interleukin-8 and Escherichia-coli-Infections

Middle East Respiratory Syndrome (MERS) is a novel respiratory illness firstly reported in Saudi Arabia in 2012. It is caused by a new corona virus, called MERS corona virus (MERS-CoV). Most people who have MERS-CoV infection developed severe acute respiratory illness.. This work is done to determine the clinical characteristics and the outcome of intensive care unit (ICU) admitted patients with confirmed MERS-CoV infection.. This study included 32 laboratory confirmed MERS corona virus infected patients who were admitted into ICU. It included 20 (62.50%) males and 12 (37.50%) females. The mean age was 43.99 ± 13.03 years. Diagnosis was done by real-time reverse transcription polymerase chain reaction (rRT-PCR) test for corona virus on throat swab, sputum, tracheal aspirate, or bronchoalveolar lavage specimens. Clinical characteristics, co-morbidities and outcome were reported for all subjects.. Most MERS corona patients present with fever, cough, dyspnea, sore throat, runny nose and sputum. The presence of abdominal symptoms may indicate bad prognosis. Prolonged duration of symptoms before patients' hospitalization, prolonged duration of mechanical ventilation and hospital stay, bilateral radiological pulmonary infiltrates, and hypoxemic respiratory failure were found to be strong predictors of mortality in such patients. Also, old age, current smoking, smoking severity, presence of associated co-morbidities like obesity, diabetes mellitus, chronic heart diseases, COPD, malignancy, renal failure, renal transplantation and liver cirrhosis are associated with a poor outcome of ICU admitted MERS corona virus infected patients.. Plasma HO-1, ferritin, p21, and NQO1 were all elevated at baseline in CKD participants. Plasma HO-1 and urine NQO1 levels each inversely correlated with eGFR (. SnPP can be safely administered and, after its injection, the resulting changes in plasma HO-1, NQO1, ferritin, and p21 concentrations can provide information as to antioxidant gene responsiveness/reserves in subjects with and without kidney disease.. A Study with RBT-1, in Healthy Volunteers and Subjects with Stage 3-4 Chronic Kidney Disease, NCT0363002 and NCT03893799.. HFNC did not significantly modify work of breathing in healthy subjects. However, a significant reduction in the minute volume was achieved, capillary [Formula: see text] remaining constant, which suggests a reduction in dead-space ventilation with flows > 20 L/min. (ClinicalTrials.gov registration NCT02495675).. 3 组患者手术时间、术中显性失血量及术后 1 周血红蛋白下降量比较差异均无统计学意义（. 对于肥胖和超重的膝关节单间室骨关节炎患者，采用 UKA 术后可获满意短中期疗效，远期疗效尚需进一步随访观察。.. Decreased muscle strength was identified at both time points in patients with hEDS/HSD. The evolution of most muscle strength parameters over time did not significantly differ between groups. Future studies should focus on the effectiveness of different types of muscle training strategies in hEDS/HSD patients.. These findings support previous adverse findings of e-cigarette exposure on neurodevelopment in a mouse model and provide substantial evidence of persistent adverse behavioral and neuroimmunological consequences to adult offspring following maternal e-cigarette exposure during pregnancy. https://doi.org/10.1289/EHP6067.. This RCT directly compares a neoadjuvant chemotherapy regimen with a standard CROSS regimen in terms of overall survival for patients with locally advanced ESCC. The results of this RCT will provide an answer for the controversy regarding the survival benefits between the two treatment strategies.. NCT04138212, date of registration: October 24, 2019.. Results of current investigation indicated that milk type and post fermentation cooling patterns had a pronounced effect on antioxidant characteristics, fatty acid profile, lipid oxidation and textural characteristics of yoghurt. Buffalo milk based yoghurt had more fat, protein, higher antioxidant capacity and vitamin content. Antioxidant and sensory characteristics of T. If milk is exposed to excessive amounts of light, Vitamins B. The two concentration of ZnO nanoparticles in the ambient air produced two different outcomes. The lower concentration resulted in significant increases in Zn content of the liver while the higher concentration significantly increased Zn in the lungs (p < 0.05). Additionally, at the lower concentration, Zn content was found to be lower in brain tissue (p < 0.05). Using TEM/EDX we detected ZnO nanoparticles inside the cells in the lungs, kidney and liver. Inhaling ZnO NP at the higher concentration increased the levels of mRNA of the following genes in the lungs: Mt2 (2.56 fold), Slc30a1 (1.52 fold) and Slc30a5 (2.34 fold). At the lower ZnO nanoparticle concentration, only Slc30a7 mRNA levels in the lungs were up (1.74 fold). Thus the two air concentrations of ZnO nanoparticles produced distinct effects on the expression of the Zn-homeostasis related genes.. Until adverse health effects of ZnO nanoparticles deposited in organs such as lungs are further investigated and/or ruled out, the exposure to ZnO nanoparticles in aerosols should be avoided or minimised.

Topics: A549 Cells; Acetylmuramyl-Alanyl-Isoglutamine; Acinetobacter baumannii; Acute Lung Injury; Adaptor Proteins, Signal Transducing; Adenine; Adenocarcinoma; Adipogenesis; Administration, Cutaneous; Administration, Ophthalmic; Adolescent; Adsorption; Adult; Aeromonas hydrophila; Aerosols; Aged; Aged, 80 and over; Aging; Agriculture; Air Pollutants; Air Pollution; Airway Remodeling; Alanine Transaminase; Albuminuria; Aldehyde Dehydrogenase 1 Family; Algorithms; AlkB Homolog 2, Alpha-Ketoglutarate-Dependent Dioxygenase; Alzheimer Disease; Amino Acid Sequence; Ammonia; Ammonium Compounds; Anaerobiosis; Anesthetics, Dissociative; Anesthetics, Inhalation; Animals; Anti-Bacterial Agents; Anti-HIV Agents; Anti-Infective Agents; Anti-Inflammatory Agents; Antibiotics, Antineoplastic; Antibodies, Antineutrophil Cytoplasmic; Antibodies, Monoclonal, Humanized; Antifungal Agents; Antigens, Bacterial; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Antimetabolites, Antineoplastic; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Antioxidants; Antitubercular Agents; Antiviral Agents; Apolipoproteins E; Apoptosis; Arabidopsis; Arabidopsis Proteins; Arsenic; Arthritis, Rheumatoid; Asthma; Atherosclerosis; ATP-Dependent Proteases; Attitude of Health Personnel; Australia; Austria; Autophagy; Axitinib; Bacteria; Bacterial Outer Membrane Proteins; Bacterial Proteins; Bacterial Toxins; Bacterial Typing Techniques; Bariatric Surgery; Base Composition; Bayes Theorem; Benzoxazoles; Benzylamines; beta Catenin; Betacoronavirus; Betula; Binding Sites; Biological Availability; Biological Oxygen Demand Analysis; Biomarkers; Biomarkers, Tumor; Biopsy; Bioreactors; Biosensing Techniques; Birth Weight; Blindness; Blood Chemical Analysis; Blood Gas Analysis; Blood Glucose; Blood Pressure; Blood Pressure Monitoring, Ambulatory; Blood-Brain Barrier; Blotting, Western; Body Mass Index; Body Weight; Bone and Bones; Bone Density; Bone Resorption; Borates; Brain; Brain Infarction; Brain Injuries, Traumatic; 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Proto-Oncogene Proteins c-ret; Proto-Oncogene Proteins p21(ras); Proton Pumps; Protons; Protoporphyrins; Pseudomonas aeruginosa; Pseudomonas fluorescens; Pulmonary Artery; Pulmonary Disease, Chronic Obstructive; Pulmonary Gas Exchange; Pulmonary Veins; Pyrazoles; Pyridines; Pyrimidines; Qualitative Research; Quinoxalines; Rabbits; Random Allocation; Rats; Rats, Sprague-Dawley; Rats, Wistar; Receptors, Histamine H3; Receptors, Immunologic; Receptors, Transferrin; Recombinant Proteins; Recurrence; Reference Values; Referral and Consultation; Regional Blood Flow; Registries; Regulon; Renal Insufficiency, Chronic; Reperfusion Injury; Repressor Proteins; Reproducibility of Results; Republic of Korea; Research Design; Resistance Training; Respiration, Artificial; Respiratory Distress Syndrome; Respiratory Insufficiency; Resuscitation; Retinal Dehydrogenase; Retreatment; Retrospective Studies; Reverse Transcriptase Inhibitors; Rhinitis, Allergic; Ribosomal Proteins; Ribosomes; Risk Assessment; 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YAP-Signaling Proteins; Yogurt; Young Adult; Zebrafish; Zebrafish Proteins; Ziziphus

Enteroaggregative Escherichia coli (EAEC) is an increasingly recognized enteric pathogen. It is a cause of both acute and persistent diarrhoea among children, adults and HIV-infected persons, in both developing and developed countries. In challenge studies, EAEC has caused diarrhoeal illness with the ingestion of 10(10) c.f.u. Outbreaks of diarrhoeal illness due to EAEC have been reported, and linked to the ingestion of contaminated food. Diarrhoeal illness due to EAEC is the result of a complex pathogen-host interaction. Some infections due to EAEC result in diarrhoeal illness and elicit an inflammatory response, whereas other infections do not result in a symptomatic infection. Many putative virulence genes and EAEC strains that produce biofilm have been identified; however, the clinical significance of these genes and of biofilm production has yet to be defined. A -251 AA single nucleotide polymorphism (SNP) in the interleukin (IL)-8 promoter region is reported to increase host susceptibility to EAEC diarrhoea. Ciprofloxacin and rifaximin continue to be an effective treatment in persons infected with EAEC. This review is intended to provide an updated review for healthcare workers on EAEC, an emerging enteric pathogen.

Topics: Adult; Animals; Anti-Infective Agents; Antidiarrheals; Bacterial Adhesion; Biofilms; Child; Child, Preschool; Clinical Trials as Topic; Diarrhea; Disease Outbreaks; DNA, Bacterial; Escherichia coli; Escherichia coli Infections; Food Microbiology; Genes, Bacterial; Genetic Predisposition to Disease; Global Health; HIV Infections; Humans; Interleukin-8; Promoter Regions, Genetic; Virulence; Virulence Factors

The aim of this article is to review recent advances in travelers' diarrhea, which remains one of the most common health problems afflicting individuals from developed countries visiting less affluent regions of the world.. A large epidemiologic study done at the point of departure provided insights into regional risk factors for travelers' diarrhea and demonstrated that visitors rarely exercised dietary precautions aimed at disease prevention. A preventive program implemented with the close interaction between public health agencies, hotel industry and academia can result in effective reduction of cases. A polymorphism in the IL-8 gene promoter is associated with susceptibility to diarrhea due to enteroaggregative Escherichia coli. New diagnostic tools assist in better understanding the role of norovirus and emerging bacterial enteric pathogens such as enteroaggregative E. coli. Rifaximin, a non-absorbable antibiotic, is a safe and effective alternative for the prevention and treatment of travelers' diarrhea due to non-invasive organisms.. Traditional public health and new antimicrobial agents can decrease the risk of travel related diarrhea.

Topics: Anti-Bacterial Agents; Diarrhea; Escherichia coli; Escherichia coli Infections; Humans; Interleukin-8; Polymorphism, Genetic; Promoter Regions, Genetic; Rifamycins; Rifaximin; Travel

Topics: Animals; Bacterial Adhesion; Bacterial Infections; Diarrhea; Enterobacteriaceae; Escherichia coli; Escherichia coli Infections; Flagellin; Humans; Inflammation Mediators; Interleukin-8; Intestinal Diseases; Intestinal Mucosa; Models, Biological; Salmonella Infections; Salmonella typhimurium

Topics: Bacterial Toxins; Cholera; Cyclic AMP; Diarrhea; Enterocolitis, Pseudomembranous; Escherichia coli Infections; Guanylate Cyclase; Humans; Interleukin-8; Nutrition Disorders; Platelet Activating Factor; Population Growth; Signal Transduction

The common response to infection is infiltration of the affected tissue by inflammatory cells. It is now recognized that the epithelium plays a crucial role in this immunological process by producing an array of proinflammatory cytokines including interleukin-8, tumour necrosis factor-alpha, monocyte chemotactic protein-1, granulocyte-macrophage colony-stimulating factor, extractable nuclear antigen-78 and others. The response of the intestinal epithelium to bacterial pathogens is particularly intriguing because it is literally bathed by normal bacterial flora and bacterial components/products yet remains immunologically quiescent despite this potentially hostile environment. In contrast, when challenged by bacterial pathogens, intestinal epithelial cells exhibit a vigorous immunological response. Our laboratory has, therefore, focused on the immune response of intestinal epithelial cells when confronted by a specific bacterial pathogen, enteropathogenic Escherichia coli.

Topics: Antigens, Nuclear; Bacterial Adhesion; Cell Line; Cytokines; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Humans; Interleukin-8; Intestinal Mucosa; Microscopy, Electron; Neutrophil Activation; NF-kappa B; Nuclear Proteins

Middle East Respiratory Syndrome (MERS) is a novel respiratory illness firstly reported in Saudi Arabia in 2012. It is caused by a new corona virus, called MERS corona virus (MERS-CoV). Most people who have MERS-CoV infection developed severe acute respiratory illness.. This work is done to determine the clinical characteristics and the outcome of intensive care unit (ICU) admitted patients with confirmed MERS-CoV infection.. This study included 32 laboratory confirmed MERS corona virus infected patients who were admitted into ICU. It included 20 (62.50%) males and 12 (37.50%) females. The mean age was 43.99 ± 13.03 years. Diagnosis was done by real-time reverse transcription polymerase chain reaction (rRT-PCR) test for corona virus on throat swab, sputum, tracheal aspirate, or bronchoalveolar lavage specimens. Clinical characteristics, co-morbidities and outcome were reported for all subjects.. Most MERS corona patients present with fever, cough, dyspnea, sore throat, runny nose and sputum. The presence of abdominal symptoms may indicate bad prognosis. Prolonged duration of symptoms before patients' hospitalization, prolonged duration of mechanical ventilation and hospital stay, bilateral radiological pulmonary infiltrates, and hypoxemic respiratory failure were found to be strong predictors of mortality in such patients. Also, old age, current smoking, smoking severity, presence of associated co-morbidities like obesity, diabetes mellitus, chronic heart diseases, COPD, malignancy, renal failure, renal transplantation and liver cirrhosis are associated with a poor outcome of ICU admitted MERS corona virus infected patients.. Plasma HO-1, ferritin, p21, and NQO1 were all elevated at baseline in CKD participants. Plasma HO-1 and urine NQO1 levels each inversely correlated with eGFR (. SnPP can be safely administered and, after its injection, the resulting changes in plasma HO-1, NQO1, ferritin, and p21 concentrations can provide information as to antioxidant gene responsiveness/reserves in subjects with and without kidney disease.. A Study with RBT-1, in Healthy Volunteers and Subjects with Stage 3-4 Chronic Kidney Disease, NCT0363002 and NCT03893799.. HFNC did not significantly modify work of breathing in healthy subjects. However, a significant reduction in the minute volume was achieved, capillary [Formula: see text] remaining constant, which suggests a reduction in dead-space ventilation with flows > 20 L/min. (ClinicalTrials.gov registration NCT02495675).. 3 组患者手术时间、术中显性失血量及术后 1 周血红蛋白下降量比较差异均无统计学意义（. 对于肥胖和超重的膝关节单间室骨关节炎患者，采用 UKA 术后可获满意短中期疗效，远期疗效尚需进一步随访观察。.. Decreased muscle strength was identified at both time points in patients with hEDS/HSD. The evolution of most muscle strength parameters over time did not significantly differ between groups. Future studies should focus on the effectiveness of different types of muscle training strategies in hEDS/HSD patients.. These findings support previous adverse findings of e-cigarette exposure on neurodevelopment in a mouse model and provide substantial evidence of persistent adverse behavioral and neuroimmunological consequences to adult offspring following maternal e-cigarette exposure during pregnancy. https://doi.org/10.1289/EHP6067.. This RCT directly compares a neoadjuvant chemotherapy regimen with a standard CROSS regimen in terms of overall survival for patients with locally advanced ESCC. The results of this RCT will provide an answer for the controversy regarding the survival benefits between the two treatment strategies.. NCT04138212, date of registration: October 24, 2019.. Results of current investigation indicated that milk type and post fermentation cooling patterns had a pronounced effect on antioxidant characteristics, fatty acid profile, lipid oxidation and textural characteristics of yoghurt. Buffalo milk based yoghurt had more fat, protein, higher antioxidant capacity and vitamin content. Antioxidant and sensory characteristics of T. If milk is exposed to excessive amounts of light, Vitamins B. The two concentration of ZnO nanoparticles in the ambient air produced two different outcomes. The lower concentration resulted in significant increases in Zn content of the liver while the higher concentration significantly increased Zn in the lungs (p < 0.05). Additionally, at the lower concentration, Zn content was found to be lower in brain tissue (p < 0.05). Using TEM/EDX we detected ZnO nanoparticles inside the cells in the lungs, kidney and liver. Inhaling ZnO NP at the higher concentration increased the levels of mRNA of the following genes in the lungs: Mt2 (2.56 fold), Slc30a1 (1.52 fold) and Slc30a5 (2.34 fold). At the lower ZnO nanoparticle concentration, only Slc30a7 mRNA levels in the lungs were up (1.74 fold). Thus the two air concentrations of ZnO nanoparticles produced distinct effects on the expression of the Zn-homeostasis related genes.. Until adverse health effects of ZnO nanoparticles deposited in organs such as lungs are further investigated and/or ruled out, the exposure to ZnO nanoparticles in aerosols should be avoided or minimised.

