL-6 content significantly rised in 6h(P<0.05), IL-6 content began to rise in 6h,came to peak in 24h, Compared the treatment group with the model group, IL-6 content significantly decreased in 24h(P<0.05); ALT, AST of the treatment group and the model group began to rise in 6h, Compared with the sham operation group, significantly higher in 6h,12h,24h(P<0.05). Compar-ed the treatment group with the model group, ALT, AST content significantly decrea-sed in 24h (P<0.05); pathological findings:Lung and terminal ileum were slightly edema with the naked eye in 3h,The light microscope findings showed pulmonary interstitial hyperemia and edema, Epithelial cells of terminal ileum get degenerated, hyperemia and edema were observed in submucosa and muscularis.With the extension of course,pathological damage got aggravated gradually.(3) Compared the treatment group and the model group with the sham operation grou-p, The expression of interleuk in-1 receptor associated kinase-4’mRNA and protein were higher in 3h, significantly higher in 6h(P<0.05). came to peak in 12h,slightly decreased in 24h,still significantly higher than the sham operation group (P<0.05); Via Pearson correlation,there are significantly positive correlations between the expression of interleuk in-1 receptor associated kinase-4 protein and endotoxin, TNF-a.Conclusion:(1) Using the concentration of 2x109cfu/ml and the bacteria level of lml/100g to create animal models,the model was stable, conform closely to MODS,It is very similar to intra-abdominal infection,It would be appropriate for experiment research.(2) gut barrier damage was the reason that enterogenous esotoxin translocation happened in rats suffered severe intra-abdominal infection.(3) Main function of gut-liver axis was to prevent endotoxin translocation,inactivate Endotoxin.(4) TNF-a rose quickly in the early stage of severe intra-abdominal infection,came to peak quickly, IL-6 rose slowly,but for a long duration,there have a positive relation with endotoxin.(5) Small doses of endotoxin in portal vein was detected in normal rats, endotoxin rose quickly in the early stage of severe intra-abdominal infection, clearance ability of liver gradually decreased in 12h, clearance ability of liver lose function in 24h.(6) The expression of interleuk in-1 receptor associated kinase-4 protein came to rise during severe intra-abdominal infection, TNF-a increased, TLRs/IL-1R pathway may be positively modulated by IRAK-4 kinase activity.(7) QingFeiChengQi Herbs can protect gut mucosa barrier,reduce enterogenous endotoxin content, the level of endotoxin and cytokine of rats suffered severe intra-abdominal infection,and inhibit IRAK-4 kinase activity, TLRs/IL-1R pathway activation.downregulate excessive release of TNF-.And above all, QingFeiChengQi Herbs can alleviate the damage degree of the rats liver during severe intra- abdominal infection.