Influence of splanchnic vascular infusion on the content of endotoxins in plasma and the translocation of intestinal bacteria in rats with acute hemorrhage necrosis pancreatitis

The main reason for the death of the patient with acute hemorrhage necrosis pancreatitis (AHNP) is pancreatic infection and multi-organ failure caused by endotoxemia and intestinal bacterial translocation[1-7]. However, the pathogenesis of endotoxemia and intestinal bacterial translocation remains a question[8-10]; moreover, no effective method of prevention and cure for it has been found till now[11 -15] In the present study, we infused low dose dopamine and low molecular weight dextran through the catheters to abdominal aorta and portal vein, and observed its influence on the endotoxin concentration in plasma and the rate of translocation of intestinal bacteria in AHNP rats.

Endotoxins in the prostatic secretions of chronic prostatitis patients:a need for further biomarkers through the use of proteomics

Dear Sir,Dai et al.[1] must be commended on their useful investigation of the clinical significance of endotoxins in the expressed prostatic secretions （EPS） of chronic prostatitis （CP） patients. However, we take issue with their conclusion. The conclusion that EPS endotoxin determination is helpful in diagnostic confirmation is plausible,

Endotoxins enhance hepatocarcinogenesis induced by oral intake of thioacetamide in rats

AIM To clarify whether endotoxin is of pathogenic importance for hepatocarcinogenesis, or the increased cancer risk results solely from the cirrhotic process. METHODS The rat model of hepatoma was treated by the intake of 0 03% thioacetamide in drinking water for six months. During induction of hepatoma, rats were additionally treated with splenectomy and/or lipopolysaccharide administration. The liver nuclear DNA index and proliferation index were quantitatively analyzed by flow cytometry. Hepatic histology was examined with light and electron microscopes. Plasmic endotoxin concentration and γ glutamyl transpeptidase activity were measured, and hepatoma incidence was recorded. RESULTS Thioacetamide induced cirrhosis and hepatoma in Wistar rats with histology or regenerative nodule, fibrosis and neoplastic foci were quite similar to the pathogenic process of human cirrhosis leading to hepatoma. In comparison with TAA controls (DNA index: 1 15±0 21), exo endotoxin increased the DNA index by 7 8% (1 24±0 25, P <0 02) and hepatoma rate by 16 7. Splenectomy induced enteric endotoxemia increased the DNA index by 25% (1 44±0 15, P <0 01) and hepatoma rate by 33%. A summation of the effects of these two factors increased the DNA index by 36% ( P <0 01)and hepatoma incidence by 50%, moreover, the level of endotoxemia showed a close relation with DNA index ( r =0 96, P <0 01), as well as with the occurrence rate of hepatoma ( r =0 00, P <0 01). Histological findings further verified such alterations. CONCLUSION Lipopolysaccharide administration and/or splenectomy induced enterogenic endotoxemia may enhance rat hepatocarcinogenesis induced by oral intake of thioacetamide.

The Antagonistic Action of Heat-Clearing and Detoxifying Chinese Drugs on Endotoxins

In the recent decade, interest in treatment and prevention of many critical, severe and acute diseases caused by bacterial endotoxins has been aroused along with the advance of the knowledge on the nature of the endotoxin and the conditions involved. In abroad, attention has been mainly payed to raising antisera and monocolonal antibodies against the endotoxin and the induced mediators. However, the allergic reactions and the cost are still the problems. Till now, there is no drug that can antagonize endotoxin with high effectiveness and low toxicity. Clinical treatments are still confined in inhibiting or killing the pathogen, and correcting the internal environmental disturbance. Being less toxic and rich in resources with low cost and less side-effects, screening of effective Chinese drugs for antagonizing endotoxin is of important and practical significance. Endotoxin belongs to the category of toxic evils, or more precisely, the heat toxin in TCM. Therefore the application of heat-clearing and detoxifying Chinese drugs to antagonizing endotoxins is consistent with the theory of TCM. Some achievements in this field are reported as follows.