Topics: A549 Cells; Acetylmuramyl-Alanyl-Isoglutamine; Acinetobacter baumannii; Acute Lung Injury; Adaptor Proteins, Signal Transducing; Adenine; Adenocarcinoma; Adipogenesis; Administration, Cutaneous; Administration, Ophthalmic; Adolescent; Adsorption; Adult; Aeromonas hydrophila; Aerosols; Aged; Aged, 80 and over; Aging; Agriculture; Air Pollutants; Air Pollution; Airway Remodeling; Alanine Transaminase; Albuminuria; Aldehyde Dehydrogenase 1 Family; Algorithms; AlkB Homolog 2, Alpha-Ketoglutarate-Dependent Dioxygenase; Alzheimer Disease; Amino Acid Sequence; Ammonia; Ammonium Compounds; Anaerobiosis; Anesthetics, Dissociative; Anesthetics, Inhalation; Animals; Anti-Bacterial Agents; Anti-HIV Agents; Anti-Infective Agents; Anti-Inflammatory Agents; Antibiotics, Antineoplastic; Antibodies, Antineutrophil Cytoplasmic; Antibodies, Monoclonal, Humanized; Antifungal Agents; Antigens, Bacterial; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Antimetabolites, Antineoplastic; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Antioxidants; Antitubercular Agents; Antiviral Agents; Apolipoproteins E; Apoptosis; Arabidopsis; Arabidopsis Proteins; Arsenic; Arthritis, Rheumatoid; Asthma; Atherosclerosis; ATP-Dependent Proteases; Attitude of Health Personnel; Australia; Austria; Autophagy; Axitinib; Bacteria; Bacterial Outer Membrane Proteins; Bacterial Proteins; Bacterial Toxins; Bacterial Typing Techniques; Bariatric Surgery; Base Composition; Bayes Theorem; Benzoxazoles; Benzylamines; beta Catenin; Betacoronavirus; Betula; Binding Sites; Biological Availability; Biological Oxygen Demand Analysis; Biomarkers; Biomarkers, Tumor; Biopsy; Bioreactors; Biosensing Techniques; Birth Weight; Blindness; Blood Chemical Analysis; Blood Gas Analysis; Blood Glucose; Blood Pressure; Blood Pressure Monitoring, Ambulatory; Blood-Brain Barrier; Blotting, Western; Body Mass Index; Body Weight; Bone and Bones; Bone Density; Bone Resorption; Borates; Brain; Brain Infarction; Brain Injuries, Traumatic; Brain Neoplasms; Breakfast; Breast Milk Expression; Breast Neoplasms; Bronchi; Bronchoalveolar Lavage Fluid; Buffaloes; Cadherins; Calcification, Physiologic; Calcium Compounds; Calcium, Dietary; Cannula; Caprolactam; Carbon; Carbon Dioxide; Carboplatin; Carcinogenesis; Carcinoma, Ductal; Carcinoma, Ehrlich Tumor; Carcinoma, Hepatocellular; Carcinoma, Non-Small-Cell Lung; Carcinoma, Pancreatic Ductal; Carcinoma, Renal Cell; Cardiovascular Diseases; Carps; Carrageenan; Case-Control Studies; Catalysis; Catalytic Domain; Cattle; CD8-Positive T-Lymphocytes; Cell Adhesion; Cell Cycle Proteins; Cell Death; Cell Differentiation; Cell Line; Cell Line, Tumor; Cell Movement; Cell Nucleus; Cell Phone Use; Cell Proliferation; Cell Survival; Cell Transformation, Neoplastic; Cell Transformation, Viral; Cells, Cultured; Cellulose; Chemical Phenomena; Chemoradiotherapy; Child; Child Development; Child, Preschool; China; Chitosan; Chlorocebus aethiops; Cholecalciferol; Chromatography, Liquid; Circadian Clocks; Circadian Rhythm; Circular Dichroism; Cisplatin; Citric Acid; Clinical Competence; Clinical Laboratory Techniques; Clinical Trials, Phase I as Topic; Clinical Trials, Phase II as Topic; Clostridioides difficile; Clostridium Infections; Coculture Techniques; Cohort Studies; Cold Temperature; Colitis; Collagen Type I; Collagen Type I, alpha 1 Chain; Collagen Type XI; Color; Connective Tissue Diseases; Copper; Coronary Angiography; Coronavirus 3C Proteases; Coronavirus Infections; Cost of Illness; Counselors; COVID-19; COVID-19 Testing; Creatine Kinase; Creatinine; Cross-Over Studies; Cross-Sectional Studies; Cryoelectron Microscopy; Cryosurgery; Crystallography, X-Ray; Cues; Cultural Competency; Cultural Diversity; Curriculum; Cyclic AMP Response Element-Binding Protein; Cyclin-Dependent Kinase Inhibitor p21; Cycloparaffins; Cysteine Endopeptidases; Cytokines; Cytoplasm; Cytoprotection; Databases, Factual; Denitrification; Deoxycytidine; Diabetes Complications; Diabetes Mellitus; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 1; Diabetes Mellitus, Type 2; Diagnosis, Differential; Diatoms; Diet; Diet, High-Fat; Dietary Exposure; Diffusion Magnetic Resonance Imaging; Diketopiperazines; Dipeptidyl Peptidase 4; Dipeptidyl-Peptidase IV Inhibitors; Disease Models, Animal; Disease Progression; Disease-Free Survival; DNA; DNA Damage; DNA Glycosylases; DNA Repair; DNA-Binding Proteins; DNA, Bacterial; DNA, Viral; Docetaxel; Dose Fractionation, Radiation; Dose-Response Relationship, Drug; Down-Regulation; Doxorubicin; Drosophila; Drosophila melanogaster; Drug Carriers; Drug Delivery Systems; Drug Liberation; Drug Repositioning; Drug Resistance, Bacterial; Drug Resistance, Multiple, Bacterial; Drug Resistance, Neoplasm; Drug Screening Assays, Antitumor; Drug Synergism; Drug Therapy, Combination; Edema; Edible Grain; Education, Graduate; Education, Medical, Graduate; Education, Pharmacy; Ehlers-Danlos Syndrome; Electron Transport Complex III; Electron Transport Complex IV; Electronic Nicotine Delivery Systems; Emergency Service, Hospital; Empathy; Emulsions; Endothelial Cells; Endurance Training; Energy Intake; Enterovirus A, Human; Environment; Environmental Monitoring; Enzyme Assays; Enzyme Inhibitors; Epithelial Cells; Epithelial-Mesenchymal Transition; Epoxide Hydrolases; Epoxy Compounds; Erythrocyte Count; Erythrocytes; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Esophagectomy; Estrogens; Etanercept; Ethiopia; Ethnicity; Ethylenes; Exanthema; Exercise; Exercise Test; Exercise Tolerance; Extracellular Matrix; Extracorporeal Membrane Oxygenation; Eye Infections, Fungal; False Negative Reactions; Fatty Acids; Fecal Microbiota Transplantation; Feces; Female; Femur Neck; Fermentation; Ferritins; Fetal Development; Fibroblast Growth Factor-23; Fibroblast Growth Factors; Fibroblasts; Fibroins; Fish Proteins; Flavanones; Flavonoids; Focus Groups; Follow-Up Studies; Food Handling; Food Supply; Food, Formulated; Forced Expiratory Volume; Forests; Fractures, Bone; Fruit and Vegetable Juices; Fusobacteria; G1 Phase Cell Cycle Checkpoints; G2 Phase Cell Cycle Checkpoints; Gamma Rays; Gastrectomy; Gastrointestinal Microbiome; Gastrointestinal Stromal Tumors; Gefitinib; Gels; Gemcitabine; Gene Amplification; Gene Expression; Gene Expression Regulation; Gene Expression Regulation, Bacterial; Gene Expression Regulation, Neoplastic; Gene Expression Regulation, Plant; Gene Knockdown Techniques; Gene-Environment Interaction; Genotype; Germany; Glioma; Glomerular Filtration Rate; Glucagon; Glucocorticoids; Glycemic Control; Glycerol; Glycogen Synthase Kinase 3 beta; Glycolipids; Glycolysis; Goblet Cells; Gram-Negative Bacterial Infections; Granulocyte Colony-Stimulating Factor; Graphite; Greenhouse Effect; Guanidines; Haemophilus influenzae; HCT116 Cells; Health Knowledge, Attitudes, Practice; Health Personnel; Health Services Accessibility; Health Services Needs and Demand; Health Status Disparities; Healthy Volunteers; Heart Failure; Heart Rate; Heart Transplantation; Heart-Assist Devices; HEK293 Cells; Heme; Heme Oxygenase-1; Hemolysis; Hemorrhage; Hepatitis B; Hepatitis B e Antigens; Hepatitis B Surface Antigens; Hepatitis B virus; Hepatitis B, Chronic; Hepatocytes; Hexoses; High-Throughput Nucleotide Sequencing; Hippo Signaling Pathway; Histamine; Histamine Agonists; Histidine; Histone Deacetylase 2; HIV Infections; HIV Reverse Transcriptase; HIV-1; Homebound Persons; Homeodomain Proteins; Homosexuality, Male; Hospice and Palliative Care Nursing; HSP70 Heat-Shock Proteins; Humans; Hyaluronan Receptors; Hydrogen; Hydrogen Peroxide; Hydrogen-Ion Concentration; Hydrolysis; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Hypoglycemia; Hypoglycemic Agents; Hypoxia; Idiopathic Interstitial Pneumonias; Imaging, Three-Dimensional; Imatinib Mesylate; Immunotherapy; Implementation Science; Incidence; INDEL Mutation; Induced Pluripotent Stem Cells; Industrial Waste; Infant; Infant, Newborn; Inflammation; Inflammation Mediators; Infliximab; Infusions, Intravenous; Inhibitory Concentration 50; Injections; Insecticides; Insulin-Like Growth Factor Binding Protein 5; Insulin-Secreting Cells; Interleukin-1; Interleukin-17; Interleukin-8; Internship and Residency; Intestines; Intracellular Signaling Peptides and Proteins; Ion Transport; Iridaceae; Iridoid Glucosides; Islets of Langerhans Transplantation; Isodon; Isoflurane; Isotopes; Italy; Joint Instability; Ketamine; Kidney; Kidney Failure, Chronic; Kidney Function Tests; Kidney Neoplasms; Kinetics; Klebsiella pneumoniae; Knee Joint; Kruppel-Like Factor 4; Kruppel-Like Transcription Factors; Lactate Dehydrogenase 5; Laparoscopy; Laser Therapy; Lasers, Semiconductor; Lasers, Solid-State; Laurates; Lead; Leukocyte L1 Antigen Complex; Leukocytes, Mononuclear; Light; Lipid Peroxidation; Lipopolysaccharides; Liposomes; Liver; Liver Cirrhosis; Liver Neoplasms; Liver Transplantation; Locomotion; Longitudinal Studies; Lopinavir; Lower Urinary Tract Symptoms; Lubricants; Lung; Lung Diseases, Interstitial; Lung Neoplasms; Lymphocyte Activation; Lymphocytes, Tumor-Infiltrating; Lymphoma, Mantle-Cell; Lysosomes; Macrophages; Male; Manganese Compounds; MAP Kinase Kinase 4; Mass Screening; Maternal Health; Medicine, Chinese Traditional; Melanoma, Experimental; Memantine; Membrane Glycoproteins; Membrane Proteins; Mesenchymal Stem Cell Transplantation; Metal Nanoparticles; Metalloendopeptidases; Metalloporphyrins; Methadone; Methane; Methicillin-Resistant Staphylococcus aureus; Mexico; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred ICR; Mice, Knockout; Mice, Nude; Mice, SCID; Mice, Transgenic; Microarray Analysis; Microbial Sensitivity Tests; Microbiota; Micronutrients; MicroRNAs; Microscopy, Confocal; Microsomes, Liver; Middle Aged; Milk; Milk, Human; Minority Groups; Mitochondria; Mitochondrial Membranes; Mitochondrial Proteins; Models, Animal; Models, Molecular; Molecular Conformation; Molecular Docking Simulation; Molecular Dynamics Simulation; Molecular Epidemiology; 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Oocytes; Open Reading Frames; Osteoclasts; Osteogenesis; Osteoporosis; Osteoporosis, Postmenopausal; Outpatients; Ovarian Neoplasms; Ovariectomy; Overweight; Oxazines; Oxidants; Oxidation-Reduction; Oxidative Stress; Oxides; Oxidoreductases; Oxygen; Oxygen Inhalation Therapy; Oxygenators, Membrane; Ozone; Paclitaxel; Paenibacillus; Pain Measurement; Palliative Care; Pancreatic Neoplasms; Pandemics; Parasympathetic Nervous System; Particulate Matter; Pasteurization; Patient Preference; Patient Satisfaction; Pediatric Obesity; Permeability; Peroxiredoxins; Peroxynitrous Acid; Pharmaceutical Services; Pharmacists; Pharmacy; Phaseolus; Phenotype; Phoeniceae; Phosphates; Phosphatidylinositol 3-Kinases; Phospholipid Transfer Proteins; Phospholipids; Phosphorus; Phosphorylation; Photoperiod; Photosynthesis; Phylogeny; Physical Endurance; Physicians; Pilot Projects; Piperidines; Pituitary Adenylate Cyclase-Activating Polypeptide; Plant Extracts; Plant Leaves; Plant Proteins; Plant Roots; Plaque, Atherosclerotic; Pneumonia; Pneumonia, Viral; Point-of-Care Testing; Polyethylene Glycols; Polymers; Polysorbates; Pore Forming Cytotoxic Proteins; Positron Emission Tomography Computed Tomography; Positron-Emission Tomography; Postprandial Period; Poverty; Pre-Exposure Prophylaxis; Prediabetic State; Predictive Value of Tests; Pregnancy; Pregnancy Trimester, First; Pregnancy, High-Risk; Prenatal Exposure Delayed Effects; Pressure; Prevalence; Primary Graft Dysfunction; Primary Health Care; Professional Role; Professionalism; Prognosis; Progression-Free Survival; Prolactin; Promoter Regions, Genetic; Proof of Concept Study; Proportional Hazards Models; Propylene Glycol; Prospective Studies; Prostate; Protein Binding; Protein Biosynthesis; Protein Isoforms; Protein Kinase Inhibitors; Protein Phosphatase 2; Protein Processing, Post-Translational; Protein Serine-Threonine Kinases; Protein Structure, Tertiary; Protein Transport; Proteoglycans; Proteome; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-myc; Proto-Oncogene Proteins c-ret; Proto-Oncogene Proteins p21(ras); Proton Pumps; Protons; Protoporphyrins; Pseudomonas aeruginosa; Pseudomonas fluorescens; Pulmonary Artery; Pulmonary Disease, Chronic Obstructive; Pulmonary Gas Exchange; Pulmonary Veins; Pyrazoles; Pyridines; Pyrimidines; Qualitative Research; Quinoxalines; Rabbits; Random Allocation; Rats; Rats, Sprague-Dawley; Rats, Wistar; Receptors, Histamine H3; Receptors, Immunologic; Receptors, Transferrin; Recombinant Proteins; Recurrence; Reference Values; Referral and Consultation; Regional Blood Flow; Registries; Regulon; Renal Insufficiency, Chronic; Reperfusion Injury; Repressor Proteins; Reproducibility of Results; Republic of Korea; Research Design; Resistance Training; Respiration, Artificial; Respiratory Distress Syndrome; Respiratory Insufficiency; Resuscitation; Retinal Dehydrogenase; Retreatment; Retrospective Studies; Reverse Transcriptase Inhibitors; Rhinitis, Allergic; Ribosomal Proteins; Ribosomes; Risk Assessment; 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STAT3 Transcription Factor; Streptomyces coelicolor; Stress, Psychological; Stroke; Stroke Volume; Structure-Activity Relationship; Students, Medical; Students, Pharmacy; Substance Abuse Treatment Centers; Sulfur Dioxide; Surface Properties; Surface-Active Agents; Surveys and Questionnaires; Survival Analysis; Survival Rate; Survivin; Sweden; Swine; Swine, Miniature; Sympathetic Nervous System; T-Lymphocytes, Regulatory; Talaromyces; Tandem Mass Spectrometry; tau Proteins; Telemedicine; Telomerase; Telomere; Telomere Homeostasis; Temperature; Terminally Ill; Th1 Cells; Thiamethoxam; Thiazoles; Thiophenes; Thioredoxin Reductase 1; Thrombosis; Thulium; Thyroid Cancer, Papillary; Thyroid Carcinoma, Anaplastic; Thyroid Neoplasms; Time Factors; Titanium; Tomography, Emission-Computed, Single-Photon; Tomography, X-Ray Computed; TOR Serine-Threonine Kinases; Transcription Factor AP-1; Transcription Factors; Transcription, Genetic; Transcriptional Activation; Transcriptome; Transforming Growth Factor beta1; Transistors, Electronic; Translational Research, Biomedical; Transplantation Tolerance; Transplantation, Homologous; Transportation; Treatment Outcome; Tretinoin; Tuberculosis, Multidrug-Resistant; Tuberculosis, Pulmonary; Tubulin Modulators; Tumor Microenvironment; Tumor Necrosis Factor Inhibitors; Tumor Necrosis Factor-alpha; Twins; Ultrasonic Therapy; Ultrasonography; Ultraviolet Rays; United States; Up-Regulation; Uranium; Urethra; Urinary Bladder; Urodynamics; Uromodulin; Uveitis; Vasoconstrictor Agents; Ventricular Function, Left; Vero Cells; Vesicular Transport Proteins; Viral Nonstructural Proteins; Visual Acuity; Vital Capacity; Vitamin D; Vitamin D Deficiency; Vitamin K 2; Vitamins; Volatilization; Voriconazole; Waiting Lists; Waste Disposal, Fluid; Wastewater; Water Pollutants, Chemical; Whole Genome Sequencing; Wine; Wnt Signaling Pathway; Wound Healing; Wounds and Injuries; WW Domains; X-linked Nuclear Protein; X-Ray Diffraction; Xanthines; Xenograft Model Antitumor Assays; YAP-Signaling Proteins; Yogurt; Young Adult; Zebrafish; Zebrafish Proteins; Ziziphus

Urinary interleukin-6 (UIL-6) and urinary interleukin-8 (UIL-8) concentrations were measured by immunoassay in 39 and 34 patients respectively, hospitalized with febrile urinary tract infection (UTI), and in 37 and 32 age-, race- and sex-matched febrile control children respectively, with negative urine cultures. UIL-6 and UIL-8 concentrations, measured in picograms per milliliter and corrected for creatinine, were compared with clinical and laboratory indicators of inflammation and bacterial virulence factors of Escherichia coli. Median UIL-6 concentrations at the time of admission were 397 pg/ml (range 0-65,789 pg/ml) in the 37 patients compared to 0 pg/ml (range 0-473.8 pg/ml) in the 37 controls (P < 0.0001). Median UIL-8 concentrations at the time of admission were 5809 pg/ml (range 0-347,368 pg/ml) in the 32 patients compared to 0 pg/ml (range 0-2231 pg/ml) in the 32 controls (P < 0.0001). UIL-6 and UIL-8 concentrations were lower (P < 0.0001 for UIL-6 and P = 0.0005 for UIL-8) in follow-up urine samples from UTI patients, obtained 48 h after the initiation of antibiotic therapy. UIL-6 and UIL-8 concentrations were statistically significantly correlated with urine white blood cells (WBC). UIL-8 concentrations were elevated in patients with E. coli organisms producing hemolysin. UIL-6 and UIL-8 are elevated in children with febrile UTI and decrease in response to antibiotic therapy. Magnitude of UIL-8 response is associated with hemolysin production, a bacterial virulence factor of E. coli. UIL-6 and UIL-8 concentrations are statistically correlated with urine WBC. UIL-6 and UIL-8 may be mediators of inflammation in children with febrile UTI.

Topics: Child; Child, Preschool; Creatinine; Escherichia coli Infections; Female; Fever; Humans; Immunoassay; Infant; Infant, Newborn; Interleukin-6; Interleukin-8; Male; Reference Values; Retrospective Studies; Urinary Tract Infections

Emerging evidence have suggested that aberrant sialylation on cell-surface carbohydrate architecture may influence host-pathogen interactions. The α2,6-sialyltransferase (ST) enzymes were found to alter the glycosylation pattern of the pathogen-infected host cell-surface proteins, which could facilitate its invasion. In this study, we assessed the role of specific α2,6-ST enzymes in the regulation of enteroaggregative E. coli (EAEC)-induced cell signaling pathways in human intestinal epithelial cells. EAEC-induced expression of α2,6-ST family genes in HCT-15 and INT-407 cell lines was assessed at mRNA level by qRT-PCR. Specific esi-RNA was used to silence the target ST-gene in each of the EAEC-infected cell type. Subsequently, the role of these enzymes in regulation of EAEC-induced cell signaling pathways was unraveled by analyzing the expression of MAPkinases (ERK1/2, p38, JNK) and transcription factors (NFκB, cJun, cFos, STAT) at mRNA and protein levels by qRT-PCR and western immunoblotting, respectively, expression of selected sialoglycoproteins by western immunoblotting along with the secretory IL-8 response using sandwich ELISA. ST6GAL-1 and ST6GAL-2 were efficiently silenced in EAEC-infected HCT-15 and INT-407 cells, respectively. Significant reduction in EAEC-induced activation of MAPKs, transcription factors, sialoglycoproteins, and IL-8 secretion was noted in ST-silenced cells in comparison to the respective control cells. We propose that ST6GAL-1 and ST6GAL-2 are quintessential for EAEC-induced stimulation of MAPK-mediated pathways, resulting in activation of transcription factors, leading to an inflammatory response in the human intestinal epithelial cells. Our study may be helpful to design better therapeutic strategies to control EAEC- infection. KEY POINTS: • EAEC induces α2,6-sialyltransferase (ST) upregulation in intestinal epithelial cells • Target STs (ST6GAL-1 & ST6GAL-2) were efficiently silenced using specific esiRNAs • Expression of MAPKs, transcription factors & IL-8 was reduced in ST silenced cells.

