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Objective To discuss the influence of Tanshinone ⅡA on the tight junction protein of intestinal mucosal epithelial cells in rat severe septic models. Methods Seventy-five Sprague-Dawley (SD) rats were randomly divided into sham operation group, model group and Tanshinone ⅡA injection high (20 mg/kg), medium (10 mg/kg) and low (5 mg/kg) dose groups, each group 15 rats. Sepsis rat models were established by cecal ligation and puncture (CLP) method, in sham operation group, only switched abdominal surgery was performed without CLP. In Tanshinone ⅡA injection groups, different doses of Tanshinone ⅡA were injected intraperitoneally after modeling for 10 minutes and 6 hours; in sham operation and model groups, equal volume of normal saline was injected intraperitoneally at the same times as above. After operation, 3 L/kg of normal saline was injected into the caudal vein in all rats for fluid resuscitation.Twelve hours after operation, the rats were killed, the abdominal lymph nodes, liver, spleen and kidney tissues were taken for bacterial culture and calculating the rate of bacterial translocation; under microscope, the histopathological changes of ileum mucosal tissues were examined and Chiu scoring was carried out; TdT-mediated dUTP nick end labeling (TUNEL) was applied to detect the ileum mucosal epithelial cell apoptosis and calculating the index (AI);fluorescence immunoassay and Western Blot methods were used to measure the contents and protein expression levels of tight junction protein, junctional adhesion molecule-1 (JAM), Claudin-1, Zonula occludens-1 (ZO-1), Occludin, c-Fos and Tryptase. Results ① In bacterial cultures of abdominal lymph node, liver, spleen and kidney, the positive rate of mesenteric lymph node was the highest, followed by liver and spleen, mainly Escherichia coli, Proteus mirabilis, etc. The highest positive rate of bacterial culture was in model group (38.8%), followed by low dose of Tanshinone ⅡA injection group (35.0%), and the lowest was 16.6% in high dose Tanshinone ⅡA injection group, the differences being statistically significant in comparisons between any pair of groups (all P < 0.05). ② Pathological examination showed that the pathological changes of ileum mucosa were obvious and the Chiu score (4.17±0.98 vs. 0) and AI (11.70±2.87 vs. 2.17±0.80) in model group were significantly higher than those in sham group (all P < 0.05); with the increase of dosage of Tanshinone ⅡA injection, the pathological changes of rat ileum mucosa were improved gradually, the Chiu score and AI were decreased gradually, and the degrees of decrease in high dose Tanshinone ⅡA group were more significant than those in model group (Chiu score: 1.12±0.79 vs. 4.17±0.98, AI: 3.65±1.98 vs. 11.70±2.87, both P < 0.05).③ Immunofluorescence staining showed that the positive staining of protein JAM, ZO-1 and c-Fos were all green in color, Claudin-1, Occludin and Tryptase were all red in color, the localizations of all of them were in the cytoplasm, the protein expression of JAM, Claudin-1, ZO-1, Occludin from strong to weak in turn were Sham group, high, medium, low dose Tanshinone ⅡA group and model group, the expression of c-Fos, Tryptase from strong to weak in turn were model group, low, medium, high dose Tanshinone ⅡA group and Sham group. ④ Western Blot showed that the expressions of ileum tissue JAM, Claudin-1, ZO-1 and Occludin in model group were all significantly lower than those of the sham group, while the expressions of c-Fos, Tryptase were obviously higher than those of the sham group, with the increase of dosage of Tanshinone ⅡA, the expressions of JAM, Claudin-1, ZO-1 and Occludin were increased gradually and the protein expressions of c-Fos and Tryptase were gradually decreased, and the changes in high dosage group of Tanshinone ⅡA were more significant than those in low and moderate groups [JAM (gray value): 25.39±1.82 vs. 12.41±1.34, 19.45±1.66, Claudin-1 (gray value): 28.44±1.56 vs.17.26±1.46, 21.23±1.34, ZO-1 (gray value): 28.84±1.59 vs. 16.45±1.21, 24.22±1.46, Occludin (gray value): 25.49±1.63 vs. 13.34±1.45, 19.45±1.37, c-Fos (gray value):15.76±1.36 vs. 27.84±1.36, 21.22±1.73, Tryptase (gray value): 14.44±1.41 vs. 28.14±1.38, 22.32±1.57], all the above comparisons of different dosage groups were statistically significant (all P < 0.05). Conclusion Tanshinone ⅡA injection may improve intestinal wall structure and reduce bacterial translocation by improving the intestinal mucosal tight junction protein in sepsis model rats, and this effect is positively correlated to Tanshinone ⅡA dosage.