Endotoxins in the prostatic secretions of chronic prostatitis patients

Aim: To evaluate the clinical significance of the quantitative determinations of endotoxins in the expressed prostatic secretions (EPS) of chronic prostatitis (CP) patients. Methods: The EPS of 45 patients with CP and 15 normal volunteers were obtained for microscopic examination, bacterial culture and endotoxin determination. The level of endotoxins was determined by the Limulus-amebocyte-lysate test with chromogenic substrate. Results: Patients with CP had higher mean levels of endotoxins in EPS than normal volunteers [52.06 ± 32.83 EU.L^(-1) vs. 4.77 ± 4.14 EU'L^(-1) (P < 0.05)]. The levels of endotoxins in CP type Ⅱ, type Ⅲa and type Ⅲb were 68.62 ± 34.78 EU.L^(-1), 45.30± 23.33 EU.L^(-1) and 15.83 ± 5.31 EU·L^(-1), respectively [type Ⅱ vs. type Ⅲa (P > 0.05), type Ⅲb vs. normal controls (P > 0.05), type Ⅱ/type Ⅲa vs. normal controls P < 0.05)]. Conclusion: CP patients have elevated levels of endotoxins in the EPS, which suggests that inflammation is a feature of this disease. EPS endotoxin determination is not only helpful in diagnostic confirmation, but also in evaluating the response to treatment in CP patients.

Comparison of result judgment algorithm of test for interfering factors in the bacterial endotoxins test among Chinese, Japanese, European, American, and Indian pharmacopeias

Background The bacterial endotoxins test （BET） is a method used to detect or quantify endotoxins （lipo-polysaccharide,LPS） and is widely used in the quality control of parenteral medicines/vaccines and clinical dialysis fluid.It is also used in the diagnosis of endotoxemia and in detection of environment air quality control.Although BET has been adopted by most pharmacopoeias,result judgment algorithms （RJAs） of the test for interfering factors in the BET still differ between certain pharmacopoeias.We have evaluated RJAs of the test for interfering factors for the revision of BET described in the Chinese Pharmacopoeia 2010 （CHP2010）.Methods Original data from 1 748 samples were judged by RJAs of the Chinese Pharmacopoeia 2010,the Japanese Pharmacopoeia 2011 （JP2011）,the European Pharmacopoeia 7.0 （EP7.0）,the United States Pharmacopoeia 36 （USP36）,and the Indian Pharmacopoeia 2010 （IP2010）,respectively.A SAS software package was used in the statistical analysis.Results The results using CHP2010 and USP36,JP2011,EP7.0,and IP2010 had no significant difference （P=-0.7740）.The results using CHP2010 of 1 748 samples showed that 132 samples （7.6%） required an additional step; nevertheless there was no such requirement when using the other pharmacopeias.The kappa value of two RJAs （CHP2010 and EP7.0） was 0.6900 （0.6297-0.7504） indicating that the CHP2010 and other pharmacopoeias have good consistency.Conclusions The results using CHP2010 and USP36,JP2011,EP7.0,and IP2010 have different characteristics.CHP2010 method shows a good performance in Specificity,mistake diagnostic rate,agreement rate,predictive value for suspicious rate,and predictive value for passed rate.The CHP2010 method only had disadvantages in sensitivity compared with other pharmacopeias.We suggest that the Chinese pharmacopoeia interference test be revised in accordance with the USP36,JP2011,EP7.0,and IP2010 judgment model.

Five simple models for interfering factors test of bacterial endotoxins test

Endotoxins have been credited for over 50% of sepsis cases, with significantly greater mortality.1 A literature indicates wide-spread agreement that early detection of endotoxemia, endotoxin in the bloodstream, is the major key for patient survival from sepsis.2 Today the most popular endotoxin detection system is bacterial endotoxins test （BET）, adopted by most pharmacopoeias. Interference test is a part of the bacterial endotoxin inspection method, used to judge whether the sample can be applied in BET. However, Limuloid resources are exhausted in China, which is an important source for LAL. Here, we reported 5 simple models for interfering factors test in the BET, and compared new models with the United States Pharmacopoeia 36 （USP36）.

