| Purpose: In this study,we analyze the pathogenesis of liver cirrhosis intestinal endotoxemia from the perspective of wet turbidity,and explore the effect of Xiang Wei combination formula on plasma endotoxin level induced by compound factor in cirrhosis rats,reveal the liver protective mechanism of the Xiang Wei Combined Formula for treating the cirrhosis rats,reveal the mechanism of prevention and treatment of intestinal endotoxemia in cirrhosis and provide new ideas and methods for it.Method: Eighty five Wistar rats were randomly divided into normal group(10 rats),combination prevention group(abbreviated as prevention group)(10 rats),lactulose prevention group(abbreviated as prevention contrast group)(10 rats),model group(55 rats).In addition to the normal group,the rest were using carbon tetrachloride complex to make cirrhosis models.40% carbon tetrachloride olive oil solution was injected subcutaneously at a dose of 0.5ml/l00 g for the first time,twice a week at a dose of 0.3ml/l00 g,supplemented with 89.5% cornmeal,10% lard and 0.5% cholesterol,and 10 % Alcohol is the only drink.During the modeling period,the prevention group was given Xiang Sha six Junzi decoction at a dose of 6.875g·kg-1d-1 and Wei Ling decoction at a dose of 6.181 g.kg-1d-1 by the method of lavage alternately qod.Prevention contrast group was lavaged lactulose liquid at a dose of 2.083g·kg-1d-1.The normal group and the modelgroup were lavaged the same amount of normal saline 1ml/100 g once a day.Modeling lasted for 8 weeks.In the process of modeling,19 rats died,including 1 in the prevention group,2 in the prevention contrast group,12 in the pure model group.4 rats were sacrificed using estimating model,among those 2 were killed in the normal group,2 in the pure model group.After the success of modeling,the pure model group was randomly divided into 5 groups: model group(9 rats),Xiang Wei combination group(referred to Xiang Wei group)(8 rats),Xiang Sha six Junzi group(referred to Xiang Sha group)(8 rats),Wei Ling Tang group(referred to Wei Ling group)(8 rats),lactulose control group(referred to treatment contrast group)(8 rats).The rats in prevention group and prevention contrast group were continued to be given the same amount of previous liquid.The normal group and the model group were given the same amount of normal saline.The Xiang Wei group was treated with Xiang Sha Six Junzi Decoction of 6.875g·kg-1d-1,Wei Ling decoction 7.188 g·kg-1d-1 alternately qod.The Xiang Sha group was treated with Xiang Sha Six Junzi Decoction at a dose of 6.875g·kg-1d-1.Wei Ling group was given 7.188g·kg-1d-1;treatment contrast group was given lactulose oral solution 2.083g·kg-1d-1,all of the medication were used for 4 weeks.During the course of the experiment,the general condition and weight change of the rats were observed.After the treatment,the rats were killed.We observed of the rats' fresh liver,calculated the index of the liver weight and spleen weight and used an automatic biochemistry analysis of the AST,ALT,ALB,GLB,A/G.The levels of plasma endotoxin were measured by limulus reagent and chromatin method.The serum NO,HA,LN,PC?,?-C,TNF-?,IL-6,IFABP,and D-lactic acid were measured by ELISA and also the Hyp(hydroxyproline)in rats' liver tissue and S-Ig A in the intestines.The pathological tissues of liver cirrhosis were assessed by HE staining and Masson staining.The intestinal tissues injury assessment of the rats was observed by HE staining under optical microscope.The ?-SMA,the TLR4 and expression of NF-? B in rat liver was detected by immunohistochemistry.The expression of TLR4 m RNA,My D88 m RNA and NF-?B m RNA in rat liver,Occludin m RNA Expression in small Intestine tissue of rats were detected by Real-time PCR.Result: 1 The Xiang Wei combination group,Xiang Sha Six Junzi Decoction,Wei Ling group and Lactulose can improve the general condition of the rats and increase the weight.Among them,the Xiang Wei group,the Xiang Sha group and the Wei Ling group have no significant difference in the aspect of gaining weight(P<0.05).There was a significant difference between the Xiang Wei combination group and treatment contrast group(P<0.01).2 The index of the liver weight and spleen weight: Compared with the model group,there was a significant difference in the normal group,the prevention group,the prevention contrast group,the Xiang Wei group,Xiang Sha group,Wei Ling group and the treatment contrast group(P<0.01).There were significant differences between the prevention group and the prevention contrast group(P<0.05).Compared with the prevention group,there were significant differences among Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P <0.01).There was no significant difference among Xiang Wei group,Wei Ling group,and Xiang Sha group(P>0.05).Xiang Wei group was very different from prevention contrast group(P<0.01).Compared with the treatment contrast group,there were significant differences among Xiang Wei group,Wei Ling group and Xiang Sha group(P<0.05);Compared with the normal group,there were significant differences among the prevention group,the prevention contrast group,Xiang Wei group,Wei Ling group,Xiang Sha group and the treatment contrast group(P<0.01).3 The levels of liver function,HA,LN,PC?,?-C,NO,TNF-?,IL-6 and Hyp in liver tissue: compared with the model group,there were significant differences among the normal group,the prevention group,the prevention contrast group,the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P<0.01).There were significant differences between the prevention group and prevention contrast group(P<0.05);Compared with the prevention group,there were significantly differences among Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P<0.01);there was no significant difference among Xiang Wei group,Wei Ling group and Xiang Sha group(P>0.05);there were significant differences between Xiang Wei group and the prevention contrast group(P<0.05 or P<0.01).Compared with the treatment contrast group,there were significant differences among Xiang Wei group,Wei Ling group,and Xiang Sha group(P<0.05);Compared with the normal group,there were significant differences among the prevention group, the prevention contrast group,the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P <0.01).4 The levels of endotoxin,IFABP and D-lactic acid:Compared with the model group,there were significant differences among the normal group,the prevention group,the prevention contrast group,the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P<0.01).There