Topics: Antigens, CD; beta-D-Galactoside alpha 2-6-Sialyltransferase; Cell Line; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Humans; Interleukin-8; RNA, Messenger; Sialyltransferases; Transcription Factors

Enterotoxigenic Escherichia coli (ETEC) is an important cause of diarrhea in piglets, which leads to great economic losses. In this study, the ternary crossbred weaned piglets were orally administered with 1.5 × 10

Topics: Animals; Enterotoxigenic Escherichia coli; Escherichia coli Infections; Interleukin-13; Interleukin-8; Intestinal Diseases; Intestinal Mucosa; Intestines; Swine; Swine Diseases; Tumor Necrosis Factor-alpha

Many bacterial and fungal pathogens cause disease across mucosal surfaces, and to a lesser extent through skin surfaces. Pathogens that potentially cause disease vaginally across epithelial cells include Staphylococcus aureus, group A and B streptococci, Escherichia coli, Neisseria gonorrhoeae, and Candida albicans. We have previously shown that staphylococcal and streptococcal superantigens induce inflammatory chemokines from vaginal epithelial cells through the immune costimulatory molecule CD40 through use of a CRISPR cas9 knockout mutant and complemented epithelial cell line. In this study, we show that the potential vaginal pathogens S. aureus, group A and B streptococci, E. coli, an Enterococcus faecalis strain, and C. albicans in part use CD40 to stimulate interleukin-8 (IL-8) production from human vaginal epithelial cells. In contrast, N. gonorrhoeae does not appear to use CD40 to signal IL-8 production. Normal flora Lactobacillus crispatus and an Enterococcus faecalis strain that produces reutericyclin do not induce IL-8. These data indicate that many potential pathogens, but no normal commensals, induce IL-8 to help disrupt the human vaginal epithelial barrier through CD40, thus providing a potential therapeutic target for drug development.

Topics: Candida albicans; Chemokines; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Female; Humans; Interleukin-8; Staphylococcal Infections; Staphylococcus aureus; Vagina

Topics: Epithelial Cells; Escherichia coli Infections; Female; Glucose; Humans; Inflammation; Interleukin-6; Interleukin-8; Male; Ribonucleases; Signal Transduction; Toll-Like Receptor 4; Urinary Bladder; Urinary Tract Infections; Uropathogenic Escherichia coli

Autophagy is crucial for the maintenance of homeostasis under stimuli related to infection. Selenium (Se) plays variable roles in defence against infection and Selenomethionine (Se-Met) is a common Se supplementation. This study aimed to understand whether Se-Met could regulate the nuclear factor-kappa B (NF-κB) signaling pathway through autophagy. Mammary alveolar cell-T (MAC-T) was challenged with Escherichia coli (E. coli). Western blotting and real-time quantitative PCR (RT-qPCR) were used to detect the protein expression and mRNA expression of cytokines. Immunofluorescence assays were performed to observe the expression of intracellular LC3. The results showed that E. coli inhibited autophagy by decreasing the LC3-Ⅱ protein levels, and the Atg5 and Beclin1 protein levels were increased after 4 h. Infection also decreased the number of LC3 puncta. E. coli increased the phosphorylation of p65 and IκBα protein. Concomitantly, the levels of interleukin (IL)-1β, IL-6, IL-8 and tumour necrosis factor (TNF)-α mRNA increased at 3 and 4 h post-infection. We further explored the regulatory role of autophagy on NF-κB-mediated inflammation with autophagy modulators and shAtg5. The results indicated that the autophagy activator reduced the phosphorylation of p65 and IκBα and the mRNA expression of IL-1β, IL-6, IL-8 and TNF-α. Additionally, activating autophagy weakened the adhesion to MAC-T of E. coli. Autophagy inhibitors exacerbated NF-κB-mediated inflammation and strengthened the adhesion of E. coli to cells. We then examined the effects of Se-Met on NF-κB-mediated inflammation through autophagy. The data suggested that Se-Met enhanced LC3-II expression, inhibited the E. coli-induced phosphorylation of p65 and IκBα, and suppressed the adhesion ability of E. coli to MAC-T and that the effects of Se-Met in attenuating NF-κB-mediated inflammation were partially blocked by an autophagy inhibitor. In summary, Se-Met alleviated NF-κB-mediated inflammation induced by E. coli by enhancing autophagy in bovine mammary epithelial cells.

Topics: Animals; Autophagy; Cattle; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Inflammation; Interleukin-6; Interleukin-8; NF-kappa B; NF-KappaB Inhibitor alpha; RNA, Messenger; Selenomethionine; Tumor Necrosis Factor-alpha

This study aimed to assess the in vitro antimicrobial effects of chlorhexidine (CHX) and povidone-iodine (PI) on clinical isolates of Escherichia coli (E. coli) and Trueperella pyogenes (T. pyogenes) from the vaginal discharge of dairy cows, as well as to compare the cytotoxicity effects of CHX and PI on bovine endometrial epithelial cells (BEnEpC). In Experiment 1, 12 E. coli and 10 T. pyogenes were isolated from the vaginal discharge of cows with a uterine infection. The MIC and MBC against CHX and PI were analyzed in vitro. In Experiment 2, the cytotoxicity effects of CHX and PI on BEnEpC were analyzed using a Viability/Cytotoxicity Kit, wound scratch healing assay, and the expression of pro-inflammatory cytokine genes (IL-6, IL-8, and TNF-α). In Experiment 1, the MIC and MBC values of CHX against E. coli were 0.0002% and 0.0002 to 0.00025%, respectively. The MIC and MBC values of PI were 1.25 to 2.5% and 1.25 to 5%, respectively. For T. pyogenes, the MIC and MBC values of CHX were 0.00002%. The MIC and MBC values of PI were 1.25%. In Experiment 2, the cell viability significantly decreased, and wound closures were significantly inhibited after treatment with ≥ 0.002% CHX and ≥ 0.025% PI. The expression of IL-6, IL-8, and TNF-α significantly increased after treatment with PI. Only IL-6 showed a significant increase after cells were treated with 0.00002% and 0.0002% CHX. The results suggested that both CHX and PI had high antibacterial effects. However, veterinarians and farmers should be aware of their cytotoxicity, which decrease viability of endometrial epithelial cells and inhibit wound healing in vitro.

Topics: Actinomycetaceae; Animals; Anti-Bacterial Agents; Cattle; Cattle Diseases; Chlorhexidine; Endometritis; Escherichia coli; Escherichia coli Infections; Female; Interleukin-6; Interleukin-8; Povidone-Iodine; Tumor Necrosis Factor-alpha; Vaginal Discharge

Bacterial flagellin is a potent powerful adjuvant, which exerts its adjuvant activity by activating the Toll-like receptor 5 (TLR5) signaling pathway to induce host pro-inflammatory responses. Flagellin of Salmonella typhimurium (S. typhimurium) has shown strong adjuvant effects for a variety of vaccine candidates, however, the adjuvanticity of different serotypes of Escherichia coli (E. coli) flagellin (FliC) is unclear. To explore the adjuvant activity of different serotypes of E. coli flagellin, FliC

Topics: Adjuvants, Immunologic; Animals; Caco-2 Cells; Escherichia coli; Escherichia coli Infections; Flagellin; Humans; Interleukin-8; Mice; Rodent Diseases; Serogroup; Toll-Like Receptor 5

Diarrheagenic. We determined the effect of supernatants obtained from CW and EA cultures on the gene expression of STEC strain 86-24 and EAEC strain 042 by RNA-seq analysis. We evaluated IL-8 secretion from T84 cells infected with these DEC strains in the presence or absence of the supernatant from EA. The effect of the supernatant from EA on the growth and adherence of STEC and EAEC to cells was also evaluated. Finally, we studied the effect of the EA supernatant on the STEC-induced inflammation mediated by the long polar fimbriae (Lpf) in T84 cells and the expression of plasmid-encoded toxin (Pet) in EAEC.. RNA-seq analysis revealed that several virulence factors in STEC and EAEC were upregulated in the presence of supernatants from CW and EA. Interestingly, an increase in the secretion of IL-8 was observed in cells infected with STEC or EAEC in the presence of a supernatant from EA. Similar results were observed with the supernatants obtained from clinical strains of. Supernatant obtained from an indicative species of DEC-positive diarrhea could modulate gene expression in STEC and EAEC, and IL-8 secretion induced by these bacteria. These data provide new insights into the effect of gut microbiota species in the pathogenicity of STEC and EAEC.

Topics: Child; Diarrhea; Escherichia coli; Escherichia coli Infections; Gastrointestinal Microbiome; Humans; Interleukin-8; Shiga Toxin; Virulence

Topics: Animals; Cytoskeletal Proteins; Enterotoxigenic Escherichia coli; Escherichia coli Infections; Interleukin-8; RNA-Seq; RNA, Long Noncoding; Swine; Swine Diseases

The intestinal mucus layer has a dual role in human health constituting a well-known microbial niche that supports gut microbiota maintenance but also acting as a physical barrier against enteric pathogens. Enterotoxigenic Escherichia coli (ETEC), the major agent responsible for traveler's diarrhea, is able to bind and degrade intestinal mucins, representing an important but understudied virulent trait of the pathogen. Using a set of complementary in vitro approaches simulating the human digestive environment, this study aimed to describe how the mucus microenvironment could shape different aspects of the human ETEC strain H10407 pathophysiology, namely its survival, adhesion, virulence gene expression, interleukin-8 induction and interactions with human fecal microbiota. Using the TNO gastrointestinal model (TIM-1) simulating the physicochemical conditions of the human upper gastrointestinal (GI) tract, we reported that mucus secretion and physical surface sustained ETEC survival, probably by helping it to face GI stresses. When integrating the host part in Caco2/HT29-MTX co-culture model, we demonstrated that mucus secreting-cells favored ETEC adhesion and virulence gene expression, but did not impede ETEC Interleukin-8 (IL-8) induction. Furthermore, we proved that mucosal surface did not favor ETEC colonization in a complex gut microbial background simulated in batch fecal experiments. However, the mucus-specific microbiota was widely modified upon the ETEC challenge suggesting its role in the pathogen infectious cycle. Using multi-targeted in vitro approaches, this study supports the major role played by mucus in ETEC pathophysiology, opening avenues in the design of new treatment strategies.

Topics: Bacteria; Caco-2 Cells; Diarrhea; Enterotoxigenic Escherichia coli; Escherichia coli Infections; Humans; Interleukin-8; Microbiota; Mucins; Mucus; Travel; Virulence

Inflammation is a cascade of events mediated by a cytokine network triggering the cellular response. In order to monitor the modulation of the crucial inflammatory proteins, e.g., Tumour Necrosis Factor-α (TNF-α), Interferon-γ (INF-γ), Interleukin-8 (IL-8) and Interleukin-10 (IL-10), upon stimulation with endotoxins, differentiated and undifferentiated THP-1 cells were treated with lipopolysaccharides (LPSs) from E. coli, key cell wall components of Gram-negative bacteria.. The multiple reaction monitoring mass spectrometry (MRM-MS) method was optimized by using the standard proteins to be quantified, in order to construct external calibration curves and define the analytical parameters. The developed method was used to quantify the above-mentioned inflammatory proteins in THP-1 differentiated cells upon stimulation with LPSs with high accuracy, sensitivity, and robustness.. The analysis of such proteins in MRM mode allowed the kinetics of stimulation along the time up to 24 h to be followed and the MS results were found to be comparable with those obtained by Western-blotting. A significant increase in TNF-α release triggered a cascade mechanism leading to the production of INF-γ and IL-8. IL-10, instead, was found to be constant throughout the process.. The developed MRM-MS method allowed the quantification of TNF-α, INF-γ, IL-8 and IL-10 along a time-course from 2 to 24 h. Hence, a trace of the kinetics of the inflammatory response in THP-1 cells upon stimulation with E. coli LPSs was obtained. Finally, the extensibility of the developed MRM method to serum samples and other matrices demonstrated the versatility of the approach and the possibility to quantify multiple target proteins in different biological samples by using a few microliters in a single analysis.

Topics: Escherichia coli; Escherichia coli Infections; Humans; Inflammation; Interferon-gamma; Interleukin-10; Interleukin-8; Kinetics; Lipopolysaccharides; Mass Spectrometry; Monocytes; THP-1 Cells; Tumor Necrosis Factor-alpha

Avian pathogenic Escherichia coli (APEC) is an important pathogen that causes avian colibacillosis in poultry. APEC infection can lead to pathological changes in chicken trachea. The type VI secretion system (T6SS) of APEC contribute to the pathogenicity of APEC. However, whether T6SS plays a role in infection of the trachea remains unclear. We constructed mutant strain Δhcp2a by the Red recombination method system. The role of hcp2a (the structural secretion components and secretory protein of the T6SS) in the infection of trachea was investigated. The mutation strain displayed a significant increase in biofilm formation and a decrease in resistance to chicken serum. Moreover, RNA sequencing analyses showed that infection of chicken tracheal epithelium by the mutant strain Δhcp2a induced differential expression of genes. The result also showed that 14 genes (13 genes were downregulated) were enriched in cytokine-cytokine receptor interaction signalling pathway at 12 and 24 h post infection. The mutation Δhcp2a resulted in significant decreases in the bacterial loads in trachea at 6 and 12 h post infection. Real-time PCR analyses showed that the hcp2a mutation downregulated the expression of IL8 and IL1β at mRNA level in chicken tracheal epithelium. Our results indicate that mutation of hcp2a influenced genes expression of the cytokine-cytokine receptor interaction pathway by decreasing APEC colonization in the trachea.

Topics: Animals; Bacterial Load; Chickens; Epithelium; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Gene Expression Regulation; Interleukin-1beta; Interleukin-8; Mutation; Poultry Diseases; Trachea; Type VI Secretion Systems; Virulence Factors

Lactic acid bacteria (LAB) "fermentates" confer a beneficial effect on intestinal function. However, the ability of new fermentations to improve LAB broth activity in preventing pathogen-induced intestinal inflammation and barrier dysfunction has not yet been studied. The objective of this study was to determine if broths of LAB fermented with

Topics: Anti-Bacterial Agents; Barbarea; Brassicaceae; Caco-2 Cells; Cell Survival; Coculture Techniques; Drug Resistance, Bacterial; Electric Impedance; Escherichia coli Infections; Escherichia coli O157; Fermentation; Gastroenteritis; Humans; Interleukin-8; Intestinal Mucosa; Lactobacillus acidophilus; Phytotherapy; Plant Extracts; Probiotics; Seeds

To prevent the onset of urosepsis and reduce mortality, a better understanding of how uropathogenic Escherichia coli (UPEC) manages to infiltrate the bloodstream through the kidneys is needed. The present study elucidates if human renal interstitial fibroblasts are part of the immune response limiting a UPEC infection, or if UPEC has the ability to modulate the fibroblasts for their own gain. Microarray results showed that upregulated genes were associated with an activated immune response. We also found that chemokines released from renal fibroblasts upon a UPEC infection could be mediated by LPS and triacylated lipoproteins activating the TLR2/1, TLR4, MAPK, NF-κB and PKC signaling pathways. Furthermore, UPEC was also shown to be able to adhere and invade renal fibroblasts, mediated by the P-fimbriae. Furthermore, it was found that renal fibroblasts were more immunoreactive than renal epithelial cells upon a UPEC infection. However, both renal fibroblasts and epithelial cells were equally efficient at inducing neutrophil migration. In conclusion, we have found that human renal fibroblasts can sense UPEC and mobilize a host response with neutrophil migration. This suggests that renal fibroblasts are not only structural cells that produce and regulate the extracellular matrix, but also highly immunoreactive cells.

Topics: Escherichia coli Infections; Escherichia coli Proteins; Fibroblasts; Humans; Interleukin-8; Kidney; NF-kappa B; Protein Kinase C; Signal Transduction; Toll-Like Receptor 1; Toll-Like Receptor 2; Toll-Like Receptor 4; Urinary Tract Infections; Uropathogenic Escherichia coli

Topics: Cytokines; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Fimbriae, Bacterial; HT29 Cells; Humans; Inflammation; Interleukin-4; Interleukin-8; Intestines; Toll-Like Receptor 2; Toll-Like Receptor 4

The objective of this study was twofold: (i) to examine the effect of Clostridium butyricum on intestinal barrier function and (ii) to elucidate the mechanisms involved in enhanced intestinal barrier function.. Forty-eight weaned piglets were assigned randomly to either a basal diet or a C. butyricum-supplemented diet. On day 15, all pigs were orally challenged with enterotoxigenic Escherichia coli (ETEC) K88 or saline. Clostridium butyricum decreased serum diamine oxidase activity and d-lactic acid concentration, as well as increased intestinal tight junction proteins (ZO-1, claudin-3 and occludin) expression in ETEC K88-infected pigs. Moreover, C. butyricum decreased IL-1β and IL-18 levels in serum and gut, whereas it increased IL-10 levels. Furthermore, C. butyricum downregulated NLRP3 and caspase-1 expression in ETEC K88-challenged pig gut, but did not affect apoptosis-associated speck-like protein expression.. Clostridium butyricum enhanced intestinal barrier function and inhibited apoptosis-associated speck-like protein-independent NLRP3 inflammasome signalling pathway in weaned piglets after ETEC K88 challenge.. The novelty of this study lies in the beneficial effects of C. butyricum on intestinal health, likely by improving intestinal barrier function and alleviating inflammatory reactions.

Topics: Animal Feed; Animals; Clostridium butyricum; Diet; Dietary Supplements; Enterotoxigenic Escherichia coli; Escherichia coli Infections; Female; Interleukin-1beta; Interleukin-8; Intestinal Mucosa; Intestines; Male; Probiotics; Swine; Swine Diseases; Weaning

Topics: Bacterial Outer Membrane Proteins; Caco-2 Cells; Cell Line, Tumor; Cell Membrane; Epithelial Cells; Escherichia coli Infections; Escherichia coli O157; Escherichia coli Proteins; Flagellin; HT29 Cells; Humans; Interleukin-8; Intestinal Mucosa; NF-kappa B; Signal Transduction; Toll-Like Receptor 4; Toll-Like Receptor 5; Virulence Factors

Mastitis caused by environmental pathogens such as Escherichia coli is highly problematic to the dairy industry because it incurs substantial cost and tends to be difficult to manage. An effective innate immune response by the host is key to controlling infection, but it should also limit collateral damage to the mammary gland. Between-animal differences in mastitis severity have been attributed to variability in the innate response. In the current study, we used primary dermal fibroblast as a model to rank animals based on composite expression of the toll-like receptor 4 gene (TLR4) and lipopolysaccharide (LPS)-induced IL-8 and IL-6 protein production. Animals ranked as high and low responders (HR and LR, respectively) were then infected with the P4 strain of E. coli to determine how difference in rank would affect response to mastitis. All animals developed an acute response to the infection with varying degrees in severity; however, HR animals had an elevated somatic cell count and fever response at 12 h post-infection and greater production of milk IL-8 at 24 h post-infection. The HR animals were also significantly more capable of limiting bacterial growth. No differences in post-infection milk production or concentrations of milk BSA were measured. The current study indicates that HR animals have an early upregulation in their innate response that is beneficial for bacterial clearance; however, they are equally susceptible to tissue damage caused by an exuberant response to the infection. The dermal fibroblast may be used in conjunction with other cell types to determine how the innate response is regulated to mitigate unnecessary injury to the mammary gland while still effectively clearing the pathogen.