Endotoxin-induced alterations of adipose tissue function:a pathway to bovine metabolic stress

During the periparturient period, dairy cows exhibit negative energy balance due to limited appetite and increased energy requirements for lactogenesis. The delicate equilibrium between energy availability and expenditure puts cows in a state of metabolic stress characterized by excessive lipolysis in white adipose tissues(AT), increased production of reactive oxygen species, and immune cell dysfunction. Metabolic stress, especially in AT, increases the risk for metabolic and inflammatory diseases. Around parturition, cows are also susceptible to endotoxemia. Bacterial-derived toxins cause endotoxemia by promoting inflammatory processes and immune cell infiltration in different organs and systems while impacting metabolic function by altering lipolysis, mitochondrial activity, and insulin sensitivity. In dairy cows, endotoxins enter the bloodstream after overcoming the defense mechanisms of the epithelial barriers, particularly during common periparturient conditions such as mastitis, metritis, and pneumonia, or after abrupt changes in the gut microbiome. In the bovine AT, endotoxins induce a pro-inflammatory response and stimulate lipolysis in AT, leading to the release of free fatty acids into the bloodstream. When excessive and protracted, endotoxin-induced lipolysis can impair adipocyte's insulin signaling pathways and lipid synthesis. Endotoxin exposure can also induce oxidative stress in AT through the production of reactive oxygen species by inflammatory cells and other cellular components. This review provides insights into endotoxins' impact on AT function, highlighting the gaps in our knowledge of the mechanisms underlying AT dysfunction, its connection with periparturient cows' disease risk, and the need to develop effective interventions to prevent and treat endotoxemia-related inflammatory conditions in dairy cattle.

A new method for concentration analysis of bacterial endotoxins in perfluorocarbon

This communication demonstrates the feasibility of the gel-clot method for the analysis of bacterial endotoxins in water extracts of perfluorocarbon which is a water insoluble liquid medical device. Perfluorocarbon （10 mL） was shaken with 10 mL water for 15 min at 2000 r/min and the endotoxin present was extracted to the aqueous phase without interference inhibition/enhancement of the product and the recovery of endotoxin added to perfluorocarbon was determined, A validation study confirmed that endotoxins presented in perfluorocarbon pass over into the aqueous phase at concentrations of 20, 10 and 5EU/mL with recoveries from 86.8% to 96.8%. Therefore, the gel-clot test is suitable for detecting bacterial endotoxins in perfluorocarbon which is a water insoluble medical device.

Value of combining the serum D-lactate, diamine oxidase, and endotoxin levels to predict gut-derived infections in cancer patients

Objective:This is a retrospective observational cohort study.The objective of this retrospective observational cohort study was to evaluate the value of the combined serum D-lactic acid,diamine oxidase(DAO),and endotoxin levels to predict intestinal barrier impairment and gut-derived infection(GDI)in cancer patients.Methods:Cancer patients receiving chemotherapy or palliative care treatment at our hospital were enrolled in the study.The serum concentrations of DAO,D-lactic acid,and endotoxin were determined using the intestinal barrier function biochemical index analysis system.The patients'infection information came from the hospital's Medicom Prescription Automatic Screening System and themedical records.Three hundred fifty-three cancer patients were included in the study(53.8%female,73.7%cancer stage IV,27.8%had bowel obstruction).Results:The total incidence of GDI was 33.4%(118/353).The median length of hospital stay was 16 days for patients with GDI,compared with 7 days for patients without GDI(P<0.001).The media hospitalization costs were¥27,362.35 for patients with GDI compared with¥11,614.08 for patients without GDI(P<0.001).The serum concentrations of DAO,D-lactic acid,and endotoxin were significantly higher in patients with GDI.As malignant bowel obstruction(MBO)worsened,the concentrations of DAO,D-lactic acid,and endotoxin increased.Multivariate logistic regression models revealed that the DAO,endotoxin,IL-6,and C-reactive protein levels were significantly associated with an increased risk of GDI.In addition,we also found a fivefold increased risk of infection in patients withMBO compared with those without bowel obstruction(OR=6.210,P<0.001).All of the areas under the receiver operating characteristic curve(AUCs)for DAO,D-lactate,and endotoxin to predict GDI were<0.7(AUC=0.648,P<0.001;AUC=0.624,P<0.01;AUC=0.620,P<0.01,respectively).However,when the parameters were combined(DAO+D-lactate+endotoxin),the predictive power increased significantly(AUC=0.797,P<0.001).Moreover,combining these intestinal barrier indicators and the presence of MBO had better power to predict GDI than either alone(AUC=0.837,P<0.001).Conclusions:Combining the serum DAO,D-lactic acid,and endotoxin levels was a better predictor of GDI than any of the indicators alone,and combining these with the diagnosis of MBO could further improve the efficacy for predicting GDI.

