Distribution of Chromate-labeled Brucella abortus Endoxin in Tolerant Mice

Endotoxin tolerance as manifested by a lesser degree of hypoferremia was demonstrated in mice when both pretreatment (10 mug per injection) and challenge (100 mug) does of Brucella abortus endotoxin were administered intraperitoneally. Qualitative and quantitative studies on the distribution of chromate-labeled endotoxin in normal mice revealed that the endotoxin localized predominately in the liver and hypoferremia could be related to a high uptake of endotoxin by this organ. In tolerant mice, the labeled endotoxin was found mainly in the mesenteric lymph nodes (MLN) with smaller quantities in the blood, spleen, and liver. Experiments with splenectomized mice provided supporting evidence that the liver was the target organ of the hypoferremic response to endotoxin. High localization of endotoxin in the MLN with lower quantities in the blood, livers, and spleens of tolerant mice indicated that tolerance may be the result of a blockage by the MLN, preventing the endotoxin from reaching the liver. This inference was supported by the finding that hypoferremic tolerance did not occur when the hypoferremia-provoking dosage of endotoxin (100 mug) was given intravenously to mice pretreated intraperitoneally. There was less hypoferremia in normal mice injected with a mixture of antiserum and 100 mug of endotoxin than in mice given the same dosage of endotoxin in saline. Distribution studies on endotoxin treated with specific antiserum revealed that the endotoxin localized principally in the MLN, thus preventing most of the endotoxin from reaching the liver, the target organ of the hypoferremic response.     

Diversity of endotoxin-induced nitrotyrosine formation in macrophage-endothelium-rich organs

The administration of bacterial lipopolysaccharide (LPS; endotoxin) can stimulate the development of the systemic inflammatory response syndrome, which can compromise the function of many organ systems, resulting in multiple organ failure. Activation of macrophages and cytokines by endotoxin and the subsequent formation of reactive oxygen and nitrogen species are of central pathogenic importance in various inflammatory diseases including sepsis. However, whether different tissues behave the same in pathological changes produced by LPS and what factors may affect pathological processes and protein tyrosine nitration in different organs, still remain to be evaluated. In the present study, we investigated the distribution of nitrotyrosine and other pathological changes induced by LPS in rat liver, spleen, and lung, all of which are rich in macrophages and endothelial cells. Our study revealed two important findings: first, a denitration activity in spleen white pulp might play a key role to protect the areas from nitration. Similar activity might also exist in endothelial cells of sinusoids and capillaries. Second, protein nitration might not induce significant tissue damage as shown in liver and spleen. However, inflammatory infiltration with increased formation NO* and other reactive species may result in severe tissue injury, as demonstrated in lung after LPS administration.     

参麦注射液对严重烫伤大鼠肠道屏障功能的保护作用

目的:探索参麦注射液对30% Ⅲ°烫伤早期肠道屏障功能的保护作用,为参麦注射液防治肠源性感染提供实验依据。方法:Wistar大鼠60只,随机分为正常对照组、模型对照组,地塞米松5 mg/kg组、参麦注射液5、10、15 mg/kg组,每组10只,使用烫伤仪建立30% Ⅲ°烫伤动物模型,立即腹腔注射相应的药物,每天1次。烫伤72 h后,检测肝脏、脾脏、肠系膜淋巴结细菌移位量、血浆内毒素、二胺氧化酶(DAO)、肿瘤坏死因子-α(TNF-α)及白细胞介素-6(IL-6)水平和肠粘膜分泌型免疫球蛋白A(sIgA)的水平。结果:与正常对照组比较,模型组肝脏、脾脏、肠系膜淋巴结细菌移位量,血浆内毒素、DAO、TNF-α及 IL-6和肠黏膜sIgA水平明显升高(P＜0.01);与模型组比较,地塞米松组和参麦注射液5、10、15 mg/kg组肝脏、脾脏、肠系膜淋巴结细菌移位量,血浆内毒素、DAO、TNF-α及 IL-6和肠黏膜sIgA水平明显降低(P＜0.05或P＜0.01)。结论:参麦注射液可减轻严重烫伤引起的肠粘膜损伤,效果与地塞米松相当,高剂量组效果更好。     

The Contribution of Gut-Derived Endotoxins to Liver Injury

The liver serves as the key organ for the removal and detoxification of bacterial endotoxins that are continously absorbed in small amounts from the gastrointestinal tract. This paper postulates that liver injury impairs this detoxification process leading to further liver damage and systemic effects as well. Evidence is reviewed to support the contention that endotoxin may be a major common pathway for liver injury by a variety of agents, and methods of reducing endotoxicity of gut origin are proposed. Finally, a new solid phase radioimmunometric assay for E. coli 026 is described and its usefulness as a gut marker suggested.     