Topics: Animals; Cattle; Cell Count; Dairying; Escherichia coli; Escherichia coli Infections; Female; Fibroblasts; Gene Expression Regulation; Immunity, Innate; Interleukin-6; Interleukin-8; Lipopolysaccharides; Mastitis, Bovine; Milk; Toll-Like Receptor 4

Cathelicidin are innate antimicrobial peptides with broad immunomodulatory functions; however, their role in regulating intestinal defenses is not well characterized. This study aimed to investigate the role of cathelicidin modulating expression of Toll-like receptors (TLRs) 4 and 9 in colonic epithelium in response to bacterial patterns. We demonstrated herein that intestinal epithelial cells, when primed by bacterial lipopolysaccharide (LPS), responded to cathelicidin by increased transcription and protein synthesis of TLR4. This cathelicidin-induced response required the interaction of LPS-TLR4 and activation of MAPK signalling pathways. However, cathelicidin blocked TLR9 responses induced by TLR9 ligand CpG oligodeoxynucleotide (CpG ODN) in these colonic epithelial cells. Modulations of TLRs triggered by cathelicidin in intestinal epithelium occurred mainly in the apical compartment of intestinal cells. Activation of TLR4 by ligands in combination with cathelicidin promoted CXCL8 chemokine secretion and epithelial antimicrobial defenses against Escherichia coli. We concluded that cathelicidin selectively modulated synthesis of TLR4 and 9 in intestinal epithelium, but only when cells were exposed to virulence factors, mostly from apical surfaces. Enhanced TLR4 expression promoted by cathelicidin in intestinal epithelium may be crucial for controlling enteric infectious diseases.

Topics: Anti-Infective Agents; Antimicrobial Cationic Peptides; Cathelicidins; Cell Line, Tumor; Colon; Escherichia coli; Escherichia coli Infections; Gene Expression Regulation; Humans; Interleukin-8; Intestinal Mucosa; Lipopolysaccharides; MAP Kinase Signaling System; Oligodeoxyribonucleotides; Toll-Like Receptor 4; Toll-Like Receptor 9

Of all B2 phylogroup strains assessed, 79% could survive and replicate in macrophages. Among them, 11/41 strains (5 CD, 2 UCs, 5 non-IBD) also adhere to and invade epithelial cells, a phenotype assigning them to the AIEC pathovar. The AIEC strains were phylogenetically heterogeneous. We did not identify a gene (or nucleic acid base composition differences) common to all, or the majority of, AIEC. Cytokine secretion and CRISPRs were not associated with the AIEC phenotype.. Comparative genomic analysis of AIEC and non-AIEC strains did not identify a molecular property exclusive to the AIEC phenotype. We recommend a broader approach to the identification of the bacteria-host interactions that are important in the pathogenesis of Crohn's disease.

Topics: Bacterial Adhesion; Cell Line; Clustered Regularly Interspaced Short Palindromic Repeats; Crohn Disease; Cytokines; DNA, Bacterial; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Genome; Host-Pathogen Interactions; Humans; Interleukin-10; Interleukin-6; Interleukin-8; Macrophages; Phenotype; Phylogeny; Sequence Analysis, DNA; Tumor Necrosis Factor-alpha

The inhibitory effects of Zanthoxylum bungeanum essential oil (ZBEO) on Escherichia coli (E. coli) in vitro and in vivo were investigated, as well as its function of improvement of intestinal health. The results of in vitro studies, such as minimal inhibitory concentration (MIC) analysis, agar disc diffusion test and growth curve analysis of E. coli, showed that ZBEO had an excellent inhibitory effect on the growth of E. coli, which may be related to the loss of the normal shape of the cell membranes and the leakage of intracellular constituents, on the basis of SEM observation and cell constituents' release assay. ZBEO also had an inhibitory effect on enteritis and intestinal dysfunction induced by infection of E. coli in vivo, and histopathological observation indicated that ZBEO could markedly ameliorate the structural destruction of intestinal tissues, which might be related to its inhibitory effect on the gene expression of inflammatory cytokines (TLR2, TLR4, TNF

Topics: Animals; Escherichia coli; Escherichia coli Infections; Humans; Interleukin-8; Intestinal Mucosa; Intestines; Mice; Oils, Volatile; Plant Extracts; Toll-Like Receptor 2; Zanthoxylum

A major cause of enteric infection, Gram-negative pathogenic bacteria activate mucosal inflammation through lipopolysaccharide (LPS) binding to intestinal toll-like receptor 4 (TLR4). Breast feeding lowers risk of disease, and human milk modulates inflammation.. This study tested whether human milk oligosaccharides (HMOSs) influence pathogenic Escherichia coli-induced interleukin (IL)-8 release by intestinal epithelial cells (IECs), identified specific proinflammatory signalling molecules modulated by HMOSs, specified the active HMOS and determined its mechanism of action.. Models of inflammation were IECs invaded by type 1 pili enterotoxigenic E. coli (ETEC) in vitro: T84 modelled mature, and H4 modelled immature IECs. LPS-induced signalling molecules co-varying with IL-8 release in the presence or absence of HMOSs were identified. Knockdown and overexpression verified signalling mediators. The oligosaccharide responsible for altered signalling was identified.. HMOSs attenuated LPS-dependent induction of IL-8 caused by ETEC, uropathogenic E. coli, and adherent-invasive E. coli (AIEC) infection, and suppressed CD14 transcription and translation. CD14 knockdown recapitulated HMOS-induced attenuation. Overexpression of CD14 increased the inflammatory response to ETEC and sensitivity to inhibition by HMOSs. 2'-fucosyllactose (2'-FL), at milk concentrations, displayed equivalent ability as total HMOSs to suppress CD14 expression, and protected AIEC-infected mice.. HMOSs and 2'-FL directly inhibit LPS-mediated inflammation during ETEC invasion of T84 and H4 IECs through attenuation of CD14 induction. CD14 expression mediates LPS-TLR4 stimulation of portions of the 'macrophage migration inhibitory factors' inflammatory pathway via suppressors of cytokine signalling 2/signal transducer and activator of transcription 3/NF-κB. HMOS direct inhibition of inflammation supports its functioning as an innate immune system whereby the mother protects her vulnerable neonate through her milk. 2'-FL, a principal HMOS, quenches inflammatory signalling.

Topics: Animals; Cell Line; Enterocytes; Enterotoxigenic Escherichia coli; Enzyme-Linked Immunosorbent Assay; Escherichia coli Infections; Female; Flow Cytometry; Humans; Interleukin-8; Lipopolysaccharide Receptors; Lipopolysaccharides; Mice; Mice, Inbred C57BL; Milk, Human; Reverse Transcriptase Polymerase Chain Reaction; Toll-Like Receptor 4; Trisaccharides

Infection with enterohemorrhagic Escherichia coli (EHEC) can result in severe disease, including hemorrhagic colitis and the hemolytic uremic syndrome. Shiga toxins (Stx) are the key EHEC virulence determinant contributing to severe disease. Despite inhibiting protein synthesis, Shiga toxins paradoxically induce the expression of proinflammatory cytokines from various cell types in vitro, including intestinal epithelial cells (IECs). This effect is mediated in large part by the ribotoxic stress response (RSR). The Shiga toxin-induced RSR is known to involve the activation of the stress-activated protein kinases (SAPKs) p38 and JNK. In some cell types, Stx also can induce the classical mitogen-activated protein kinases (MAPKs) or ERK1/2, but the mechanism(s) by which this activation occurs is unknown. In this study, we investigated the mechanism by which Stx activates ERK1/2s in IECs and the contribution of ERK1/2 activation to interleukin-8 (IL-8) expression. We demonstrate that Stx1 activates ERK1/2 in a biphasic manner: the first phase occurs in response to StxB1 subunit, while the second phase requires StxA1 subunit activity. We show that the A subunit-dependent ERK1/2 activation is mediated through ZAK-dependent signaling, and inhibition of ERK1/2 activation via the MEK1/2 inhibitors U0126 and PD98059 results in decreased Stx1-mediated IL-8 mRNA. Finally, we demonstrate that ERK1/2 are activated in vivo in the colon of Stx2-intoxicated infant rabbits, a model in which Stx2 induces a primarily neutrophilic inflammatory response. Together, our data support a role for ERK1/2 activation in the development of Stx-mediated intestinal inflammation.

Topics: Animals; Butadienes; Cell Line; Enterohemorrhagic Escherichia coli; Enzyme Activation; Epithelial Cells; Escherichia coli Infections; Extracellular Signal-Regulated MAP Kinases; Flavonoids; Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating); HEK293 Cells; Hemolytic-Uremic Syndrome; Humans; Inflammation; Interleukin-8; Intestinal Mucosa; JNK Mitogen-Activated Protein Kinases; MAP Kinase Kinase Kinases; Nitriles; p38 Mitogen-Activated Protein Kinases; Protein Kinases; Rabbits; RNA Interference; RNA, Small Interfering; Shiga Toxin 1; Shiga Toxin 2

Krill oil is a marine derived oil rich in phospholipids, astaxanthin and omega-3 fatty acids. Several studies have found benefits of krill oil against oxidative and inflammatory damage.. We aimed at assessing the ability of krill oil to reduce intestinal inflammation by improving epithelial barrier integrity, increasing cell survival and reducing pathogenicity of adherent-invasive Escherichia coli.. CACO2 and HT29 cells were exposed to cytomix (TNFα and IFNγ) to induce inflammation and co-exposed to cytomix and krill oil. E-cadherin, ZO-1 and F-actin levels were analyzed by immunofluorescence to assess barrier integrity. Scratch test was performed to measure wound healing. Cell survival was analyzed by flow cytometry. Adherent-invasive Escherichia coli LF82 was used for adhesion/invasion assay.. In inflamed cells E-cadherin and ZO-1 decreased, with loss of cell-cell adhesion, and F-actin polymerization increased stress fibres; krill oil restored initial conditions and improved wound healing, reduced bacterial adhesion/invasion in epithelial cells and survival within macrophages; krill oil reduced LF82-induced mRNA expression of pro-inflammatory cytokines.. Krill oil improves intestinal barrier integrity and epithelial restitution during inflammation and controls bacterial adhesion and invasion to epithelial cells. Thus, krill oil may represent an innovative tool to reduce intestinal inflammation.

Topics: Actins; Animals; Bacterial Adhesion; Caco-2 Cells; Cadherins; Cell Adhesion; Cell Death; Cell Survival; Escherichia coli; Escherichia coli Infections; Euphausiacea; Fatty Acids, Omega-3; HT29 Cells; Humans; Interferon-gamma; Interleukin-8; Mice; Microbial Viability; RAW 264.7 Cells; RNA, Messenger; Tumor Necrosis Factor-alpha; Wound Healing; Zonula Occludens-1 Protein

Diabetes mellitus (DM) during pregnancy causes congenital malformation, macrosomia, respiratory distress syndrome, and other abnormalities in neonates, but whether maternal DM affects the neonatal innate immune system is unknown. Therefore we aimed to reveal the influence of DM in pregnancy on the toll-like receptor (TLR)-mediated innate immune response in neonates. Cord blood was collected after full-term vaginal or cesarean delivery and classified into a DM group (n=8) and non-DM (control) group (n=7). Mononuclear cells were harvested from cord blood by using density gradient centrifugation, after which anti-CD14 magnetic beads were used to isolate monocytes from the mononuclear population. After monocytes were cultured with lipopolysaccharide (TLR4 ligand), flagellin (TLR5 ligand), Pam3CSK4 (TLR1/TLR2 ligand), zymosan (TLR2/TLR6 ligand), or macrophage-activating lipopeptide (TLR2/TLR6 ligand) for 12h, the cytokine levels (interleukin [IL]-8, IL-6, IL-1β, IL-10, tumor necrosis factor alpha and IL-12) in the culture supernatants were measured. Compared with the control group, the DM group had higher concentrations of IL-8 (P=0.01) and tumor necrosis factor alpha (P=0.02) after monocyte cultures were stimulated with Pam3CSK4 and higher concentrations of IL-8 (P=0.01) after flagellin treatment. In contrast, stimulation with lipopolysaccharide, zymosan, or macrophage-activating lipopeptide did not lead to any difference in cytokine profiles between the two groups. These data indicate that maternal DM induces excessive inflammatory activation in neonates via a TLR5- or TLR1/2-mediated innate immune response.

Topics: Adult; Cells, Cultured; Escherichia coli Infections; Female; Humans; Immunity, Innate; Infant, Newborn; Infant, Newborn, Diseases; Interleukin-8; Listeriosis; Monocytes; Obesity; Pregnancy; Pregnancy in Diabetics; Toll-Like Receptor 1; Toll-Like Receptor 2; Toll-Like Receptor 5; Tumor Necrosis Factor-alpha

Enterohemorrhagic E.coli (EHEC) is an important foodborne pathogen in the developed world and can cause life-threatening disease particularly in children. EHEC persists in the human gut by adhering intimately to colonic epithelium and forming characteristic attaching/effacing lesions. In this study, we investigated the innate immune response to EHEC infection with particular focus on antimicrobial peptide and protein expression by colonic epithelium. Using a novel human colonic biopsy model and polarized T84 colon carcinoma cells, we found that EHEC infection induced expression of human β-defensin 2 (hBD2), whereas hBD1, hBD3, LL-37, and lysozyme remained unchanged. Infection with specific EHEC deletion mutants demonstrated that this was dependent on flagellin, and apical exposure to purified flagellin was sufficient to stimulate hBD2 and also interleukin (IL)-8 expression ex vivo and in vitro. Flagellin-mediated hBD2 induction was significantly reduced by inhibitors of NF-κB, MAP kinase p38 and JNK but not ERK1/2. Interestingly, IL-8 secretion by polarized T84 cells was vectorial depending on the side of stimulation, and apical exposure to EHEC or flagellin resulted in apical IL-8 release. Our results demonstrate that EHEC only induces a modest immune response in human colonic epithelium characterized by flagellin-dependent induction of hBD2 and low levels of IL-8.

Topics: Adhesins, Bacterial; Anti-Infective Agents; Bacterial Adhesion; beta-Defensins; Biopsy; Cell Line, Tumor; Colon; Colonic Neoplasms; Enterohemorrhagic Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Flagellin; Gene Expression Regulation, Bacterial; Humans; Immunity, Innate; Interleukin-8; Intestinal Mucosa; NF-kappa B; Sequence Deletion

Enteroaggregative Escherichia coli (EAEC) is an important diarrhoeal pathogen causing diseases in multiple epidemiological and clinical settings. In developing countries like India, diarrhoeal diseases are one of the major killers among paediatric population and oddly, few studies are available from Indian paediatric population on the variability of EAEC virulence genes. In this study, we examined the distribution of plasmid and chromosomal-encoded virulence determinants in EAEC isolates, and analysed cytokines response generated against EAEC with specific aggregative adherence fimbriae (AAF) type in duodenal biopsies using in vitro organ culture (IVOC) mimicking in vivo conditions. Different virulence marker combinations among strains were reflected as a function of specific adhesins signifying EAEC heterogeneity. fis gene emerged as an important genetic marker apart from aggA and aap Further, EAEC infection in IVOC showed upregulation of IL-8, IL-1β, IL-6, TNF-α and TLR-5 expression. EAEC with AAFII induced significant TLR-5 and IL-8 response, conceivably owing to more pathogenicity markers. This study sheds light on the pattern of EAEC pathotypes prevalent in North Indian paediatric population and highlights the presence of unique virulence combinations in pathogenic strains. Thus, evident diversity in EAEC virulence and multifaceted bacteria-host crosstalk can provide useful insights for the strategic management of diarrhoeal diseases in India, where diarrhoeal outbreaks are more frequent.

Topics: Bacterial Adhesion; Biopsy; Child; Cytokines; Diarrhea; Duodenum; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Factor For Inversion Stimulation Protein; Feces; Fimbriae Proteins; Fimbriae, Bacterial; Host-Pathogen Interactions; Humans; India; Inflammation; Interleukin-6; Interleukin-8; Organ Culture Techniques; Tumor Necrosis Factor-alpha; Virulence; Virulence Factors

Bacterial colonization of epithelial surfaces and subsequent transmigration across the mucosal barrier are essential for the development of infection. We hypothesized that the methyl-accepting proteins (MCPs), known as chemoreceptors expressed on Escherichia coli (E. coli) bacterial surface, play an important role in mediating bacterial transmigration. We demonstrated a direct interaction between human interleukin-8 (IL-8) and Tsr receptor, a major MCP chemoreceptor. Stimulation of human lung epithelial cell monolayer with IL-8 resulted in increased E. coli adhesion and transmigration of the native strain (RP437) and a strain expressing only Tsr (UU2373), as compared to a strain (UU2599) with Tsr truncation. The augmented E. coli adhesion and migration was associated with a higher expression of carcinoembryonic antigen-related cell adhesion molecule 6 and production of inflammatory cytokines/chemokines, and a lower expression of the tight junction protein claudin-1 and the plasma membrane protein caveolin-1 in lung epithelial cells. An increased E. coli colonization and pulmonary cytokine production induced by the RP437 and UU2373 strains was attenuated in mice challenged with the UU2599 strain. Our results suggest a critical role of the E. coli Tsr chemoreceptor in mediating bacterial colonization and transmigration across human lung epithelium during development of pulmonary infections.

Topics: Antigens, CD; Bacterial Adhesion; Bacterial Proteins; Caveolin 1; Cell Adhesion Molecules; Cells, Cultured; Claudin-1; Cytokines; Escherichia coli; Escherichia coli Infections; GPI-Linked Proteins; Humans; Inflammation Mediators; Interleukin-8; Lung; Membrane Proteins; Organisms, Genetically Modified; Respiratory Mucosa; Sequence Deletion; Transendothelial and Transepithelial Migration

Inadequate and delayed antibiotic treatment of extended spectrum beta-lactamase (ESBL)-producing isolates have been associated with increased mortality of affected patients. The purpose of this study was to compare the host response of human renal epithelial cells and polymorphonuclear leucocyte (PMN) cells when infected by ESBL-producing uropathogenic Escherichia coli (UPEC) isolates in the presence or absence of ineffective antibiotics. The renal epithelial cell line A498 and PMN cells were stimulated with ESBL-producing UPEC isolates in the presence or absence of three different antibiotics (trimetoprim, ceftibuten and ciprofloxacin). Host cell responses were evaluated as release of cytokines (IL-6, IL-8), reactive oxygen species (ROS), ATP and endotoxins. Bacterial morphology and PMN phagocytosis were evaluated by microscopy. In the presence of ceftibuten, 2 out of 3 examined ESBL-isolates changed their morphology into a filamentous form. The presence of ceftibuten enhanced IL-6, IL-8 and ROS-production from host cells, but only from cells stimulated by the filamentous isolates. The bacterial supernatant and not the filamentous bacteria per se was responsible for the increased release of IL-6, IL-8 and ROS. Increased endotoxin and ATP levels were found in the bacterial supernatants from filamentous isolates. Apyrase decreased IL-6 secretion from A498 cells and polymyxin B abolished the increased ROS-production from PMN cells. PMN were able to inhibit the bacterial growth of some ESBL-isolates in the presence of ceftibuten. In conclusion, antibiotic-induced filamentation of ESBL-producing UPEC isolates and the associated release of ATP and endotoxins can alter the host cell response in the urinary tract.