Effect of hepatocyte apoptosis induced by TNF-α on acute severe hepatitis in mouse models

AIM To study the effect of hepatocyteapoptosis and necrosis induced by TNF-α on thepathogenesis of acute severe hepatitis(ASH).METHODS The model of ASH was prepared inD-galactosamine(GAIN)sensitized BALB/c miceby injection of either endotoxin(ET)or tumornecrosis factor-α(TNF-α).Morphologicalchanges of apoptotic hepatocytes were studiedby both light and electron microscope and in siteend labeling method(ISEL).Molecular biologicalchanges of DNA ladder were observed byelectrophoresis of extract from liver tissues.Biochemical changes were measured by alanineaminotransferase(ALT),asparticaminotransferase(AST)and TNF-α.The relationbetween apoptosis and necrosis was evaluatedsimultaneously.RESULTS The sequence of hepatocyteapoptosis,necrosis,and final death from ASHwas observed both in GAIN/ET and GAIN/TNF-agroup.Apoptosis was prominent at 3.5 h and 5 hafter injection of inducer,while necrosis becamedominant at 9 h after challenge.The appearanceof apoptosis was earlier in GAIN/TNF-α groupthan that in GAIN/ ET group.Pretreatment ofmice with antiTNF IgG1 may completely preventthe liver injury induced by GalN/ET.CONCLUSION TNF-α can cause liver damageby inducing hepatic apoptosis and necrosis inmice with endotoxemia.

Diet high in fructose leads to an overexpression of lipocalin-2 in rat fatty liver

AIM: To explore lipocalin-2 (LCN-2) expression and its possible role and mechanism(s) of production in rat models of diet-inducible fatty liver.

Possible relationship between intestinal barrier function and formation of pigment gallstones in hamsters

BACKGROUND: The presence of bacteria in bile is an important factor in the formation of pigment gallstones. The bile of healthy people is sterile and bacteria in the biliary system come from endogenous infection from the gut. Yet, the route of bacterial translocation into the bile duct is still unclear. Theoretically, two routes exist: one is through the intestinal barrier and the other is by direct reflux from the sphincter of Oddi. This study was undertaken to explore the relationship between the effectiveness of intestinal barrier and the formation of pigment gallstones in hamsters. METHODS: Thirty-two hamsters were divided into an experimental and a control group, with 16 hamsters in each group. A low protein and high cellulose diet was given for 6 weeks to induce the formation of pigment gallstones in the experimental group (PS) and a normal diet was given to the control group (CON). Morphological changes, changes in the levels of serum endotoxin and diamine oxidase, and changes in the numbers of B lymphocytes, plasma cells and secretory immunoglobin A (sIgA) in the intestinal mucosa were assessed after 6 weeks. RESULTS: Four hamsters died during lithogenesis and body weight decreased in the PS group. Pigment gallstones were found in 11 hamsters at the end of the experiment, giving a lithogenesis rate of 91.67%. The serum endotoxin level before and after gallstone formation in the PS group was 0.2960 +/- 0.1734 U/ml and 8.2964 +/- 4.6268 U/ml, respectively (P<0.05). The blood diamine oxidase level before and after gallstone formation in the PS group was 2.6333 +/- 0.8037 U/ml and 3.3642 +/- 0.9545 U/ml, respectively (P<0.05). The numbers of B lymphocytes, plasma cells and sIgA in the intestinal mucosa in the PS group were 71.56 +/- 2.89, 68.65 +/- 2.09 and 27.56 +/- 1.07, respectively, and were significantly decreased compared with the corresponding values in the CON group (94.25 +/- 3.69, 93.47 +/- 3.98 and 42.57 +/- 1.96, respectively, P<0.05). CONCLUSIONS: A low protein and high cellulose diet can markedly reduce intestinal barrier function and facilitate the formation of pigment gallstones. The decrease of intestinal barrier function may take part in the formation of pigment gallstones.