A potential role of GW4064 to inhibit gut bacterial overgrowth by activating FXR in suppression of ethanol-induced liver injury

Hepatic parenchymal and nonparenchymal cells are highly susceptible to ethanol and its metabolites, and excessive alcohol consumption results in damage to the liver. Ethanol induces an increased prevalence for bacterial overgrowth in the small intestine and translocation of endotoxin into the portal blood. Some studies have pointed to a role for activation of Kupffer cells by gut bacteria-derived endotoxin as a primary event in mechanisms of alcoholic liver injury (ALD). GW4064, a potent farnesoid X receptor (FXR) agonist, has been developed as a hepatoprotective agent, and has been used in animal models of a variety of liver diseases. At the same time, previous experimental results showed that BAs and GW4064 inhibit bacterial overgrowth in the small intestine. It is logical to postulate that GW4064 may control or alleviate the ethanol-induced liver injury through inhibiting gut bacterial overgrowth. GW4064 activates FXR, which induces the expression of several genes with potential functions in mucosal defense to prevent intestinal bacteria overgrowth and translocation into the circulation induced by ethanol, and then will alleviate ethanol-induced liver injury. The hypothesis will provide the brand-new direction that we may prevent and treat ALD by using GW4064 through activating FXR to control gut bacteria overgrowth.     

Endotoxin-induced ATP depletion in thyrotoxic rats

Effect of endotoxin from E. coli on the ATP content in heart muscle, the liver and the kidney of thyrotoxic rats was studied. When endotoxin (200-400 micrograms) was intravenously injected to rats taking drinking water containing 2-7.5 micrograms T3 per ml, body temperature rose and the heart rate increased. At the same time, a marked decrease in the ATP content in heart muscle and the kidney was observed together with an increase in Na+-K+-ATPase activity. Such changes were not observed or seen only to a small extent in euthyroid rats after endotoxin administration. Endotoxin-induced ATP depletion in T3-treated rats was prevented by administration of 5 mg hydrocortisone just prior to endotoxin injection. These findings indicate that endotoxin easily causes ATP depletion in some tissue or organs in thyrotoxicosis, even if the dose of endotoxin is not enough to produce such an effect in the euthyroid. These observations are of interest in relation to thyroid storm associated with bacterial infection.     

Effect of superoxide dismutase,allopurinol and glucocorticoids on liver and lung metallothionein induction by endotoxin in the rat

Liver and lung metallothionein (MT) levels were increased by endotoxin. The administration of superoxide dismutase (SOD) or allopurinol (ALLO) before (30–60 min) or after (24–32 h) the endotoxin treatment either increased or did not affect the effect of endotoxin on MT levels, depending on the particular treatment and tissue. SOD and ALLO also increased liver and lung MT levels in control rats. In contrast, liver MT levels tended to be decreased by the glucocorticoid prednisolone (PRED) when administered before the endotoxin and were significantly decreased when it was administered after endotoxin. The effect of PRED on lung MT levels was completely different, since it decreased the effect of endotoxin when injected before the lipopolysaccharide, but increased it when injected after the endotoxin. Liver lipid peroxidation, as measured by thiobarbituric acid reactants (TBARs), increased after endotoxin in the liver but not in the lung, an effect even potentiated in some cases by the antioxidants studied. As expected, tissue MT and TBARs could not be correlated.     

The effect of taurine on mesenteric blood flow and organ injury in sepsis

Endotoxin decreases mesenteric blood flow and inflicts organ injury via free radicals. We investigated whether taurine, an endogenous antioxidant and vasodilator, could attenuate the deleterious effects of endotoxin in a mouse model of sepsis. Swiss albino mice were allocated into four groups and treated either with taurine (150 mg/kg, i.p. at 0(th), 8(th), 16(th) h) or its solvent sterile saline (NaCl 0.9%, w/v) while E. coli endotoxin (20 mg/kg, i.p.) or its solvent saline were also given at 8(th) h. At 24(th) h the animals were anaesthetized and the mesenteric blood flow was measured by using perivascular ultrasonic Doppler-flowmeter. The animals were then exsanguinated, the spleen, liver, and kidneys were isolated for histopathological examination. Thiobarbituric acid-reacting substances (TBARS), glutathione, and myeloperoxidase activity were determined in the liver samples. Endotoxin significantly decreased the mesenteric blood flow and glutathione levels in liver while TBARS and myeloperoxidase activity were increased. However, taurine did not block the deleterious effects of endotoxin nor it did attenuate the histopathological injury. Therefore, we concluded that endotoxin-induced organ injury via free radicals is resistant to blockade by taurine.     