Topics: Adenosine Triphosphate; Anti-Bacterial Agents; beta-Lactamases; Ceftibuten; Cell Line; Cephalosporins; Escherichia coli Infections; Escherichia coli Proteins; Humans; Interleukin-6; Interleukin-8; Reactive Oxygen Species; Uropathogenic Escherichia coli

Ileal lesions of Crohn's disease [CD] patients are colonised by adherent-invasive Escherichia coli [AIEC] able to survive in macrophage cell lines. We analysed the ability of monocyte-derived macrophages [MDM] from CD patients to control AIEC intracellular replication and the pro-inflammatory cytokine response of the infected-MDM.. Peripheral blood MDM were obtained from 24 CD genotyped for NOD2 and ATG16L1 mutations, 5 ulcerative colitis [UC] patients and 12 healthy controls [HC]. The numbers of intracellular bacteria were determined using gentamicin assay. Cytokine secretion was quantified by ELISA assay.. We observed that higher levels of bacteria were internalised within MDM from CD patients than MDM from HC or UC patients. MDM from CD patients were unable to restrict AIEC intracellular replication. Infection of MDM from CD patients with AIEC resulted in significantly increased secretion of IL-6 and tumour necrosis factor alpha [TNF α] than did infection with non-pathogenic E. coli. AIEC-infected MDM from CD patients exhibited a disordered cytokines secretion compared with MDM from UC patients and HC. AIEC-infected MDM from patients with quiescent CD released significantly higher amounts of IL-6 and TNF-alpha than those with active disease or those from HC. The level of secreted TNF-alpha was correlated to the number of intracellular AIEC in MDM from CD patients. Treatment of MDM with infliximab did not change the MDM behaviour.. MDM from CD patients are unable to restrict intracellular AIEC replication, leading to disordered inflammatory response influenced by disease activity.

Topics: Adult; Anti-Inflammatory Agents, Non-Steroidal; Autophagy-Related Proteins; Bacterial Adhesion; Bacterial Load; Carrier Proteins; Cells, Cultured; Colitis, Ulcerative; Crohn Disease; Escherichia coli; Escherichia coli Infections; Female; Genotype; Humans; Infliximab; Interleukin-6; Interleukin-8; Macrophages; Male; Middle Aged; Nod2 Signaling Adaptor Protein; Polymorphism, Single Nucleotide; Tumor Necrosis Factor-alpha

Increased reactive oxygen species (ROS) production is associated with inflamed ileal lesions in Crohn's disease colonized by pathogenic adherent-invasive Escherichia coli LF82. We investigated whether such ileal bacteria can modulate ROS production by epithelial cells, thus impacting on inflammation and mucin expression.. Ileal bacteria from patients with Crohn's disease were incubated with cultured epithelial T84 cells, and ROS production was assayed using the luminol-amplified chemiluminescence method. The gentamicin protection assay was used for bacterial invasion of T84 cell. The expression of NADPH oxidase (NOX) subunits, mucin, and IL-8 was analyzed by quantitative real-time PCR and Western blots. Involvement of NOX and ROS was analyzed using diphenyleneiodonium (DPI) and N-acetylcysteine (NAC).. Among different bacteria tested, only LF82 induced an increase of ROS production by T84 cells in a dose-dependent manner. This response was inhibited by DPI and NAC. Heat- or ethanol-attenuated LF82 bacteria and the mutant LF82ΔFimA, which does not express pili type 1 and poorly adheres to epithelial cells, did not induce the oxidative response. The LF82-induced oxidative response coincides with its invasion in T84 cells, and both processes were inhibited by DPI. Also, we observed an increased expression of NOX1 and NOXO1 in response to LF82 bacteria versus the mutant LF82ΔFimA. Furthermore, LF82 inhibited mucin gene expression (MUC2 and MUC5AC) in T84 cells while increasing the chemotactic IL-8 expression, both in a DPI-sensitive manner.. Adherent-invasive E. coli LF82 induced ROS production by intestinal NADPH oxidase and altered mucin and IL-8 expression, leading to perpetuation of inflammatory lesions in Crohn's disease.

Topics: Colorectal Neoplasms; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Humans; Ileum; Immunoenzyme Techniques; Interleukin-8; Intestinal Mucosa; Intestines; Mucin 5AC; Mucin-2; NADPH Oxidase 1; NADPH Oxidases; Reactive Oxygen Species; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tumor Cells, Cultured

To study the role of outer membrane protein T (OmpT) in the pathogenesis of uropathogenic Escherichia.coli.. In cultured human bladder epithelial cell line 5637, we examined the adhesion ability of wild-type (CFT073), ompT gene knockout (COTD), and revertant (pST) strains of E.coli to the cells and the extracellular matrix (ECM). The expressions of the adhesion gene iha and virulence gene iroN were detected by real-time PCR. Murine models of urinary tract infection with the 3 strains were established to evaluate the bacterial burden of the bladder and kidney tissue and bacterial counts in blood. We also detected the expressions of interleukin-6 (IL-6) and IL-8 in the bladder and kidney tissues of the mice.. The COTD strain showed a significantly lower cell adhesion rate than CFT073 strain [(4.62∓0.39)% vs (8.81∓1.13)%, P<0.05] with also a lower ECM-adhesion rate [(4.95∓0.59)% vs (8.85∓0.79)%, P<0.05]. The mRNA expressions of iha and iroN in CFT073 strain were 2.1 and 3.8 times that of COTD strain. In the mouse model, the mean bacterial load of CFT073 strain in the bladder tissue was 6.36∓0.06, significantly greater than that of COTD (6.01∓0.07) and revertant (6.29∓0.06) strains (P<0.05); the bacterial load of CFT073 strain in the kidney tissue was also significantly higher than that of COTD strain (6.25∓0.05 vs 5.87∓0.06, P<0.05). In mice infected with the wild-type, knockout, and revertant strains, the detection rates of IL-6, which were identical to those of IL-8, in the inflammatory bladder and kidney tissues were 60%, 12.5%, and 50%, respectively.. OmpT may regulate the expression of the adhesion gene iha and the transferrin gene iroN to affect the adhesion of uropathogenic E.coli to host cells.

Topics: Animals; Bacterial Adhesion; Bacterial Load; Bacterial Outer Membrane Proteins; Cell Line, Tumor; Escherichia coli Infections; Escherichia coli Proteins; Gene Knockout Techniques; Humans; Inflammation; Interleukin-6; Interleukin-8; Kidney; Mice; Peptide Hydrolases; Receptors, Cell Surface; Urinary Bladder; Urinary Tract Infections; Uropathogenic Escherichia coli

Crohn's disease is a multifactorial disease in which an aberrant immune response to commensal intestinal microbiota leads to chronic inflammation. The small intestine of patients with Crohn's disease is colonized by a group of adherent-invasive Escherichia coli strongly able to adhere and invade intestinal epithelial cells lactoferrin is an iron-binding glycoprotein known to have anti-bacterial and anti-inflammatory activities.. We explore the ability of bovine lactoferrin to modulate the interactions between the adherent-invasive E. coli strain LF82 and intestinal epithelial cells as well as the inflammatory response.. Bacterial adhesion and invasion assays were used to assess the antimicrobial activity of lactoferrin. Electron microscopy was used to characterize bacteria-cell interactions. The mRNA expression of pro-inflammatory cytokines was measured both in cultured cells and in biopsies taken from intestine of patients affected by Crohn's disease.. Lactoferrin inhibited bacterial invasion through minimally affecting adhesion. This divergence was due to a mannose-dependent lactoferrin binding to the bacterial type 1 pili and consequent bacterial aggregation on the intestinal epithelial cell surface. Expression of pro-inflammatory cytokines, such as TNF-alpha, IL-8, and IL-6, was markedly inhibited by lactoferrin both in cultured and Crohn-derived intestinal cells.. Bovine lactoferrin might function via an antibacterial and/or anti-inflammatory mechanism in the treatment of Crohn's disease.

Topics: Animals; Anti-Infective Agents; Bacterial Adhesion; Caco-2 Cells; Cattle; Crohn Disease; Escherichia coli; Escherichia coli Infections; Gene Expression; Humans; Interferon-gamma; Interleukin-6; Interleukin-8; Intestinal Mucosa; Lactoferrin; Mannose; Microscopy, Electron, Scanning; Microscopy, Electron, Transmission; Receptors, Immunologic; RNA, Messenger; Tumor Necrosis Factor-alpha

To identify a fast, economic and reliable method for preselecting lactic acid-producing bacterial (LAB) isolates to control enterotoxigenic Escherichia coli (ETEC).. Two assays with porcine intestinal epithelial IPEC-J2 cells or Caenorhabditis elegans for selecting effective probiotic candidates were compared. Both assays were based on measuring death of cells or worms caused by ETEC strain JG280. Six of 13 LAB isolates showed ≥50% protection in each assay, among which only four isolates (≥50% protection) were consistently selected by both assays. Isolate CL9 (Lactobacillus reuteri) was further studied. It reduced gene expression of estA, estB and elt in JG280 in both assays. Furthermore, the isolate protected IPEC-J2 and C. elegans from cell and worm death caused by STa, STb or LT enterotoxin expressed in E. coli DH5α. CL9 also promoted host defensive responses by decreasing IL-8 and increasing IL-10 production in IPEC-J2 cells and expression of antimicrobial peptide genes clec-60, clec-85 in C. elegans.. Caenorhabditis elegans is useful for preselecting probiotic candidates to control ETEC after initial screening with IPEC-J2 cells.. A combination of IPEC-J2 cell and C. elegans assays can improve the effectiveness in preselecting probiotic candidates.

Topics: Animals; Antibiosis; Antimicrobial Cationic Peptides; Caenorhabditis elegans; Cell Line; Enterotoxigenic Escherichia coli; Enterotoxins; Epithelial Cells; Escherichia coli Infections; Interleukin-10; Interleukin-8; Intestines; Limosilactobacillus reuteri; Models, Biological; Probiotics; Swine; Swine Diseases

A Shiga toxin type 2a (Stx2a)-producing enteroaggregative Escherichia coli (EAEC) strain of serotype O104:H4 caused a large outbreak in 2011 in northern Europe. Pathogenic mechanisms for this strain are unclear. We hypothesized that EAEC genes encoded on the pAA virulence plasmid promoted the translocation of Stx2a across the intestinal mucosa.. We investigated the potential contribution of pAA by using mutants of Stx-EAEC strain C227-11, either cured of the pAA plasmid or deleted for individual known pAA-encoded virulence genes (ie, aggR, aggA, and sepA). The resulting mutants were tested for their ability to induce interleukin 8 (IL-8) secretion and translocation of Stx2a across a polarized colonic epithelial (T84 cell) monolayer.. We found that deletion of aggR or aggA significantly reduced bacterial adherence and (independently) translocation of Stx2a across the T84-cell monolayer. Moreover, deletion of aggR, aggA, sepA, or the Stx2a-encoding phage from C227-11 resulted in reduced secretion of IL-8 from the infected monolayer.. Our data suggest that the AggR-regulated aggregative adherence fimbriae I enhance inflammation and enable the outbreak strain to both adhere to epithelial cells and translocate Stx2a across the intestinal epithelium.

Topics: Bacterial Adhesion; Cell Line; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Gene Deletion; Genotype; Germany; Humans; Interleukin-8; Plasmids; Protein Transport; Serogroup; Shiga Toxin 2; Trans-Activators; Virulence Factors

Enterohemorrhagic Escherichia coli (EHEC) strains are causative agents of diarrhea and hemorrhagic colitis, both diseases associated with intestinal inflammation and cell damage. Several studies have correlated EHEC virulence factors to high levels of intestinal pro-inflammatory cytokines and we have previously described that the Long polar fimbriae (Lpf) is involved in the secretion of interleukin-8 (IL-8) and up-regulation of genes belonging to the NF-κB pathway using non-polarized epithelial intestinal T84 cells. In the current study, we evaluated the two EHEC O157 Lpf fimbriae (Lpf1 and Lpf2) for their ability to induce intestinal secretion of IL-8 and the activation of IL8, CCL20, and ICAM1 genes on polarized T84 cells. We also determined the participation of Lpf1 and Lpf2 in transepithelial migration of polymorphonuclear neutrophils (PMNs). Polarized T84 cells infected with EHEC revealed that both, Lpf1 and Lpf2, were required for the secretion of IL-8 and the induction of IL8, CCL20, and ICAM1 genes. Both fimbriae also played a role in the migration of PMNs trough the intestinal cells monolayer. Overall, the present work further demonstrated that the fimbriae Lpf1 and Lpf2 are important bacterial virulence factors that might be involved in the inflammatory responses associated with EHEC infections.

Topics: Cell Movement; Chemokine CCL20; Epithelial Cells; Escherichia coli Infections; Escherichia coli O157; Escherichia coli Proteins; Fimbriae Proteins; Fimbriae, Bacterial; Humans; Intercellular Adhesion Molecule-1; Interleukin-8; Intestinal Diseases; Intestines; Neutrophils

Bacterial sepsis induced immunsuppression via antigen hyporesponsibility increases the risk of nosokomial infections and mortality. Pattern recognition receptors (PRR) might have a central role in the pathophysiology of hyporesponsibility.. In this study we evaluated in a human E. coli sepsis cohort, the role of PRR including TLR's and Trem-1. Expression of Trem-1, TLR2, TLR4, CD14 and HLA-DR on blood monozytes and neutrophils were examined using flow cytometry from 22 patients with E. coli sepsis and 6 healthy controls. LPS and LTA stimulated TNF alpha, IL-10, IL-8 and IL-6 production was studied in a 24 h whole blood assay. Free cytokine serum concentration of TNF alpha, PCT and IP-10 were evaluated.. We found a significant higher expression of Trem-1 and TLR-2 on monocytes and neutrophils in patients compared to healthy volunteers. TLR2 expression (p < 0.05) was higher and HLA-DR lower (p < 0.05) on monocytes of patients with severe sepsis compared to patients with simple sepsis. Trem-1 expression was tendentially higher (p = 0,07) on monocytes and lower on neutrophils of patients with severe sepsis. Trem-1 expression on neutrophils was associated with the IL-10 (LPS: r = 0,61, p < 0.02) and TNF-α inducibility (LPS: r = 0,78, p < 0,002). In addition Trem-1 expression on neutrophils shows a negative correlation to the serum levels of TNF alpha (r = -0,63; p < 0,005), IP-10 (r = -0,5; p < 0,035) and procalcitonin (r = -0,59; p < 0,007).. Patients with E. coli sepsis are characterized by an association of Trem-1 expression on blood neutrophils with cytokine inducibility. The TREM-1 pathway on neutrophils might play a role in producing an adequate inflammatory and bactericidal response in bacterial sepsis.. The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/4441869398748313.

Topics: Aged; Case-Control Studies; Cytokines; Escherichia coli Infections; Female; Flow Cytometry; HLA-DR Antigens; Humans; Immune Tolerance; Interleukin-10; Interleukin-6; Interleukin-8; Lipopolysaccharide Receptors; Male; Membrane Glycoproteins; Monocytes; Neutrophils; Prospective Studies; Receptors, Immunologic; Sepsis; Toll-Like Receptor 2; Triggering Receptor Expressed on Myeloid Cells-1; Tumor Necrosis Factor-alpha; Up-Regulation

Complement and the TLR family constitute two important branches of innate immunity. We previously showed attenuating effects on inflammation and thromogenicity by inhibiting the TLR coreceptor CD14 in porcine sepsis. In the present study, we explored the effect of the C5 and leukotriene B4 inhibitor Ornithodoros moubata complement inhibitor (OmCI; also known as coversin) alone and combined with anti-CD14 on the early inflammatory, hemostatic, and hemodynamic responses in porcine Escherichia coli-induced sepsis. Pigs were randomly allocated to negative controls (n = 6), positive controls (n = 8), intervention with OmCI (n = 8), or with OmCI and anti-CD14 (n = 8). OmCI ablated C5 activation and formation of the terminal complement complex and significantly decreased leukotriene B4 levels in septic pigs. Granulocyte tissue factor expression, formation of thrombin-antithrombin complexes (p < 0.001), and formation of TNF-α and IL-6 (p < 0.05) were efficiently inhibited by OmCI alone and abolished or strongly attenuated by the combination of OmCI and anti-CD14 (p < 0.001 for all). Additionally, the combined therapy attenuated the formation of plasminogen activator inhibitor-1 (p < 0.05), IL-1β, and IL-8, increased the formation of IL-10, and abolished the expression of wCD11R3 (CD11b) and the fall in neutrophil cell count (p < 0.001 for all). Finally, OmCI combined with anti-CD14 delayed increases in heart rate by 60 min (p < 0.05) and mean pulmonary artery pressure by 30 min (p < 0.01). Ex vivo studies confirmed the additional effect of combining anti-CD14 with OmCI. In conclusion, upstream inhibition of the key innate immunity molecules, C5 and CD14, is a potential broad-acting treatment regimen in sepsis as it efficiently attenuated inflammation and thrombogenicity and delayed hemodynamic changes.

Topics: Animals; Antithrombin III; Arterial Pressure; Arthropod Proteins; Carrier Proteins; CD11b Antigen; Complement C5; Escherichia coli; Escherichia coli Infections; Granulocyte-Macrophage Colony-Stimulating Factor; Heart Rate; Hemodynamics; Immunity, Innate; Inflammation; Interleukin-10; Interleukin-1beta; Interleukin-6; Interleukin-8; Leukocyte Count; Leukotriene B4; Lipopolysaccharide Receptors; Neutrophils; Peptide Hydrolases; Plasminogen Activator Inhibitor 1; Sepsis; Sus scrofa; Tumor Necrosis Factor-alpha

Evidence suggests that adjunctive treatment with intravenous immunoglobulin preparations enriched with IgA and IgM reduce mortality in sepsis. The mode of action of polyvalent immunoglobulin is complex, including neutralization of toxins and modulation of complement activation and cytokine formation toward an anti-inflammatory profile. In this study we explored the effect of Pentaglobin, containing IgG, IgA and IgM, on the initial inflammatory reaction as well as on hemodynamics, using a well characterized and standardized porcine model of sepsis. Anesthetized and mechanically ventilated pigs, mean weight 14.9 kg, were allocated into two groups of 8 animals, receiving either Pentaglobin or saline, before sepsis was induced by intravenous Escherichia coli infusion. Five negative controls received saline only. All animals were observed for 4 h under extensive invasive monitoring. Pentaglobin significantly (p < 0.05) attenuated IL-1β formation by 38% at the end of the experiment, and markedly increased (p < 0.05) the formation of IL-10 at 60 min. TNF-α, IL-6, IL-8 and expression of the cell surface marker wCD11R3 were lower in the Pentaglobin group, but the differences were not significant. The serum concentration of LPS was three times higher in the Pentaglobin group (p < 0.005), indicating binding of LPS to Pentaglobin. Complementary in vitro experiments showed a higher binding affinity for IgM and IgA to LPS than for IgG. LPS-induced formation of IL-6 was significantly (p < 0.05) attenuated by Pentaglobin in an in vitro whole blood model. In conclusion, Pentaglobin decreased the key inflammasome IL-1β molecule in an E. coli-model of pigs sepsis.

Topics: Animals; Antigen-Antibody Complex; Biomarkers; Escherichia coli; Escherichia coli Infections; Hemodynamics; Immunoglobulin A; Immunoglobulin M; Immunoglobulins, Intravenous; Inflammation; Interleukin-10; Interleukin-1beta; Interleukin-6; Interleukin-8; Lipopolysaccharides; Protein Binding; Sepsis; Swine; Tumor Necrosis Factor-alpha

The pathogenesis of diffusely adherent Escherichia coli (DAEC) remains to be elucidated. Previously, we found that afimbrial adhesin gene (afa)-positive motile DAEC strains isolated from patients with diarrhoea induce high levels of IL-8 secretion in Caco-2 cells via toll-like receptor 5 (TLR-5), while non-motile strains did not. The aim of this study was to compare virulence properties, including the phylogenetic groups, afa subtypes, IL-8 secretion levels, and the effects on tight junctions, of DAEC strains isolated from healthy persons with those isolated from patients with diarrhoea. Induction of IL-8 secretion in Caco-2 cells was examined for a total of 36 afa-positive strains: 19 from diarrhoeal patients and 17 from healthy carriers. Irrespective of the source, all strains were classified into the phylogenetic group B2 or D, with the exception of two strains. All 7 motile strains isolated from diarrhoeal patients induced high levels of IL-8 secretion, while only 6 of 15 motile strains from healthy carriers induced IL-8 secretion to the same levels as the diarrhoeal strains. We speculated that additional virulence factors other than afa and motility cause the loosening of tight junctions that allows flagellin to reach TLR-5 located on the basolateral side of the epithelium. However, no differences in the TER and dextran permeability were observed between cells infected with diarrhoeal strains and those from healthy persons. Thus, diarrhoeagenic DAEC seems to possess additional factors, in addition to adhesin and flagellin, which can induce high IL-8 secretion.