was significant difference between the prevention group and the prevention contrast group(P<0.05).Compared with prevention group,there were significant differences among the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P<0.01).Compared with the Xiang Wei group,there were significant differences between Wei Ling group and Xiang Sha group(P<0.05),and there were significant differences between the prevention group and the prevention contrast group(P<0.01).There was significant difference between the Wei Ling group and the Xiang Sha group compared with the treatment contrast group(P<0.05).There was no significant difference between the Wei Ling group and the Xiang Sha group(P>0.05).Compared with the normal group,there were significant differences in the prevention group,the prevention contrast group,the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P <0.01).5 Liver histopathology: Percentage of collagen fibers in liver tissue of each group showed that compared with the model group,there were significant differences in the normal group,the prevention group,the prevention contrast group,the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P <0.01).There was significant difference between the prevention group and the prevention contrast group(P<0.05).Compared with the prevention group,there were significant differences among the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P<0.01).There was no significant difference among Xiang Wei group,Wei Ling group and Xiang Sha group(P>0.05);There were significant differences between Xiang Wei group and prevention contrast group(P<0.01).There were significant differences among the Xiang Wei group,Wei Ling group and Xiang Sha group compared with the treatment contrast group(P<0.05).Compared with the normal group,the differences were significant in the prevention group,the prevention contrast group,the Xiang Wei group,Wei Ling group,Xiang Sha group and the treatment contrast group(P <0.01).6 Small intestinal histopathology: The small intestinal mucosal epithelial cell degeneration,necrosis,a large number of intestinal villi shedding,lamina propria exposure,intestinal mucosal heavy injury in the model group;the small intestinal mucosa villi apical subcutaneous was showed cystic space and the intestinal mucosal injury of rats in the prevention group were lighter than that of the prevention contrast group.A few fluff shedding of the the small intestinal mucosa villi apical happened in the Xiang Wei group,Wei Ling group and Xiang Sha group,and there an obvious edema in lamina propria.The intestinal mucosal injury degree were lighter in the Xiang Wei group than that in the Xiang Sha group and Wei Ling group,and more serious than the one in the prevention contrast group.The treatment contrast group degree of intestinal mucosal injury was between the model group and the treatment group.7 The expression results of ?-SMA,TLR4,NF-? B immunohistochemistry and TLR4 m RNA,My D88 m RNA and NF-?B m RNA in liver tissue: compared with the model group,there were significant differences in the normal group,the prevention group,the prevention contrast group,the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P<0.01).There was a significant difference between the prevention group and the prevention contrast group(P<0.05).Compared with the prevention group,there were significant differences among the Xiang Wei group,Wei Ling group,Xiang Sha group and treatment contrast group(P <0.01).There was no significant difference among the Xiang Wei group,Wei Ling group and Xiang Sha group(P > 0.05);There were significant differences between Xiang Wei group and prevention contrast group(P<0.01);Compared with the treatment contrast group,there were significant differences in the Xiang Wei group,Wei Ling group,and Xiang Sha group(P<0.05 or P<0.01).Compared with the normal group,there were significant differences in the prevention group,the prevention contrast group,the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P<0.05 or P<0.01).8 The expression results of S-Ig A and Occludin m RNA in small intestine: compared with the model group,there were significant differences in the normal group,the prevention group,the prevention contrast group,the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P<0.01).There was a significant difference between the prevention group and the prevention contrast group(P<0.05).Compared with the prevention group,there were significant differences among the Xiang Wei group,Wei Ling group and Xiang Sha group and treatment contrast group(P<0.01).Compared with the Xiang Wei group,there were significant differences between Wei Ling group and Xiang Sha group(P<0.05).Compared with the Xiang Wei group,there was a significant difference between the prevention group and the treatment contrast group((P<0.05 or P<0.01).Compared with the treatment contrast group,there were significant differences between Wei Ling group and Xiang Sha group(P<0.05).There was no significant difference between Wei Ling group and Xiang Sha group(P>0.05).Compared with the normal group,the differences were significantly in the prevention group,the prevention contrast group,the Xiang Wei group,Wei Ling group,Xiang Sha group,and the treatment contrast group(P<0.01).Conclusion: Xiang Wei combination Formula can remove intestinal endotoxin effectively,improve intestinal mucosal permeability,reduce plasma endotoxin levels significantly,reduce liver damage,and delay the progress of rat cirrhosis.It will be better if use medicine early for prevention.The mechanism of action may be like this: 1 The Xiang Wei combination Formula can reduce the level of serum D-lactic acid and small intestinal fatty acid binding protein in cirrhotic rats,reduce the intestinal mucosal permeability,increase the expression of Occludin protein and secretion of S-Ig A,thus it can decrease plasma endotoxin levels in cirrhotic rats.2 Xiang Wei combination Formula can suppress TLR4-My D88-NF-?B signal pathway caused by endotoxin in the plasma.It also can decrease expression of TLR4 protein,My D88 protein and NF-?B protein,and the levels of TNF-?,IL-6 and NO.Thereby it can reduce endotoxin damage to the liver of cirrhotic rats.3 Xiang Wei combination Formula not only can improve the liver function,but also can decrease the levels of plasma endotoxin and serum liver fibrosis markers and portion of Hyp and ?-SMA in liver tissue,thus it can delay the progress of liver cirrhosis in rats. |
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