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Objective To investigate the effects of ω-3 polyunsaturated fatty acids (ω-3 PUFA) on inflammatory response of intestine and bacteria translocation in rats with traumatic shock (TS) in order to explore the underlying mechanism.Methods A total of 36 male Wistar rats provided by Academy of Military Medical Sciences Animal Center were assigned randomly (random number) into 3 groups (n =12 in each group):sham operation group,TS model group and PUFA pretreatment group.Rat models of IS were established by comminuted fracture of femur and depletion of blood,and 2 mg/kg ω-3 PUFA or normal saline were injected 12 hours and 2 hours before modeling.Blood specimens were collected and intestinal tissue samples were obtained 120 min after modeling.The serum levels of tumor necrosis factor-o (TNF-α),IL-1β,IL-10 and 8-iso-prostaglandin F 2α (8-iso-PGF2α) were measured with ELISA.Light microscopic examination was carried out for histopathological assessment of the intestina tissue and the intestinal mucosa damage index ( IMDI ) was calculated.The number of marked bacilli found in mesenteric lymph nodes,lung,liver,spleen,and kidney tissues were counted under a fluorescent microscope.The percentages for categorical variables and mean ± SD for continuous variables were expressed. Chi-square test and unpaired t-test were used for comparisons among groups,and statistical significance defined as P ＜ 0.05.Results The levels of TNF-α,IL-1β,IL-10 and 8-iso-PGF2α,the IMDI and the positive rates of bacteria translocation in TS model group were [ (325.14 ±21.17) ng/ml,(26.93 +2.58) μg/L,(7.59 ± 1.26) μg/L,(259.73 +61.32) pg/ml,(4.15 +0.37) and 58.33％,respectively] and those in PUFA group were [ (251.47 + 19.16) ng/ml,(17.81±1.94) μg/L,(9.44±1.85) μg/L,(171.44±39.25) pg/ml,(3.28±0.43) and 36.67％,respectively ].And those biomarkers in both TS group and PUFA group were higher obviously than those in sham group [ (37.02 ±5.54) ng/ml,(2.49 ±0.67) μg/L,(2.93 ±0.74) μg/L,(81.26 ± 15.18) pg/ml,(0.33 ±0.12) and 6.67％,respectively,P＜0.01].Compared with TS model group,the levels of TNF-α,IL-1β and 8-iso-PGF2α,the IMDI and the positive rates of bacteria translocation were lower,and the levels of IL-10 were higher in PUFA group ( P ＜ 0.01 or P ＜ 0.05 ).Conclusions The supplementation of ω-3 PUFA lessens the injury of intestina mucosa after traumatic shock,and it may be associated with the inhibition of inflammatory response by intestine and bacteria translocation.was carried out for histopathological assessment of the intestina tissue and the intestinal mucosa damage index ( IMDI ) was calculated.The number of marked bacilli found in mesenteric lymph nodes,lung,liver,spleen,and kidney tissues were counted under a fluorescent microscope.The percentages for categorical variables and mean ± SD for continuous variables were expressed. Chi-square test and unpaired t-test were used for comparisons among groups,and statistical significance defined as P ＜ 0.05.Results The levels of TNF-α,IL-1β,IL-10 and 8-iso-PGF2α,the IMDI and the positive rates of bacteria translocation in TS model group were [ (325.14 ±21.17) ng/ml,(26.93 +2.58) μg/L,(7.59 ± 1.26) μg/L,(259.73 +61.32) pg/ml,(4.15 +0.37) and 58.33％,respectively] and those in PUFA group were [ (251.47 + 19.16) ng/ml,(17.81±1.94) μg/L,(9.44±1.85) μg/L,(171.44±39.25) pg/ml,(3.28±0.43) and 36.67％,respectively ].And those biomarkers in both TS group and PUFA group were higher obviously than those in sham group [ (37.02 ±5.54) ng/ml,(2.49 ±0.67) μg/L,(2.93 ±0.74) μg/L,(81.26 ± 15.18) pg/ml,(0.33 ±0.12) and 6.67％,respectively,P＜0.01].Compared with TS model group,the levels of TNF-α,IL-1β and 8-iso-PGF2α,the IMDI and the positive rates of bacteria translocation were lower,and the levels of IL-10 were higher in PUFA group ( P ＜ 0.01 or P ＜ 0.05 ).Conclusions The supplementation of ω-3 PUFA lessens the injury of intestina mucosa after traumatic shock,and it may be associated with the inhibition of inflammatory response by intestine and bacteria translocation.