Rapid Determination of Bacteriological Water Quality by Using Limulus Lysate

The Limulus lysate assay was used to measure the endotoxin content in stream water and was found to reflect the degree of bacterial contamination as measured by coliform, enteric, gram-negative, and heterotrophic bacteria. The firm-clot method was found to be a less sensitive and reproducible technique for the detection of endotoxin than was the spectrophotometric modification of the Limulus lysate assay. Bound endotoxin, as determined by the spectrophotometric modification of the Limulus lysate assay, was found to be a better measure of the endotoxin associated with bacterial cells than was total endotoxin.     

The effect of bacterial endotoxin on synthesis of (Cu,Zn)superoxide dismutase in lungs of oxygen-exposed rats

Administration of bacterial endotoxin to rats exposed to greater than 95% O2 results in increased lung superoxide dismutase activity, decreased O2-induced lung damage, and a 3- to 4-fold improvement in survival rate (Frank, L., Yam, J., and Roberts, R. J. (1978) J. Clin. Invest, 61, 269-275). Antibodies to rat liver (Cu,Zn) superoxide dismutase were prepared and utilized to investigate the mechanism by which endotoxin treatment leads to increased lung superoxide dismutase activity. Assay of enzyme activity and of immunodetectable enzyme showed that the increased activity is due to an increase in the number of enzyme molecules rather than activation of existing enzyme. Compared to air controls, lung slices from rats exposed to greater than 95% O2 and treated with endotoxin have elevated rats of synthesis of (Cu,Zn)superoxide dismutase (51%) and of total protein (100%). Lung slices from untreated rats exposed to greater than 95% O2 have no such elevations. Endotoxin treatment thus appears to stimulate lung protein synthesis, leading to greater (Cu,Zn)superoxide dismutase activity due to an increased number of enzyme molecules.     

Sequential hydration-dehydration studies of lysozyme by the thermally stimulated depolarization currents (TSDC) technique.

To investigate the physical state of water in hydrating biological macro-molecules, the dielectric properties of water in hen egg lysozyme pellets with various moisture contents were studied using the thermally stimulated depolarisation currents technique. The water dipoles appeared to be directly involved in the relaxation processes, such that, by increasing the content of water of sorption from ho = 0.075 to ho = 0.29, the current density recorded increased abruptly at moisture content above 0.075. At a fixed starting hydration level, the time evolution of water content was also studied by isothermal sample aging in dynamic vacuum: the TSDC spectra changed in both intensity and position of their main peaks (TM = 245 K, 190 K, 150 K) with moisture content and showed hysteresis. The complex behaviour of the TSDC response can be compared with the results obtained with the same technique on other biological macromolecules and suggests possible models for water configurations and rearrangements.     

Protective effect of bile acids on the onset of fructose-induced hepatic steatosis in mice

Fructose intake is being discussed as a key dietary factor in the development of nonalcoholic fatty liver disease (NAFLD). Bile acids have been shown to modulate energy metabolism. We tested the effects of bile acids on fructose-induced hepatic steatosis. In C57BL/6J mice treated with a combination of chenodeoxycholic acid and cholic acid (100 mg/kg body weight each) while drinking water or a 30% fructose solution for eight weeks and appropriate controls, markers of hepatic steatosis, portal endotoxin levels, and markers of hepatic lipogenesis were determined. In mice concomitantly treated with bile acids, the onset of fructose-induced hepatic steatosis was markedly attenuated compared to mice only fed fructose. The protective effects of the bile acid treatment were associated with a downregulation of tumor necrosis factor (TNF)α, sterol regulatory element-binding protein (SREBP)1, FAS mRNA expression, and lipid peroxidation in the liver, whereas hepatic farnesoid X receptor (FXR) or short heterodimer partner (SHP) protein concentration did not differ between groups fed fructose. Rather, bile acid treatment normalized occludin protein concentration in the duodenum, portal endotoxin levels, and markers of Kupffer cell activation to the level of water controls. Taken together, these data suggest that bile acids prevent fructose-induced hepatic steatosis in mice through mechanisms involving protection against the fructose-induced translocation of intestinal bacterial endotoxin.     