Topics: Adhesins, Escherichia coli; Bacterial Adhesion; Caco-2 Cells; Diarrhea; DNA, Bacterial; Electric Impedance; Escherichia coli; Escherichia coli Infections; Humans; Interleukin-8; Phylogeny; Polymerase Chain Reaction; Tight Junctions

Wnt signaling plays an essential role in gastrointestinal epithelial proliferation. Most investigations have focused on developmental and immune responses. Bacterial infection can be chronic and increases the risk of inflammatory bowel disease and colitis-associated cancer. However, we lack studies on how bacteria regulate Wnt proteins and how Wnts modulate the host responses to enteric bacteria. This study investigated the effects of Salmonella and Escherichia coli on Wnt2, one of the Wnt family members, in intestinal epithelia cells.. Using cultured epithelial cells, a Salmonella-colitis mouse model, and a gnotobiotic mouse model, we found that Wnt2 mRNA and protein expression levels were elevated after bacterial infection. Enteric bacteria regulate Wnt2 location in the intestine. Furthermore, we found that elevation of Wnt2 was a strategy for host defense by inhibiting cell apoptosis and inflammatory responses to infection.. Using Wnt2 siRNA analysis, we show enhanced inflammatory cytokine IL-8 in epithelial cells. Cells overexpressed Wnt2 had less bacterial-induced IL-8 secretion. AvrA is a bacterial protein that inhibits inflammation by stabilizing β-catenin, the downstream target of Wnt. We found that the stabilization of Wnt2 was regulated through ubiquitination. Moreover, the bacterial protein AvrA from Salmonella and E. coli stabilized Wnt2 protein expression in vivo. In an ex-germ-free system, E. coli F18 expressing AvrA increased Wnt2 expression and changed Wnt2 distribution in intestine.. Wnt2 contributes to host protection in response to enteric bacteria. Our findings thus reveal a previously undefined role of Wnt for host-pathogen interaction and inflammation.

Topics: Animals; Apoptosis; Bacterial Proteins; Cells, Cultured; Colitis; Epithelial Cells; Escherichia coli Infections; Host-Pathogen Interactions; Interleukin-8; Intestinal Mucosa; Mice; RNA, Messenger; RNA, Small Interfering; Salmonella Infections; Salmonella typhimurium; Ubiquitination; Wnt Signaling Pathway; Wnt2 Protein

Crohn's disease (CD) and ulcerative colitis (UC), known as inflammatory bowel diseases (IBD), are characterized by an abnormal immunological response to commensal bacteria colonizing intestinal lumen and mucosa. Among the latter, strains of adherent-invasive Escherichia coli (AIEC), capable of adhering to and invading epithelium, and to replicate in macrophages, have been described in CD adults. We aimed at identifying and characterizing AIEC strains in pediatric IBD.. In all, 24 CD children, 10 UC, and 23 controls were investigated. Mucosal biopsies, taken during colonoscopy, were analyzed for the presence of AIEC strains by an adhesive-invasive test. Protein expression of the specific AIEC receptor, the carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6), was evaluated by western blot and immunohistochemistry, while tumor necrosis factor alpha (TNF-α) and interleukin (IL)-8 mRNA expression was detected by real-time polymerase chain reaction (PCR), after bacterial infection. Transmission electron microscopy and trans-epithelial electric resistance assays were performed on biopsies to assess bacteria-induced morphological and functional epithelial alterations.. Two bacterial strains, EC15 and EC10, were found to adhere and invade the Caco2 cell line, similar to the well-known AIEC strain LF82 (positive control): they upregulated CEACAM6, TNF-α, and IL-8 gene/protein expression, in vitro and in cultured intestinal mucosa; they could also survive inside macrophages and damage the epithelial barrier integrity. Lesions in the inflamed tissues were associated with bacterial infection.. This is the first study showing the presence of adhesive-invasive bacteria strains in the inflamed tissues of children with IBD. Collective features of these strains indicate that they belong to the AIEC spectrum, suggesting their possible role in disease pathogenesis.

Topics: Adolescent; Animals; Antigens, CD; Bacterial Adhesion; Blotting, Western; Case-Control Studies; Cell Adhesion Molecules; Cells, Cultured; Child; Colitis, Ulcerative; Crohn Disease; Escherichia coli; Escherichia coli Infections; Female; Fluorescent Antibody Technique; GPI-Linked Proteins; Humans; Immunoenzyme Techniques; Interleukin-8; Intestinal Mucosa; Macrophages; Male; Mice; Organ Culture Techniques; Real-Time Polymerase Chain Reaction; RNA, Messenger; Tumor Necrosis Factor-alpha

Bacterial flagella contribute to pathogen virulence; however, the role of flagella in the pathogenesis of F18ab E. coli-mediated swine edema disease (ED) is not currently known. We therefore evaluated the role of flagella in F18ab E. coli adhesion, invasion, biofilm formation, and IL-8 production using an in vitro cell infection model approach with gene-deletion mutant and complemented bacterial strains. We demonstrated that the flagellin-deficient fliC mutant had a marked decrease in the ability to adhere to and invade porcine epithelial IPEC-J2 cells. Surprisingly, there was no difference in adhesion between the F18 fimbriae-deficient ΔfedA mutant and its parent strain. In addition, both the ΔfedA and double ΔfliCΔfedA mutants exhibited an increased ability to invade IPEC-J2 cells compared to the wild-type strain, although this may be due to increased expression of other adhesins following the loss of F18ab fimbriae and flagella. Compared to the wild-type strain, the ΔfliC mutant showed significantly reduced ability to form biofilm, whereas the ΔfedA mutant increased biofilm formation. Although ΔfliC, ΔfedA, and ΔfliCΔfedA mutants had a reduced ability to stimulate IL-8 production from infected Caco-2 cells, the ΔfliC mutant impaired this ability to a greater extent than the ΔfedA mutant. The results from this study clearly demonstrate that flagella are required for efficient F18ab E. coli adhesion, invasion, biofilm formation, and IL-8 production in vitro.

Topics: Animals; Bacterial Adhesion; Caco-2 Cells; Edema Disease of Swine; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Fimbriae Proteins; Fimbriae, Bacterial; Flagella; Humans; Interleukin-8; Swine; Virulence; Virulence Factors

Modulation of the immune response is an established feature of certain lactobacilli. CXCL8 is an inflammatory chemokine released by the urinary tract mucosa after contact with uropathogenic Escherichia coli during urinary tract infection and is crucial for proper infiltration of immune cells. Nevertheless, persistently high levels of CXCL8 are associated with pathogenicity and malignancy. In this study, we tested twelve Lactobacillus strains for their ability to influence CXCL8 release from urothelial cells. We evaluated how strains from different Lactobacillus species could regulate CXCL8 in human 5637 urothelial cells, either resting cells or cells concomitantly challenged with heat-killed E. coli. A majority of the tested species altered CXCL8 release from the urothelial cells after 24 hours of stimulation. Most species increased CXCL8 release, whereas a few lactobacilli efficiently suppressed CXCL8 secretion from E. coli-challenged cells. While strong CXCL8 modulators such as Lactobacillus reuteri and Lactobacillus delbrueckii were unable to degrade CXCL8 in the extracellular environment, effects on IL8 transcription were evident for selected lactobacilli. Although IL8 transcription was affected by lactobacilli, the influence on mRNA transcript did not correlate to the impact on CXCL8 release. Phylogenetic analysis based on a 16S rRNA dendrogram of the tested lactobacilli and their effect on CXCL8 revealed some linkage to specific Lactobacillus groups. Testing the immunomodulatory nature of lactobacilli can prove important when selecting new probiotic microbes. Moreover, we believe that phylogenetic and phenotypic similarities could be used to analyse the traits governing such modulation.

Topics: Cell Line; Escherichia coli; Escherichia coli Infections; Gene Expression Regulation; Humans; Interleukin-8; Lactobacillus; Molecular Sequence Data; Phylogeny; Probiotics; Urothelium

Proinflammatory cytokines are related to the pathogenesis of enterohemorrhagic Escherichia coli (EHEC) infection and hemolytic-uremic syndrome (HUS). We assessed the kinetics of the release of cytokines such as neopterin, interleukin (IL)-6, IL-8 and tumour necrosis factor (TNF)-α and the soluble forms of type I and II TNF receptors during EHEC O111-induced HUS (EHEC O111/HUS). Fourteen patients with EHEC O111/HUS were enrolled in this study. Serum concentrations of all cytokines other than TNF-α were significantly elevated in patients with severe HUS compared with those in patients with mild HUS. Although serum concentrations of TNF-α were not significantly higher in patients with severe HUS, most patients with acute encephalopathy showed elevated TNF-α levels. Serum concentrations of these cytokines rapidly and markedly increased, and massive hypercytokinaemia developed 1 day before the diagnosis of HUS in patients with severe HUS. Changes in the number of white blood cells and concentration of serum lactate dehydrogenase were significantly larger between the onset of hemorrhagic colitis and the time of the diagnosis of HUS in patients with severe HUS compared with those in patients with mild HUS. Proinflammatory cytokines play an important role in the pathogenesis of EHEC infection and development of severe complications, including HUS and encephalopathy. Monitoring the cytokine profile may be useful for assessing disease activity of EHEC O111 infections.

Topics: Adolescent; Adult; Child; Cytokines; Enterohemorrhagic Escherichia coli; Escherichia coli Infections; Female; Hemolytic-Uremic Syndrome; Humans; Infant; Interleukin-6; Interleukin-8; L-Lactate Dehydrogenase; Leukocyte Count; Male; Neopterin; Receptors, Tumor Necrosis Factor; Tumor Necrosis Factor-alpha; Young Adult

Rab GTPases are frequent targets of vacuole-living bacterial pathogens for appropriate trafficking of the vacuole. Here we discover that bacterial effectors including VirA from nonvacuole Shigella flexneri and EspG from extracellular Enteropathogenic Escherichia coli (EPEC) harbor TBC-like dual-finger motifs and exhibits potent RabGAP activities. Specific inactivation of Rab1 by VirA/EspG disrupts ER-to-Golgi trafficking. S. flexneri intracellular persistence requires VirA TBC-like GAP activity that mediates bacterial escape from autophagy-mediated host defense. Rab1 inactivation by EspG severely blocks host secretory pathway, resulting in inhibited interleukin-8 secretion from infected cells. Crystal structures of VirA/EspG-Rab1-GDP-aluminum fluoride complexes highlight TBC-like catalytic role for the arginine and glutamine finger residues and reveal a 3D architecture distinct from that of the TBC domain. Structure of Arf6-EspG-Rab1 ternary complex illustrates a pathogenic signaling complex that rewires host Arf signaling to Rab1 inactivation. Structural distinctions of VirA/EspG further predict a possible extensive presence of TBC-like RabGAP effectors in counteracting various host defenses.

Topics: ADP-Ribosylation Factors; Amino Acid Sequence; Animals; Autophagy; Dysentery, Bacillary; Enteropathogenic Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Fibroblasts; GTPase-Activating Proteins; Interleukin-8; Mice; Models, Molecular; Molecular Sequence Data; Protein Structure, Tertiary; Sequence Alignment; Shigella flexneri; Virulence; Virulence Factors

Urinary tract infections (UTIs) are frequently polymicrobial diseases mainly sustained by Escherichia coli in association with other opportunistic pathogens. Cystitis and pyelonephritis are usually accompanied by an inflammatory response, which includes neutrophil recruitment. Uropathogenic E. coli possess the ability to evade host defenses, modulating the innate immune response. The aim of this study was to determine whether particular E. coli strains correlate with polymicrobial bacteriuria and whether escape from the early host defenses and microbial synergy could lead to mixed UTIs. We evaluated 188 E. coli-positive urine samples and assessed the relationships among polymicrobism, neutrophil presence and several traits of E. coli isolates (virulence factors such as hlyA, fimA, papC and their relative products, i.e. hemolysin, type 1 and P fimbriae, and cnf1, their phylogenetic group) and their ability to suppress cytokine response in 5637 bladder epithelial cells. Escherichia coli susceptibility toward quinolones and fluoroquinolones, known to be linked to the pathogenicity of this species, was also considered. We found significant correlations among polymicrobial bacteriuria, absence of pyuria and quinolone/fluoroquinolone susceptibility of E. coli isolates and their enhanced capability to suppress interleukin-8 urothelial production when compared with the patterns induced by the resistant strains.

Topics: Aged; Anti-Bacterial Agents; Bacteriuria; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Female; Humans; Interleukin-8; Male; Microbial Sensitivity Tests; Middle Aged; Opportunistic Infections; Quinolones; Tumor Cells, Cultured; Urinary Bladder; Urinary Tract Infections

Four efficiently translocating Escherichia coli (TEC) strains isolated from the blood of humans (HMLN-1), pigs (PC-1) and rats (KIC-1 and KIC-2) were tested for their ability to adhere and translocate across human gut epithelial Caco-2 and HT-29 cells, to elicit a proinflammatory response and for the presence of 47 pathogenic E. coli virulence genes. HMLN-1 and PC-1 were more efficient in adhesion and translocation than rat strains, had identical biochemical phenotype (BPT) and serotype (O77:H18) and phylogenetic group (D). KIC-2 adhered more than KIC-1, belonged to different BPT and serotype but the same phylogenetic group as KIC-1. TEC strains elicited significantly higher IL-8 response in both cell lines (P < 0.05) and monocytic THP-1 (P < 0.0001) cells than non-TEC strains. KIC-2 induced the highest IL-8 response which may be associated with its immunostimulatory flagellin. Apart from adhesin genes fimH and bmaE that were carried by all strains, HMLN-1 and PC-1 carried capsule synthesis gene kpsMT III and KIC-2 carried the EAST1 toxin gene. The lack of known virulence genes and the ability of TEC to efficiently adhere and translocate whilst causing proinflammatory response suggests that these strains may carry as yet unidentified genes that enable their translocating ability.

Topics: Animals; Bacterial Adhesion; Bacterial Translocation; Caco-2 Cells; Escherichia coli; Escherichia coli Infections; HT29 Cells; Humans; Interleukin-8; Molecular Sequence Data; Phylogeny; Rats; Rodent Diseases; Swine; Swine Diseases; Virulence; Virulence Factors

The aim of this study was to investigate the effects of carbon dioxide (CO(2)) pneumoperitoneum on the intra-abdominal spread of bacteria, the local and systemic cytokine expression, and oxidant/antioxidant status in young rats with bacterial peritonitis.. Young Sprague-Dawley rats, aging 20-27 days and weighing around 50 g, were allocated to six groups of six to nine animals in each. Intra-abdominal infection model was developed by intraperitoneal injection with 1 cc of Escherichia coli (E. coli) (10(8) CFU/mL, ATCC25922 strain) via right lower abdominal wall. Carbon dioxide (CO(2)) pneumoperitoneum was applied to the rats via umbilical pit insufflation with 20 cc CO(2) for 30 min. All survived rats underwent laparotomy and were killed 24 h or 3 days later. Serum levels of CO(2) and CRP were measured. Left lower abdomen peritoneum, peritoneal fluid, mesenteric lymph node of terminal ileum, and liver were taken for bacterial culture. Liver and plasma levels of TNF-α, IL-1β, and IL-6 were examined for the level of local and systemic immunologic response. Oxidant/antioxidant status in liver and plasma were assessed by measuring malondialdehyde (MDA), and reduced to oxidized glutathione ratio (GSH/GSSG).. Carbon dioxide (CO(2)) pneumoperitoneum does not facilitate E. coli dissemination to other intra-abdominal organs in rats with localized E. coli peritonitis. Peritonitis rats that underwent abdominal CO(2) insufflation have insignificantly higher CRP or lower CO(2) levels. Plasma and liver TNF-α, IL-1β concentrations were not significantly different among the four groups, but plasma IL-6 was significantly increased in rats with E. coli peritonitis and CO(2) pneumoperitoneum that were killed 3 days later as compared with that of rats that were killed 24 h later. In rats with E. coli peritonitis, CO(2) pneumoperitoneum was significantly associated with decreased hepatic GSH/GSSG ratio. However, plasma and liver MDA levels were not altered after CO(2) pneumoperitoneum.. Carbon dioxide (CO(2)) pneumoperitoneum is not associated with E. coli dissemination in the presence of local intra-abdominal infection. CO(2) pneumoperitoneum elicited systemic anti-inflammatory response at a specific time period and decreased hepatic antioxidant status in young rats with E. coli peritonitis.

Topics: Analysis of Variance; Animals; C-Reactive Protein; Carbon Dioxide; Escherichia coli Infections; Glutathione; Insufflation; Interleukin-6; Interleukin-8; Liver; Malondialdehyde; Oxidative Stress; Peritonitis; Pneumoperitoneum; Rats; Rats, Sprague-Dawley; Statistics, Nonparametric; Tumor Necrosis Factor-alpha

Two major arms of the inflammatory response are the NF-κB and c-Jun N-terminal kinase (JNK) pathways. Here, we show that enteropathogenic Escherichia coli (EPEC) employs the type III secretion system to target these two signalling arms by injecting host cells with two effector proteins, NleC and NleD. We provide evidence that NleC and NleD are Zn-dependent endopeptidases that specifically clip and inactivate RelA (p65) and JNK, respectively, thus blocking NF-κB and AP-1 activation. We show that NleC and NleD co-operate and complement other EPEC effectors in accomplishing maximal inhibition of IL-8 secretion. This is a remarkable example of a pathogen using multiple effectors to manipulate systematically the host inflammatory response signalling network.

Topics: Apoptosis; Enteropathogenic Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Gene Expression; HeLa Cells; Host-Pathogen Interactions; Humans; Interleukin-8; JNK Mitogen-Activated Protein Kinases; Mitogen-Activated Protein Kinase 9; NF-kappa B; Transcription Factor RelA; Transcription, Genetic; Tumor Necrosis Factor-alpha

Pathogen contact induces quickly in Mammary Epithelial Cells (MEC) the expression of the proinflammatory cytokine IL-8 and delayed that of the bactericidal β-defensin LAP. Both genes encoding these factors feature on their proximal promoter a composite NF-κB/CEBP binding site. We compare here in MEC the role of NF-κB and C/EBP factors in regulating basal and pathogen-induced expression of both genes from cattle. Abrogating NF-κB binding to that site by introduction of a single point mutation blocks promoter activity of both genes in reporter gene assays. Chromatin accessibility PCR and Chromatin immunoprecipitation reveal that the chromatin of the resting LAP promoter is tightly packed and NF-κB p50 homodimer binding prevails. Infection results in chromatin decompaction accompanied by predominant recruitment of NF-κB p65 for promoter activation. Overexpression of transcription factors confirms a stimulatory role of NF-κB p65 but also a repressive function of C/EBPβ for LAP promoter activity. These factors reverse roles to control IL-8 expression. NF-κB p65 homodimers already reside on the resting IL-8 promoter and induction recruits NF-κB p50. Overexpression of both NF-κB factors represses the promoter in MEC, but not in HEK293 cells. Inhibitors of NF-κB activation and nuclear recruitment both tremendously increase basal and pathogen stimulated IL-8 mRNA concentrations in MEC. Mutation of the C/EBP-binding site blocks and overexpression of C/EBPβ stimulates IL-8-promoter activity. Thus, the pathogen-induced fast activation of diverse transcription factors acting through a common promoter binding site is gene specifically differentiated into opposite functional significance for swiftly (IL-8) or slowly (LAP) induced genes in MEC.