Innate immune reactivity of the liver in rats fed a choline-deficient L-amino-acid-defined diet

AIM： To investigate the innate immune reactivity of tumor necrosis factor-alpha （TNF-α）, Toll-like receptor 4 （TLR4）, and CD14 in the liver of non-alcoholic steatohepatitis （NASH） model rats. METHODS： Male F344 rats were fed a cholinedeficient L-amino-acid-defined （CDAA） diet. The rats were killed after 4 or 8 wk of the diet, and their livers were removed for immunohistochemical investigation and RNA extraction. The liver specimens were immunostained for TNF-α, TLR4, and CD14. The gene expressions of TNF-α, TLR4, and CD14 were determined by reverse-transcriptase polymerase chain reaction （RT-PCR）. Kupffer cells were isolated from the liver by Percoll gradient centrifugation, and were then cultured to measure TNF-α production. RESULTS： The serum and liver levels of TNF-~ in the CDAA-fed rats increased significantly as compared with the control group, as did the immunohistochemical values and gene expressions of TNF-α, TLR4, and CD14 with the progression of steatohepatitis. TNF-α production from the isolated Kupffer cells of the CDAAfed rats was elevated by lipopolysaccharide stimulation. CONCLUSION： The expressions of TNF-α, TLR4, and CD14 increased in the NASH model, suggesting that

Expression of TNF mRNA in the internal organs after severe burn injury in rats

Tumor necrosis factor(TNF) mRNA was determined with dot blotting in various viscera 24 h after severe burn injury in rats. It was found that TNF mRNA was detected in the liver, kidneys, spleen,lungs and small intestines in normal conditions. After burn in

Endotoxin receptor CD14 gene variants and histological features in chronic HCV infection

AIM:To analyze the correlation between CD14 rs2569190/C-159T single nucleotide polymorphism (SNP) and disease progression in chronic hepatitis C.METHODS: Liver biopsy specimens from a total of 137 and 349 patients with chronic hepatitis C were separately evaluated with respect to necroinflammatory activity (grading) and architectural changes (staging). In one group, further histological lesions characteristic for hepatitis C, hepatitis C virus subtypes, and biochemical parameters of liver disease were also investigated. Samples of genomic DNA were genotyped for the respective SNP by 5'-nuclease assays using fluorescent dye-labeled allele-specif ic probes.RESULTS: Genotype distribution did not deviate from the Hardy-Weinberg equilibrium. In the first group, patients homozygous for the variant allele T were found to be younger than C allele carriers (39.6±12.5 vs 45.7±11.5, P=0.008). Among the histological lesions studied, portal lymphoid aggregates were more frequently observed among TT homozygotes than among C carriers (21/37 vs 32/100, P=0.008). The presence of portal lymphoid aggregates was closely correlated with hepatic inflammation (P=0.003) and with bile duct damage (P<0.001). The degree of fibrosis, in contrast, was not found to be related to the CD14 gene C-159T polymorphism.CONCLUSION: The data suggest a possible relationship between CD14 C-159T polymorphism and the formation of portal lymphoid aggregates, but not liver fibrosis progression in chronic hepatitis C.

Potential roles of functional bacterial amyloid proteins,bacterial biosurfactants and other putative gut microbiota products in the etiopathogeny of Parkinson’s Disease

An increasing number of studies provide evidence for the existence of a microbiota-gut-brain axis and its potential involvement in the development of sporadic Parkinson’s disease and other neurodegenerative conditions.The neuropathologic hallmark of Parkinson’s disease is the presence of brain intraneuronal aggregates of misfolded alpha-synuclein,known as Lewy bodies.Some gut microbiota products may trigger alpha-synuclein conformational changes in the neurons of the enteric nervous system,which can then spread to the brain in a prion-like fashion through the vagus nerve.Others may interfere with neuroinflammatory pathways and susceptibility to neurodegeneration.In this review,we assess the potential role of putative gut microbiota products in the etiopathogeny of Parkinson’s disease,with a special emphasis on functional bacterial amyloid proteins,bacterial biosurfactants,endotoxins and short-chain fatty acids.The possible roles of molecular hydrogen,a common byproduct of bacterial fermentation,are also addressed.