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Objective To investigate the effect of probiotic supplement on the microbiology,bacterial translocation,and gut barrier function of the rats with abdominal infection. Methods After making the models of cecal ligation and perforation,the SD rats were divided into two groups.The rats in the control group were administrated with parenteral nutrition.The rats in the experimental group were administrated with parenteral nutrition and probiotics via the needle jejunostomy and neck vein for 5 d.The feces in the cecum were cultured by anaerobic bacterial growth.The vena cava blood and the homogenated tissues of the lives,hungs and mesenteric lymph nodes were cultured to determine bacterial translocation. Results The quantity of normal intestinal bacteria in the experimental group and the control group had significant difference except Enterobacteriaceae.The quantity of L.acidophilus and Bifidobacteria in the experimental group were higher than those in the control group.The quantity of C.perfrigens,the potential pathogenic germ in the experimental group were lower than those in the control group(P
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Objective To explore the effect of glutamine on immune function of rat with obstructive jaundice and its possible mechanism. Methods Fifty male Wistar rats were randomly divided into three groups: Control group (n=10), obstructive jaundice group (n=20) and glutamine treatment group (n=20). The serum concentration of TNF-?, IL-10 was detected by using radioimmune method. Liver function was measured through automated biochemistry analyzer. The animal model of obstructive jaundice was established by ligating the rat's common bile duct. Bacteria cultures were performed with the rat's tissues of lung, spleen, liver and kidney respectively. Resu- lts Compared with control group, obstructive jaundice group showed statistically lower serum level of TNF-?, and statistically higher serum level of IL-10, TBIL, ALT and AST during the first and the second week after ligation of common bile duct. During the first and second week after administration of glutamine, the serum TNF-? of glutamine treatment group was statistically higher than that in control group and obstructive jaundice group. Meanwhile, glutamine treatment group showed statistically lower serum level of IL-10, TBIL, ALT and AST than obstructive jaundice group. There were statistically less bacteria translocations in glutamine treatment group than those in obstructive jaundice group. Conclusion Glutamine can increase the immune function by changing serum concentration of TNF-?, IL-10 and decrease the bacteria translocation.
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Currently,the popular notions on the mechanisms of gut-derived-infection are described as follows.The stress may cause the damage of intestinal mucosal barrier and dysfunction of intestinal immune response,which leads to the alteration of intestinal flora,abnormal proliferation of opportunistic pathogen as well as the translocation of alive bacteria and their toxin into systemic compartment.As a result,the proinflammatory cytokines are released to induce the outbreak of intestinal inflammation or systemic inflammatory response syndrome,even the MODS.This review focuses on the relationship between biological behavior of intestinal bacteria and gut-derived-infection.
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Objective To investigate the relationship between the plasma cytokines and the translocation of intestinal bacteria and endotoxin after gut barrier injury in severe acute pancreatitis (SAP) rats. Methods SD rats were divided randomly into sham operation group(n=36) and SAP group (n=36). The rat model of SAP was set up by retrograde injection of 4% sodium taurocholate in biliopancreatic duct. Morphological changes of pancreas and ileum were observed. The plasma levels of TNF-a,IL-6 and IL-10 were determined by ELISA. The plasma levels of DAO activities and LPS were measured at various time points. The rates of bacterial translocation to abdominal organs were also calculated. Results The plasma levels of TNF-a and IL-6 obviously elevated immediately after SAP induction and reached peak value at 48 hours, and the plasma IL-10 level significantly increased only 6 hours after SAP induction. Plasma DAO activities increased at the early stage of SAP and obviously decreased at 24 hours. Plasma LPS levels also increased significantly at the early stage of SAP and reached peak value at 48 hours. The rates of bacterial translocation to organs sharply increased 24 hours after SAP induction and reached 58.3% at 72 hours. Conclusion The levels of cytokines increased and gut barrier function was injured in the early stage of SAP. Cytokines may impair the intestinal microcirculation and gut barrier function, which could promote the intestinal bacteria and endotoxin translocation. Simultaneously, intestinal bacteria-endotoxin translocation could also induce excessive release of cytokines and aggravate the gut barrier damage, which might cause systemic inflammatory response syndrome and multiple organ disfunction syndrome. There was a close relationship beween cytokines and the translocation of intestinal bacteria and endotoxin in SAP.
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Objective To observe the changes in gut mucosal barrier and gut-origin bacteria-endotoxin translocation in acute necrotizing pancreatitis (ANP) rats. Methods Wistar rats were divided randomly into normal group (n=6), sham operation group (n=30) and ANP group (n=39). ANP was introduced by infusion of artificial bile into biliopancreatic duct. Morphology of pancreas and intestine were observed and tight junction on ileum epithelia were assessed by cryofracture replicas electroscopy. Plasma levels of D-lactic acid and endotoxin were examined at various time points. The rates of bacterial translocation to abdominal organs were also calculated. Results Mucosal and tight junction damages of the gut were found during early stage of ANP. Simultaneously, plasma D-lactate levels increased and endotoxemia occurred. The rate of bacterial translocation to organs was 59.5% 72h after ANP occurred. Conclusions Gut barrier function can be injured in the early stage of ANP, and resulting in gut origin bacteria-endotoxin translocation, which may be the originator of systemic inflammatory reaction and secondary infection of the pancreas.