Topics: Animals; beta-Defensins; Binding Sites; Cattle; CCAAT-Enhancer-Binding Protein-beta; Chromatin; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Female; Gene Expression Regulation; HEK293 Cells; Humans; Interleukin-8; Kinetics; Mammary Glands, Animal; Mice; NF-kappa B p50 Subunit; Promoter Regions, Genetic; Protein Binding; Repressor Proteins; RNA, Messenger; Transcription Factor RelA

Magnesium sulfate is proposed to have neuroprotective effects in the offspring. We examined the effects of maternal magnesium sulfate administration on maternal and fetal inflammatory responses in a rat model of maternal infection.. Pregnant rats were injected with saline, Gram-negative bacterial endotoxin lipopolysaccharide or lipopolysaccharide with magnesium sulfate (pre- and/or after lipopolysaccharide) to mimic infection. Maternal blood, amniotic fluid, fetal blood, and fetal brains were collected 4 hours after lipopolysaccharide and assayed for tumor necrosis factor, interleukin-6, monocyte chemoattractant protein-1, and growth-related oncogene-KC. In addition, the effect of magnesium sulfate on cytokine production by an astrocytoma cell line was assessed.. Lipopolysaccharide administration induced tumor necrosis factor, interleukin-6, monocyte chemoattractant protein-1, and growth-related oncogene-KC expression in maternal and fetal compartments. Maternal magnesium sulfate treatment significantly attenuated lipopolysaccharide-induced multiple proinflammatory mediator levels in maternal and fetal compartments.. Antenatal magnesium sulfate administration significantly ameliorated maternal, fetal, and gestational tissue-associated inflammatory responses in an experimental model of maternal infection.

Topics: Amniotic Fluid; Animals; Brain; Chemokine CCL2; Chemokine CXCL1; Disease Models, Animal; Escherichia coli Infections; Female; Inflammation; Interleukin-6; Interleukin-8; Magnesium Sulfate; Neuroprotective Agents; Pregnancy; Pregnancy Complications, Infectious; Rats; Rats, Sprague-Dawley; RNA, Messenger; Tumor Necrosis Factor-alpha

Escherichia coli gene fimA was the most frequent gene that occurred in the intestine of all investigated groups. All subjects with fimA gene had significantly higher values of tumor necrosis factor alpha (TNF-α) and CRP than those with other E. coli genes. There was also a tendency to increased serum interleukin (IL)-6 levels in patients carrying the fimA gene; however, no relation was observed to serum IL-8 and IL-10. Patients with Crohn's disease had significantly higher IL-6 than those with ulcerative colitis (UC) and controls. The highest levels of TNF-α were detected in the UC group. There were no significant differences in serum IL-8 and IL-10 between all three groups. The presence of E. coli gene fimA in the large bowel of patients with IBD is related to the immunological activity of the disease which may be important from the aspect of therapeutical strategy.

Topics: Adult; Aged; Anti-Bacterial Agents; Bacterial Typing Techniques; Case-Control Studies; Colitis, Ulcerative; Crohn Disease; Czech Republic; Escherichia coli; Escherichia coli Infections; Female; Fimbriae Proteins; Humans; Inflammation; Interleukin-10; Interleukin-6; Interleukin-8; Intestines; Male; Middle Aged; Polymerase Chain Reaction; Tumor Necrosis Factor-alpha

Effective response to mammary gland infection depends on efficient early innate immune response. The desired response would be one that is sufficient to clear the infection with a rapid return to the production of high-quality milk and limited tissue damage. In this study, 43 early lactation cows were ranked based on the ability of their fibroblasts to produce IL-8 in response to Escherichia coli lipopolysaccharide. Subsequently, the effect of a low or high response phenotype on the response to E. coli mastitis was determined. Untreated fibroblasts produced no detectable IL-8, whereas the range of IL-8 production in response to LPS (100 ng/mL) was approximately 7-fold between the lowest and highest responding cultures. Similar patterns of between-cow variation were observed in fibroblast production of IL-8 and IL-6 in response to IL-1β and Pam2CSK4 (a synthetic diacylated lipopeptide ligand). Four low and 4 high responder cows were challenged in late lactation with intramammary infusion of E. coli. All cows developed clinical mastitis in the challenged quarters and all cows cleared the infection within 8 d. However, somatic cell count began to decline earlier in the low responder group, and milk BSA concentration (an indicator of tissue damage) was also lower in low responders compared with high responders. Milk production from the challenged quarter was markedly depressed in both groups, but returned toward prechallenge values earlier in low responder cows. Dermal fibroblast cells appear predictive of a cow's response to mastitis. In this study, the low responder phenotype was sufficient to contain an E. coli infection with a more rapid return to the production of high quality milk.

Topics: Animals; Cattle; Cells, Cultured; Enzyme-Linked Immunosorbent Assay; Escherichia coli; Escherichia coli Infections; Female; Fibroblasts; Inflammation; Interleukin-6; Interleukin-8; Lipopeptides; Lipopolysaccharides; Mastitis, Bovine; Phenotype; Skin

Enteroaggregative Escherichia coli (EAEC) is emerging as a cause of acute and persistent diarrhea in developing countries. An important feature of EAEC pathogenesis is the induction of profound inflammatory response in the intestinal epithelium. In this article, we have shown that EAEC-induced activation of mitogen-activated protein kinases (MAPK) (ERK-1/2, JNK and p38MAPK) in cultured human intestinal epithelial cells (INT-407) leads to the induction of DNA-binding activity of NF-kappaB and AP-1, resulting in IL-8 production. Plasmid-cured EAEC could also activate the MAPK and the transcription factors leading to IL-8 secretion, but to a lesser extent than that of wild-type EAEC. Further, pretreatment of these cells with the highly specific MEK inhibitor (PD 098059), the JNK inhibitor (SP 600125), and the p38MAPK inhibitor (SB 203580) resulted in inhibition of the IL-8 secretion by EAEC (wild type as well as plasmid cured)-infected INT-407 cells. These findings demonstrate that the inflammatory response induced by EAEC may be due to the specific stimulation of MAPK signaling pathways leading to nuclear responses. To our knowledge, this is the first article regarding the detailed mechanism of IL-8 secretion from the EAEC-infected human intestinal epithelial cell line.

Topics: Cell Line; Embryo, Mammalian; Enzyme Activation; Escherichia coli; Escherichia coli Infections; Extracellular Signal-Regulated MAP Kinases; Humans; Interleukin-8; Intestinal Mucosa; Intestine, Small; MAP Kinase Signaling System; NF-kappa B; Protein Binding; Protein Kinase Inhibitors; RNA, Messenger; Transcription Factor AP-1; Transcription Factors; Up-Regulation

Escherichia coli are widespread in the environment and pathogenic strains cause diseases of mucosal surfaces including the female genital tract. Pelvic inflammatory disease (PID; metritis) or endometritis affects approximately 40% of cattle after parturition. We tested the expectation that multiple genetically diverse E. coli from the environment opportunistically contaminate the uterine lumen after parturition to establish PID.. Distinct clonal groups of E. coli were identified by Random Amplification of Polymorphic DNA (RAPD) and Multilocus sequence typing (MLST) from animals with uterine disease and these differed from known diarrhoeic or extra-intestinal pathogenic E. coli. The endometrial pathogenic E. coli (EnPEC) were more adherent and invasive for endometrial epithelial and stromal cells, compared with E. coli isolated from the uterus of clinically unaffected animals. The endometrial epithelial and stromal cells produced more prostaglandin E(2) and interleukin-8 in response to lipopolysaccharide (LPS) purified from EnPEC compared with non-pathogenic E. coli. The EnPEC or their LPS also caused PID when infused into the uterus of mice with accumulation of neutrophils and macrophages in the endometrium. Infusion of EnPEC was only associated with bacterial invasion of the endometrium and myometrium. Despite their ability to invade cultured cells, elicit host cell responses and establish PID, EnPEC lacked sixteen genes commonly associated with adhesion and invasion by enteric or extraintestinal pathogenic E. coli, though the ferric yersiniabactin uptake gene (fyuA) was present in PID-associated EnPEC. Endometrial epithelial or stromal cells from wild type but not Toll-like receptor 4 (TLR4) null mice secreted prostaglandin E(2) and chemokine (C-X-C motif) ligand 1 (CXCL1) in response to LPS from EnPEC, highlighting the key role of LPS in PID.. The implication arising from the discovery of EnPEC is that development of treatments or vaccines for PID should focus specifically on EnPEC and not other strains of E. coli.

Topics: Animals; Cattle; Cattle Diseases; Cells, Cultured; DNA, Bacterial; Endometritis; Endometrium; Escherichia coli; Escherichia coli Infections; Female; Genotype; Host-Pathogen Interactions; Interleukin-8; Lipopolysaccharides; Mice; Mice, Inbred C57BL; Mice, Knockout; Pelvic Inflammatory Disease; Phylogeny; Random Amplified Polymorphic DNA Technique; Species Specificity; Toll-Like Receptor 4; Uterus

Shiga toxins expressed in the intestinal lumen during infection with Shiga-toxigenic Escherichia coli must translocate across the epithelium and enter the systemic circulation to cause systemic (pathological) effects, including hemolytic uremic syndrome. The transepithelial migration of polymorphonuclear leukocytes in response to chemokine expression by intestinal epithelial cells is thought to promote uptake of Stx from the intestinal lumen by compromising the epithelial barrier. In the present study, we investigated the hypothesis that flagellin acts in conjunction with Shiga toxin to augment this chemokine expression. We investigated the relative contributions of nuclear factor kappaB (NF-kappaB) and mitogen-activated protein kinase (MAPK) signaling to transcription and translation of interleukin-8. Using reporter gene constructs, we showed that flagellin-mediated interleukin-8 gene transcription is heavily dependent on both NF-kappaB and extracellular signal-regulated kinase 1 and 2 (ERK-1/2) activation. In contrast, inhibition of p38 has no detectable effect on interleukin-8 gene transcription, even though flagellin-mediated activation of host p38 is critical for maximal interleukin-8 protein expression. Inhibition of MAPK-interacting kinase 1 suggests that p38 signaling affects the posttranscriptional regulation of interleukin-8 protein expression induced by flagellin. Cotreatment with Stx2 and flagellin results in a synergistic upregulation of c-Jun N-terminal protein kinases (JNKs), p38 activation, and a superinduction of interleukin-8 mRNA. This synergism was also evident at the protein level, with increased interleukin-8 protein detectable following cotreatment with flagellin and Stx2. We propose that flagellin, in conjunction with Shiga toxin, synergistically upregulates stress-activated protein kinases, resulting in superinduction of interleukin-8 and, ultimately, absorption of Stx into the systemic circulation.

Topics: Cell Line; Cloning, Molecular; Enzyme Activation; Enzyme-Linked Immunosorbent Assay; Escherichia coli Infections; Flagellin; Fluorescent Antibody Technique, Indirect; Humans; Interleukin-8; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; NF-kappa B; p38 Mitogen-Activated Protein Kinases; Reverse Transcriptase Polymerase Chain Reaction; Shiga Toxin 2; Shiga-Toxigenic Escherichia coli; Signal Transduction

Many bacterial pathogens utilize a type III secretion system to deliver multiple effector proteins into host cells. Here we found that the type III effectors, NleE from enteropathogenic E. coli (EPEC) and OspZ from Shigella, blocked translocation of the p65 subunit of the transcription factor, NF-kappaB, to the host cell nucleus. NF-kappaB inhibition by NleE was associated with decreased IL-8 expression in EPEC-infected intestinal epithelial cells. Ectopically expressed NleE also blocked nuclear translocation of p65 and c-Rel, but not p50 or STAT1/2. NleE homologues from other attaching and effacing pathogens as well OspZ from Shigella flexneri 6 and Shigella boydii, also inhibited NF-kappaB activation and p65 nuclear import; however, a truncated form of OspZ from S. flexneri 2a that carries a 36 amino acid deletion at the C-terminus had no inhibitory activity. We determined that the C-termini of NleE and full length OspZ were functionally interchangeable and identified a six amino acid motif, IDSY(M/I)K, that was important for both NleE- and OspZ-mediated inhibition of NF-kappaB activity. We also established that NleB, encoded directly upstream from NleE, suppressed NF-kappaB activation. Whereas NleE inhibited both TNFalpha and IL-1beta stimulated p65 nuclear translocation and IkappaB degradation, NleB inhibited the TNFalpha pathway only. Neither NleE nor NleB inhibited AP-1 activation, suggesting that the modulatory activity of the effectors was specific for NF-kappaB signaling. Overall our data show that EPEC and Shigella have evolved similar T3SS-dependent means to manipulate host inflammatory pathways by interfering with the activation of selected host transcriptional regulators.

Topics: Active Transport, Cell Nucleus; Caco-2 Cells; Dysentery, Bacillary; Escherichia coli Infections; Escherichia coli O157; Escherichia coli Proteins; HeLa Cells; Humans; I-kappa B Proteins; Interleukin-8; Intestinal Mucosa; NF-kappa B p50 Subunit; Proto-Oncogene Proteins c-rel; RNA, Messenger; Shigella boydii; Shigella flexneri; STAT1 Transcription Factor; STAT2 Transcription Factor; Transcription Factor RelA; Transcriptional Activation; Virulence; Virulence Factors

Infections with F4(+) enterotoxigenic Escherichia coli (ETEC) causes severe diarrhoea in piglets, resulting in morbidity and mortality. F4 fimbriae are the key virulence factors mediating the attachment of F4(+) ETEC to the intestinal epithelium. Intestinal epithelial cells (IEC) are recently being recognized as important regulators of the intestinal immune system through the secretion of cytokines, however, data on how F4(+) ETEC affect this cytokine secretion are scarce. By using ETEC strains expressing either polymeric, monomeric or F4 fimbriae with a reduced polymeric stability, we demonstrated that polymeric fimbriae are essential for adhesion to porcine IEC and the secretion of IL-6 and IL-8 by IEC. Remarkably, this cytokine secretion was not abrogated following stimulation with an F4-negative strain. Since this strain expresses flagellin, TLR5 mediated signalling could be involved. Indeed, porcine IEC express TLR5 and purified flagellin induced IL-6 and IL-8 secretion, indicating that, as for other pathogens, flagellin is the dominant virulence factor involved in the induction of proinflammatory responses in IEC. These results indicate a potential mucosal adjuvant capacity of ETEC-derived flagellin and may improve rational vaccine design against F4(+) ETEC infections.

Topics: Animals; Animals, Newborn; Antigens, Bacterial; Bacterial Adhesion; Cell Line; Diarrhea; Enterotoxigenic Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Fimbriae Proteins; Flagellin; Inflammation; Interleukin-6; Interleukin-8; Intestinal Mucosa; Mutant Proteins; Protein Engineering; Swine; Toll-Like Receptor 5; Virulence Factors

In this study, we investigated the interaction of 19 benign strains of lactic acid bacteria (LAB), bifidobacteria and staphylococci with enterohemorrhagic Escherichia coli (EHEC) strains of different serotypes and virulence gene spectrum in a HT29 cell culture infection model. As markers of infection, the secretion of interleukin 8 (IL-8) and the activation of the transcription factor NF-κB by the infected cells were determined. With 12 of 19 tested strains, a weak reduction <30% of IL-8 secretion of HT29 cells after co-infection with EHEC O157:H7 strain EDL933 was observed. Six strains reduced the IL-8 secretion up to 60% and the strain B. adolescentis DSMZ 20086 decreased the IL-8 production about 73%. In further co-infection assays with EHEC strains of the serotypes O103:H2, O26:H⁻, 0157:H⁻ and O113:H21, different abilities of the LAB strains to influence the infection with the different EHEC strains were noted. Therefore, the protective anti-inflammatory effect is strain specific for LAB and also depends on the application of EHEC strains with different sero- and virulence types. The differences in efficacy of protective bacteria against certain EHEC strains were unexpected and have not been shown so far. Furthermore, we could show that the inhibitory effects were not attributed to lower adhesion abilities of EHEC to the production of organic acids by the benign bacteria. In addition, viable bacteria are needed to inhibit the IL-8 secretion. Moreover, the NF-κB activation was reduced significantly by all tested LAB strains in co-infection trials, but was not strain-specific. The model described here is useful to screen for basic effects of protective bacteria that are able to counteract EHEC-mediated effects on human cells, and to study the molecular interaction between bacteria as well as between bacteria and human cultured cells.

Topics: Acids; Bacterial Adhesion; Bifidobacterium; Enterohemorrhagic Escherichia coli; Escherichia coli Infections; Gene Expression Regulation; HT29 Cells; Humans; Interleukin-8; Lactobacillus; NF-kappa B; Probiotics; Species Specificity; Staphylococcus

Although transmembrane C-type lectins (CLs) are known to initiate immune signaling, the participation and mechanism of action of soluble CLs have remained enigmatic. In this study, we found that M-ficolin, a conserved soluble CL of monocyte origin, overcomes its lack of membrane-anchor domain by docking constitutively onto a monocyte transmembrane receptor, G protein-coupled receptor 43 (GPCR43), to form a pathogen sensor-cum-signal transducer. On encountering microbial invaders, the M-ficolin-GPCR43 complex activates the NF-κB cascade to upregulate IL-8 production. We showed that mild acidosis at the local site of infection induces conformational changes in the M-ficolin molecule, which provokes a strong interaction between the C-reactive protein (CRP) and the M-ficolin-GPCR43 complex. The collaboration among CRP-M-ficolin-GPCR43 under acidosis curtails IL-8 production thus preventing immune overactivation. Therefore, we propose that a soluble CL may become membrane-associated through interaction with a transmembrane protein, whereupon infection collaborates with other plasma protein to transduce the infection signal and regulate host defense. Our finding implies a possible mechanism whereby the host might expand its repertoire of immune recognition-cum-regulation tactics by promiscuous protein networking. Furthermore, our identification of the pH-sensitive interfaces of M-ficolin-CRP provides a powerful template for future design of potential immunomodulators.

Topics: Acidosis; Animals; C-Reactive Protein; Cell Line; Chlorocebus aethiops; COS Cells; Escherichia coli Infections; Ficolins; Humans; Immunity, Innate; Interleukin-8; Lectins; Macromolecular Substances; Membrane Proteins; Monocytes; Pseudomonas Infections; Receptor Cross-Talk; Receptors, Cell Surface; Salmonella Infections; Signal Transduction; Staphylococcal Infections; U937 Cells; Up-Regulation

Infection with the enteric pathogen enterohemorrhagic Escherichia coli (EHEC) causes a variety of symptoms ranging from nonbloody diarrhea to more severe sequelae including hemorrhagic colitis, altered sensorium and seizures, and even life-threatening complications, such as hemolytic uremic syndrome and thrombotic thrombocytopenic purpura. The more severe consequences of EHEC infection are attributable to the production of Shiga toxin (Stx) and its subsequent effects on the vasculature, which expresses high levels of the Stx receptor, Gb3. Interestingly, the intestinal epithelium does not express Gb3. Despite the lack of Gb3 receptor expression, intestinal epithelial cells translocate Stx. The effect of Stx on intestinal epithelial cells is controversial with some studies demonstrating induction of inflammation and others not. This may be difficult to resolve because EHEC expresses both proinflammatory molecules, such as flagellin, and factor(s) that dampen the inflammatory response of epithelial cells. The goal of our study was to define the effect of Stx on the inflammatory response of intestinal epithelial cells and to determine whether infection by EHEC modulates this response. Here we show that Stx is a potent inducer of the inflammatory response in intestinal epithelial cells and confirm that EHEC attenuates the induction of IL-8 by host-derived proinflammatory cytokines. More importantly, however, we show that infection with EHEC attenuates the inflammatory response by intestinal epithelial cells to its own toxin. We speculate that the ability of EHEC to dampen epithelial cell inflammatory responses to Stx and cytokines facilitates intestinal colonization.