Local and systemic autonomic nervous effects on cell migration to the spleen.

This work is based on the hypothesis that sympathetic nerves regulate the uptake of circulating cells by the spleen by affecting splenic blood flow and that the quantity of cells sequestered depends on whether changes in noradrenergic transmission occur at local or systemic levels. Fluorescently labeled lymphoid cells were injected into rats, and organ blood flow was measured by the microsphere method. Increased retention of cells in the spleen paralleled by increased blood flow was detected after local denervation of this organ or administration of bacterial endotoxin. A comparable enhanced splenic blood flow was observed after general sympathectomy. However, the redistribution of blood perfusion during general vasodilatation resulted in deviation of leukocyte flow from the spleen, thus resulting in reduced uptake of cells by this organ. These results indicate that, although the uptake of cells by the spleen depends on arterial blood supply, enhanced perfusion does not always result in increased cell sequestration because general vasodilatation reduces cell uptake by this organ and even overrides stimulatory effects of endotoxin.     

The role of macrophages in the uptake of endotoxin by the mouse liver.

The purpose of this investigation was to analyse the macrophage subpopulations involved in the uptake of endotoxin in the liver. The results show that in normal B10.D2 mice the liver macrophages constitute a heterogeneous population of cells which, depending on their state of differentiation, are distinguished by their differential distribution in the liver acinus and by their ability to phagocytose latex. Following the intravenous administration of endotoxin (lipopolysaccharide = LPS) from Salmonella abortus equi, endotoxin-carrying non-parenchymal cells of the liver (NPLC) were investigated immunohistochemically (in situ) and immunocytochemically (after isolation) between 1 h and 14 days after the injection. The endotoxin content of the blood and of isolated NPLC was also determined, using radioactivity labeled LPS. Following LPS injection, the total number of macrophages in the liver increased, reaching a maximum after 3 days. There was a striking increase in the ratio of mature to immature macrophages. After day 3, the number of macrophages decreased again, returning to the pre-injection values by day 14. 1 h after the administration of LPS, 41% of the isolated NPLC were already endotoxin-positive, a percentage which remained constant until the 3rd day. Thereafter, the number of LPS-bearing cells increased to a maximum of about 52% on the 5th day. This increase mostly involved macrophages which had taken up endotoxin. Concurrent with these changes there was a threefold increase in radioactivity-labeled LPS from the 7th h to the 5th day after injection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of bacterial endotoxin on migration of gonadotropin-releasing, hormone producing neurons in rat embryogenesis

The effect of bacterial lipopolysaccharide endotoxin (LPS), immune system activator, on differentiation and migration of gonadotropin-releasing, hormone producing neurons in rat embryogenesis has been studied. Intraperitoneal introduction of LPS (18 jg/kg) to pregnant rats on the 12th day of pregnancy led to 50% decrease in total number of GRH-neurons in the forebrain of 17-day-old embryos and 17% decrease in 19-day-old embryos. At the same time, the number of GRH-neurons in the nasal area of the head of 17- and 19-day-old embryos increased by 40 and 50%, respectively, whereas it increased by 20% in olfactory bulbs of 17-day-old embryos and did not changed in olfactory bulbs of 19-day-old embryos. Neither the total number of neurons nor their distribution patterns were affected by the introduction of LPS into pregnant rats on the 15th day of pregnancy. Singular localization of GRH-neurons in embryo forebrain was observed after LPS administration, whereas the neurons were located by groups of 3-4 cells in rostral areas. Therefore, at the early stages of pregnancy, LPS was shown to suppress initial stages of differentiation and migration of GRH producing neurons. The effects observed in our study may be mediated by LPS-induced, proinflammatory cytokines.     

Acute Binge Drinking Increases Serum Endotoxin and Bacterial DNA Levels in Healthy Individuals

Binge drinking, the most common form of alcohol consumption, is associated with increased mortality and morbidity; yet, its biological consequences are poorly defined. Previous studies demonstrated that chronic alcohol use results in increased gut permeability and increased serum endotoxin levels that contribute to many of the biological effects of chronic alcohol, including alcoholic liver disease. In this study, we evaluated the effects of acute binge drinking in healthy adults on serum endotoxin levels. We found that acute alcohol binge resulted in a rapid increase in serum endotoxin and 16S rDNA, a marker of bacterial translocation from the gut. Compared to men, women had higher blood alcohol and circulating endotoxin levels. In addition, alcohol binge caused a prolonged increase in acute phase protein levels in the systemic circulation. The biological significance of the in vivo endotoxin elevation was underscored by increased levels of inflammatory cytokines, TNFα and IL-6, and chemokine, MCP-1, measured in total blood after in vitro lipopolysaccharide stimulation. Our findings indicate that even a single alcohol binge results in increased serum endotoxin levels likely due to translocation of gut bacterial products and disturbs innate immune responses that can contribute to the deleterious effects of binge drinking.     