African Dust Storms Reaching Puerto Rican Coast Stimulate the Secretion of IL-6 and IL-8 and Cause Cytotoxicity to Human Bronchial Epithelial Cells (BEAS-2B)

African dust storm events (ADE) travel across theAtlantic Ocean(ADEAO) and reach the Puerto Rican coast (ADEPRC), potentially impacting air quality and human health. To what extent seasonal variations in atmospheric particulate matter (PM) size fractions, composition and sources trigger respiratory-adverse effects to Puerto Ricans is still unclear. In the present study, we investigated the pro-inflammatory and cytotoxic effects of PM samples harvested during ADEAO (PM10), ADEPRC (PM2.5 and PM10) and Non-ADE (Pre-and Post-ADEAO and Non-ADEPRC), using BEAS-2B cells. Endotoxins (ENX) in PM2.5 and PM10 extracts and traces of metals (TMET) in PM2.5 extracts were also examined. IL-6 and IL-8 secretion and cytotoxicity were used as endpoints. ADEAO and ADEPRC extracts were found to be more cytotoxic than Non-ADE and ADEAO were more toxic than ADEPRC extracts. PM10 extracts from ADEAO and Post-ADEAO caused significant secretion of IL-8. IL-6 and IL-8 secretion was higher following treatment with PM10 and PM2.5 ADEPRC than with Non-ADEPRC extracts. ENX levels were found to be higher in PM10 ADEAO than in the rest of the samples tested. TMET levels were higher in PM2.5 ADEPRC than in Non-ADEPRC extracts. Deferoxamine significantly reduced cytotoxicity and IL-6 and IL-8 secretion whereas Polymyxin B did not. TMET in PM2.5 fractions is a major determinant in ADEPRC-induced toxicity and work in conjunction with ENX to cause toxicity to lung cells in vitro. ENX and TMET may be responsible, in part, for triggering PM-respiratory adverse responses in susceptible and predisposed individuals.

Lack of new antiinfective agents: Passing into the pre-antibiotic age?

The lack of newly developed antibiotics, together with the increase in multi-resistance of relevant pathogenic bacteria in the last decades, represents an alarming signal for human health care worldwide. The number of severely infected persons increases not only in developing but also in highly industrialized countries. This relates in first line to the most severe form of a bacterial infection, sepsis and the septic shock syndrome, with high mortality on critical care units. No particular anti-sepsis drug is available, and the therapy with conventional antibiotics more and more fails to provide a survival benefit. Due to the fact that the pharmaceutical industry has withdrawn to a high degree from the development of anti-infectious agents, a huge challenge for health care is approaching in the 21 st century. In this article, these problems are outlined and possible alternatives are presented which may be helpful to solve the problem.

Changes of mucosal permeability to lipopolysaccharide in the colon of chronic alcoholic rats

AIM To evaluate the effects of chronic alcohol abuse on the mucosal permeability to lipopolysaccharide in the colon of rats. METHODS Escherichia coli lipopolysaccharide (LPS, 20mg/L) was injected into the colon of chronic alcoholic rats ( n =10) which were supplied with Lieber diets every other day for 6 weeks. Before LPS injection and 5, 10, 20, 30 minutes after the injection, blood samples from the portal vein were obtained and contents of LPS in the blood were measured. The distribution of LPS in the colon tissues was observed with a confocal laser scanning microscope by immunofluorescent technique using the monoclonal antibody specific to the lipid A region of LPS. Normal rats were used as controls ( n =6). RESULTS Before LPS injection in the colon, LPS levels in the blood of portal vein of chronic alcoholic rats were significantly higher than those of normal controls (3 56ng/L±0 67ng/L, vs 2 45ng/L±0 15ng/L, P <0 01). At 5, 10, 20, 30 minutes after the injection of LPS, LPS contents were significantly higher than those before LPS injection (173 56ng/L±3 45ng/L, 154 78ng/L±0 57ng/L, 43 89ng/L±0 67ng/L, 45 38ng/L±0 89ng/L vs 3 56ng/L±0 67ng/L, P <0 01, respectively). Most mucosal cells showed strong positive reactions to LPS in the rats of chronic alcohol abuse, but no significant changes of LPS contents in blood from the portal vein and fluorescent reactions to LPS in mucosal cells of normal rats were found after LPS injection. CONCLUSION Chronic alcohol abuse resulted in a significant increase of permeability to LPS in colon mucosal cells in rats.