华支睾吸虫感染对大鼠肠道菌群移位和内毒素血症的影响

目的研究肠道细菌移位在华支睾吸虫病致病机制中的作用。方法建立华支睾吸虫感染大鼠模型。分别在造模后48 h(后尾蚴期)、18 d(童虫期)和35 d(成虫期),取肝、肺、淋巴结和血液组织,采用平板培养法进行细菌移位的检测;采用鲎三肽基质染色定量法检测血浆内毒素含量。结果感染18 d后,实验组肠道细菌移位率开始增高,至感染35 d时,细菌移位率为70%,明显高于对照组的10%,差异有统计学意义(P=0.0230.05);感染鼠以童虫期、成虫期细菌移位现象明显,总移位率为65%,与对照组10%比较,差异有统计学意义(u=3.59,P0.01),且在肝、肺、淋巴结和血液组织中,移位发生率分别为60%、15%、25%和10%,以肝脏部位最高;造模后18 d血浆内毒素水平明显增高,造模后35 d血浆LPS水平略有下降,但仍明显高于对照组,差异有统计学意义(t=7.612,P0.01)。结论华支睾吸虫感染可引发宿主肠道菌群移位,以肝组织多发,从而参与致病机制。     

Endotoxin stimulates glycogenolysis in the liver by means of intercellular communication

Escherichia coli endotoxin (lipopolysaccharide) was shown to increase glycogenolysis in the perfused liver 2-3-fold. In isolated parenchymal liver cells, however, endotoxin did not influence glycogenolysis, whereas stimulation by endotoxin of glycogenolysis in the perfused liver could be blocked by aspirin. This suggests that the effect of endotoxin on liver glycogenolysis is mediated by eicosanoids. The amount of prostaglandin D2 (which is the major prostanoid formed by Kupffer cells) in the liver perfusates was increased 5-fold upon endotoxin addition, with a time course which preceded the increase in glucose output. It is concluded that endotoxin stimulates glycogenolysis in the liver by stimulating prostaglandin D2 release from Kupffer cells, with a subsequent activation of glycogenolysis in parenchymal liver cells. This mechanism of intercellular communication may be designed to provide the carbohydrate source of energy necessary for the effective destruction of invaded microorganisms, by phagocytic cells, including the Kupffer cells.     

Neutrophil adhesion is impaired in the mesentery but not in the liver sinusoids of portal hypertensive rats

Altered leukocyte/cytokine response to inflammation has been observed in human and experimental portal hypertension. The aim of this study was to characterize leukocyte adhesion in portal hypertensive (PPVL) rats stimulated with endotoxin. Leukocyte rolling, adhesion, and migration assessed by intravital microscopy were impaired in mesenteric venules after lipopolysaccharide administration (150 microg/kg) in PPVL vs. sham-operated rats. Analysis of leukocyte L-selectin expression and soluble L-selectin showed that this defective adhesion was related to increased L-selectin shedding. In vitro experiments using isolated leukocytes treated with phorbol 12-myristate 13-acetate showed that monocytes and neutrophils but not lymphocytes were hyperreactive to cell activation, as measured by CD11b overexpression and increased L-selectin shedding in PPVL rats. However, neutrophil emigration in liver sinusoids and in the lung 3 h after endotoxin injection were similar in both groups of animals. Thus the alterations in leukocyte activation and adhesion molecule expression observed in this study may contribute to a better understanding of the higher susceptibility and severity of bacterial infections in cirrhotic patients with portal hypertension.     

Accumulation of activated lymphocytes in liver of endotoxin treated rabbits.

Intravenous administration of endotoxin in doses, 1.0 mg/kg, 0.1 mg/kg or 0.01 mg/kg caused activation (in the sense of nucleolar RNA synthesis) of the lymphocytes in peripheral blood, kidney and liver in rabbits. The evaluation of the ratio of lymphocyte count to the number of organ tissue cells leads to the conclusion that the accumulation of endotoxin activated lymphocytes occurs in liver.