Topics: Cytokines; Enteritis; Enterohemorrhagic Escherichia coli; Epithelial Cells; Escherichia coli Infections; Host-Pathogen Interactions; HT29 Cells; Humans; I-kappa B Proteins; Inflammation Mediators; Interleukin-1beta; Interleukin-8; Intestinal Mucosa; NF-KappaB Inhibitor alpha; Protein Transport; Shiga Toxins; Trihexosylceramides; Tumor Necrosis Factor-alpha

The ability of some typical enteropathogenic Escherichia coli (EPEC) strains to adhere to, invade, and increase interleukin-8 (IL-8) production in intestinal epithelial cells in vitro has been demonstrated. However, few studies regarding these aspects have been performed with atypical EPEC (aEPEC) strains, which are emerging enteropathogens in Brazil. In this study, we evaluated a selected aEPEC strain (1711-4) of serotype O51:H40, the most prevalent aEPEC serotype in Brazil, in regard to its ability to adhere to and invade Caco-2 and T84 cells and to elicit IL-8 production in Caco-2 cells. The role of flagella in aEPEC 1711-4 adhesion, invasion, and IL-8 production was investigated by performing the same experiments with an isogenic aEPEC mutant unable to produce flagellin (FliC), the flagellum protein subunit. We demonstrated that this mutant (fliC mutant) had a marked decrease in the ability to adhere to T84 cells and invade both T84 and Caco-2 cells in gentamicin protection assays and by transmission electron microscopy. In addition, the aEPEC 1711-4 fliC mutant had a reduced ability to stimulate IL-8 production by Caco-2 cells in early (3-h) but not in late (24-h) infections. Our findings demonstrate that flagella of aEPEC 1711-4 are required for efficient adhesion, invasion, and early but not late IL-8 production in intestinal epithelial cells in vitro.

Topics: Bacterial Adhesion; Brazil; Cell Line; Colony Count, Microbial; Cytoplasm; DNA, Bacterial; Enterocytes; Enteropathogenic Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Flagella; Flagellin; Gene Deletion; Humans; Interleukin-8; Microscopy, Electron; Molecular Sequence Data; Sequence Analysis, DNA

Bovine colonic crypt cells express CD77 molecules that potentially act as receptors for Shiga toxins (Stx). The implication of this finding for the intestinal colonization of cattle by human pathogenic Stx-producing Escherichia coli (STEC) remains undefined. We used flow cytometric and real-time PCR analyses of primary cultures of colonic crypt cells to evaluate cell viability, CD77 expression, and gene transcription in the presence and absence of purified Stx1. A subset of cultured epithelial cells had Stx receptors which were located mainly intracellularly, with a perinuclear distribution, and were resistant to Stx1-induced apoptosis and Stx1 effects on chemokine expression patterns. In contrast, a population of vimentin-positive cells, i.e., mesenchymal/nonepithelial cells that had high numbers of Stx receptors on their surface, was depleted from the cultures by Stx1. In situ, CD77(+) cells were located in the lamina propria of the bovine colon by using immunofluorescence staining. A newly established vimentin-positive crypt cell line with high CD77 expression resisted the cytolethal effect of Stx1 but responded to Stx1 with a significant increase in interleukin-8 (IL-8), GRO-alpha, MCP-1, and RANTES mRNA. Combined stimulation with lipopolysaccharide and Stx1 increased IL-10 mRNA. Our results show that bovine colonic crypt cells of epithelial origin are resistant to both the cytotoxic and modulatory effects of Stx1. In contrast, some mucosal mesenchymal cells, preliminarily characterized as mucosal macrophages, are Stx1-responsive cells that may participate in the interaction of STEC with the bovine intestinal mucosa.

Topics: Animals; Cattle; Cell Survival; Cells, Cultured; Chemokine CCL5; Chemokine CXCL1; Colon; Epithelial Cells; Escherichia coli Infections; Flow Cytometry; Fluorescent Antibody Technique; Gene Expression; Interleukin-8; Intestinal Mucosa; Mesoderm; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Shiga Toxin 1; Shiga-Toxigenic Escherichia coli; Transcription, Genetic; Transforming Growth Factor beta; Trihexosylceramides

In this study we tested how a combination of early and late paraclinic markers could predict early onset neonatal sepsis (EONS).. The first 24 hours after the suspicion of EONS, we measured interleukine (IL)-6, IL-8, IL-10, IL-18, tumor necrosis factor-alpha (TNF-alpha), interferon gamma (INF-gamma), procalcitonin (PCT) and C-reactive protein (CRP) at 8-hour intervals on 123 neonates clinically suspected for EONS. The neonates were divided into two groups. The sepsis group: 1A with blood culture verified bacteraemia and 1B strongly suspected sepsis (29 patients). The no sepsis group: 2A treated with antibiotics (37 patients) and 2B not treated with antibiotics (57 patients).. Combined evaluation of each of the early markers with PCT > 25 ng/ml for prediction of EONS at time 0, gave the following sensitivities and specificities: IL-6 > 250 pg/ml: 71% and 88%; IL-8 > 900 pg/ml: 50% and 88%; IL-10 > 40 pg/ml: 43% and 87%; and immature/total (I/T) ratio > 0.35: 59% and 88%. The results of IL-18, TNF-alpha and IFN-gamma did not predict EONS.. IL-6 combined with PCT values is a fair way to evaluate EONS at the time of suspicion of infection. The "old" early marker, I/T ratio, is almost as efficient as IL-6. By combining an early and a late marker it may be possible to reduce the diagnostic "non-conclusive" period of paraclinic values.

Topics: Anti-Bacterial Agents; Bacteremia; Biomarkers; C-Reactive Protein; Calcitonin; Calcitonin Gene-Related Peptide; Cytokines; Escherichia coli Infections; Female; Humans; Infant, Newborn; Inflammation Mediators; Interferon-gamma; Interleukin-10; Interleukin-18; Interleukin-6; Interleukin-8; Leukocyte Count; Male; Neutrophils; Protein Precursors; Retrospective Studies; Sensitivity and Specificity; Sepsis; Staphylococcal Infections; Streptococcal Infections; Streptococcus agalactiae; Tumor Necrosis Factor-alpha

Staphylococcus aureus, but not E. coli pathogens frequently cause subclinical, chronic infections of the mammary gland. We examined here, if inadequate activation of the bovine TLR2 and TLR4 pathogen receptors by ligands derived from S. aureus pathogens might contribute to molecular mechanisms underpinning the escape strategies from mammary immune defence of this pathogen. We show that infections with live E. coli, but not S. aureus pathogens induce strongly IL-8 and TNFalpha gene expression in the udders. Yet, preparations of heat-killed bacteria from both pathogens activate equally well bovine TLR2 and TLR4 receptors to induce NF-kappaB activation, as shown in the HEK293 reconstitution system of TLR-signal transduction. LTA prepared from the S. aureus strain used to infect the cows activates the bovine TLR2 as strongly as the entire, heat-killed pathogen. Both pathogens induce in primary bovine mammary epithelial cells (pbMEC) IL-8 and TNFalpha gene expression, but S. aureus to less than 5% of the degree caused by E. coli. This impaired proinflammatory activation is paralleled by a complete lack of NF-kappaB activation in pbMEC by S. aureus or LTA. In contrast, E. coli and LPS activate strongly NF-kappaB in these cells. A large proportion of this activation is attributable to TLR-mediated signalling, since a dual transdominant negative DN-MyD88-DN-TRIF factor blocks >80% of the pathogen-related NF-kappaB activation in pbMEC. Our results prove that impaired binding of TLR-ligands from the pathogenic S. aureus strain are not the cause for the inadequate mammary immune response elicited by this pathogen. Rather, the pathogen causing subclinical mastitis impairs NF-kappaB activation in MEC thereby severely weakening the immune response in the udder.

Topics: Animals; Animals, Domestic; Cattle; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Immunity; Interleukin-8; Mammary Glands, Animal; NF-kappa B; Signal Transduction; Staphylococcal Infections; Staphylococcus aureus; Toll-Like Receptor 2; Toll-Like Receptor 4; Tumor Necrosis Factor-alpha

Based on the controversial findings of clinical studies regarding the influence of multidrug resistance on mortality, 10 susceptible and 10 multidrug-resistant (MDR) and extended-spectrum beta-lactamase-producing isolates of Escherichia coli were applied to stimulate monocytes isolated from healthy donors. Immune mediators were estimated in supernatants. Four susceptible isolates (Group A) and four MDR isolates (Group B) were used to initiate acute pyelonephritis in 48 rabbits following inoculation of the pathogen into the right renal pelvis. Survival was recorded and blood monocytes were isolated and incubated to estimate the ex vivo release of tumour necrosis factor-alpha (TNFalpha). Release of TNFalpha, interleukin (IL)-6 and IL-8 was higher after 2 h and 4 h of stimulation by MDR isolates compared with susceptible isolates. The opposite occurred for the release of IL-12. Death occurred in 22 rabbits in Group A (91.7%) compared with 12 in Group B (50.0%) (P=0.003). Monocytes isolated at 24 h from Group A rabbits released significantly higher TNFalpha than monocytes from Group B. Tissue bacterial load after animal death was significantly higher in the kidneys of Group A rabbits. It is concluded that susceptible and MDR E. coli stimulate monocytes resulting in a different pattern of release of pro-inflammatory cytokines, which is accompanied by prolonged survival following experimental sepsis by MDR isolates.

Topics: Animals; beta-Lactamases; Cells, Cultured; Colony Count, Microbial; Drug Resistance, Multiple, Bacterial; Escherichia coli; Escherichia coli Infections; Female; Humans; Interleukin-12; Interleukin-6; Interleukin-8; Kidney; Male; Monocytes; Pyelonephritis; Rabbits; Survival Analysis; Tumor Necrosis Factor-alpha

Mastitis is one of the most prevalent diseases in cattle and remains among the most costly diseases to the dairy industry. Various surveys have indicated a greater prevalence of and risk for mastitis in Holstein cows than in Jersey cows. The innate immune system comprises the immediate host defense mechanisms that respond to infection, and differences in the magnitude and rapidity of this response are known to influence susceptibility to and clearance of infectious pathogens. The reported differences in the prevalence of mastitis between Holstein and Jersey cows may suggest the occurrence of breed-dependent differences in the innate immune response to intramammary infection. The objective of the current study was to compare the acute phase and cytokine responses of Holstein and Jersey cows following intramammary infection by the bacterial pathogen Escherichia coli, a leading cause of clinical mastitis. All cows in the study were in similar stages of lactation, of the same parity, subjected to the same housing and management conditions, and experimentally infected on the same day with the same inoculum preparation. Before and after infection, the following innate immune parameters were monitored: bacterial clearance; febrile response; induction of the acute phase proteins serum amyloid A and lipopolysaccharide-binding protein; alterations in total and differential white blood cell counts; changes in milk somatic cell counts and mammary vascular permeability; and induction of the cytokines IFN-gamma, IL-1beta, IL-8, IL-12, and tumor necrosis factor-alpha. Overall innate immune responses were similar between the 2 breeds; however, temporal differences in the onset, cessation, and duration of several responses were detected. Despite these differences, intramammary clearance of E. coli was comparable between the breeds. Together, these data demonstrate a highly conserved innate immune response of Holstein and Jersey cows to E. coli intramammary infection.

Topics: Acute-Phase Proteins; Acute-Phase Reaction; Animals; Breeding; Carrier Proteins; Cattle; Cytokines; Disease Susceptibility; Escherichia coli; Escherichia coli Infections; Female; Immunity, Innate; Interferon-gamma; Interleukin-1; Interleukin-12; Interleukin-8; Mammary Glands, Animal; Mastitis, Bovine; Membrane Glycoproteins; Prevalence; Risk Factors; Time Factors

The defense against mucosal infections relies on chemokines that recruit inflammatory cells to the mucosa. This study examined if the chemokine response to uro-pathogenic Escherichia coli is influenced by fimbrial expression. The CXC (CXCL1, CXCL5, CXCL8, CXCL9, CXCL10) and CC chemokines (CCL2, CCL3, CCL5) were quantified after in vitro infection of uro-epithelial cells with a fimbriated E. coli pyelonephritis isolate, or with P or type 1 fimbriated transformants of an avirulent E. coli K-12 strain. The response profile was shown to vary with the fimbrial type. Type 1 fimbriated E. coli elicited mainly CXCL1 and CXCL8, whereas P fimbriated E. coli stimulated CCL2 and CCL5 and class II were more potent chemokine inducers than class III P fimbriae. Chemokines were also quantified in urine samples from 73 patients with febrile urinary tract infection, and analyzed as a function of disease severity and fimbrial expression by the strain infecting each patient. A complex CXC and CC chemokine response was detected in patient urine, with a significant influence of the fimbrial type. The results show that virulence factors like fimbriae may modify the mucosal chemokine response. This mechanism may allow the host to adjust the inflammatory cell infiltrate to fit the infecting strain.

Topics: Adhesins, Escherichia coli; Adult; Aged; Aged, 80 and over; Cell Line; Chemokines; Escherichia coli; Escherichia coli Infections; Female; Fever; Fimbriae Proteins; Fimbriae, Bacterial; Genotype; Humans; Interleukin-8; Kinetics; Lectins; Male; Middle Aged; Mucous Membrane; RNA, Messenger; Urinary Tract Infections; Urothelium

Escherichia coli are the most common etiological agents of urinary tract infections (UTIs). Uropathogenic E. coli (UPECs) produce specific toxins including the cytotoxic necrotizing factor-1 (CNF1) and the alpha-hemolysin (alpha-Hly). CNF1 triggers, through Rho protein activation, a specific gene response of host cells, which results in the production for instance of interleukin-8 (IL-8), monocyte chemoattractant protein-1 (MCP-1) and the macrophage inflammatory protein-3alpha (MIP-3alpha). The alpha hemolysin alpha-Hly also triggers the production of inflammatory mediators. Cnf1 is always associated with alpha-hly in a pathogenicity island conserved among UPECs. Using two complementary approaches we have investigated whether alpha-hly and cnf1 bearing UPECs are associated with a specific type of UTI both in term of pathology and host response. Here we report that UPECs bearing alpha-hly/cnf1 have a prevalence of 50% in UPECs isolated from hemorrhagic UTIs, as compared to 30% in the overall UPEC population. In addition, we observed that MCP-1, and IL-8 to a lower extent, is produced in urine at higher concentrations in UTIs caused by UPECs carrying alpha-hly/cnf1.

Topics: Bacterial Toxins; Chemokine CCL2; Chemokine CCL20; Chemokines, CC; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Health; Hemolysin Proteins; Hemorrhage; Humans; Interleukin-8; Macrophage Inflammatory Proteins

The role of diffusely adherent Escherichia coli (DAEC) in diarrheal disease has been controversial. However, DAEC strains were recently implicated in diarrheal disease in developing countries. To clarify whether DAEC are prevalent among sporadic cases of diarrheal illness in Osaka City, Japan, E. coli strains isolated between July 1997 and March 2000 during diarrheagenic E. coli (DEC) investigation were retrospectively examined. DAEC strains were recognized among 41 (4.4%) of 924 patients and formed the biggest subgroup of DEC. Previously, we reported that some DAEC strains caused epithelial cells to secrete as much IL-8 as enteroaggregative E. coli strains did. In this study, we attempted to evaluate epidemiologically whether the ability of DAEC to induce IL-8 was involved in the pathogenesis. Relationship among patient age, symptoms, Afa adhesins, season and IL-8 induction were examined. The subgroup of DAEC that possessed Afa genes and/or induced a high level of IL-8 was significantly prevalent among patients age 1 to 4 years; however total DAEC was not significantly high among the children compared to other age group. IL-8 inducing DAEC seems to play a role in causing sporadic diarrheal illnesses, particularly in pediatric fields. Investigations highlighting the relationship between IL-8 induction and enteropathogenicity are clearly necessary to confirm the role of DAEC in infectious enteritis.

Topics: Adhesins, Escherichia coli; Adolescent; Adult; Age Distribution; Bacterial Adhesion; Child; Child, Preschool; Diarrhea; Escherichia coli; Escherichia coli Infections; Feces; Female; Humans; Infant; Interleukin-8; Intestines; Japan; Male; Prevalence; Retrospective Studies; Seasons

Uropathogenic Escherichia coli (UPEC), the most frequent cause of urinary tract infection (UTI), is associated with an inflammatory response which includes the induction of cytokine/chemokine secretion by urothelial cells and neutrophil recruitment to the bladder. Recent studies indicate, however, that UPEC can evade the early activation of urothelial innate immune response in vitro. In this study, we report that infection with the prototypic UPEC strain NU14 suppresses tumor necrosis factor alpha (TNF-alpha)-mediated interleukin-8 (CXCL-8) and interleukin-6 (CXCL-6) secretion from urothelial cell cultures compared to infection with a type 1 piliated E. coli K-12 strain. Furthermore, examination of a panel of clinical E. coli isolates revealed that 15 of 17 strains also possessed the ability to suppress cytokine secretion. In a murine model of UTI, NU14 infection resulted in diminished levels of mRNAs encoding keratinocyte-derived chemokine, macrophage inflammatory peptide 2, and CXCL-6 in the bladder relative to infection with an E. coli K-12 strain. Furthermore, reduced stimulation of inflammatory chemokine production during NU14 infection correlated with decreased levels of bladder and urine myeloperoxidase and increased bacterial colonization. These data indicate that a broad phylogenetic range of clinical E. coli isolates, including UPEC, may evade the activation of innate immune response in the urinary tract, thereby providing a pathogenic advantage.

Topics: Animals; Chemokine CXCL2; Chemokines; Colony Count, Microbial; Escherichia coli; Escherichia coli Infections; Female; Immunity, Innate; Interleukin-6; Interleukin-8; Mice; Mice, Inbred C57BL; Peroxidase; RNA, Messenger; Tumor Necrosis Factor-alpha; Urinary Bladder; Urinary Tract Infections; Urine; Urothelium; Virulence

The technetium 99 m (99mTc)-radiolabeled, leukocyte-avid peptide-glycoseaminoglycan complex, [99mTc]P1827DS, has been synthesized as an improved infection/inflammation imaging agent to [99mTc]P483H (LeukoTect, Diatide). In a phase I/II clinical trail, [99mTc]P483H images were equivalent to those obtained with 111In ex vivo labeled leukocytes. However, there was physiologic accumulation of radioactivity in the body that could hamper interpretation of the images. In this study, the potential of [99mTc]P1827DS for infection imaging was assessed in comparison with [99mTc]P483H and the well-described imaging agent [99mTc] hydrazinonicotinamide (HYNIC)-interleukin 8 (IL-8).. The binding of [99mTc]P1827DS to human blood cell was studied in vitro. A rabbit Escherichia coli infection model was used to perform the biodistribution and imaging studies with [99mTc]P1827DS, [99mTc]P483H and [99mTc]HYNIC-IL-8.. [99mTc]P1827DS binds to leukocytes but not to erythrocytes. The leukocyte binding was not saturable up to an investigated concentration of 10 microM. The accumulation of [99mTc]P1827/DS at the infection site strongly depends on the P1827/DS ratio and was optimal at a molar ratio of 10:1. [99mTc]P1827DS shows improved biodistribution over [99mTc]P483H with similar uptake at the infection site. Abscess uptake of [99mTc]HYNIC-IL-8 was approximately three times higher than that of [99mTc]P1827DS. [99mTc]HYNIC-IL-8 showed high accumulation in the kidneys, whereas [99mTc]P1827DS showed high lung uptake and slightly higher accumulation in the liver and spleen.. [99mTc]P1827DS is a potential new inflammation imaging agent, which clearly visualized the abscess in the rabbit E. coli infection model and showed improved biodistribution compared to [99mTc]P483H. However, the infection uptake and biodistribution of [99mTc]P1827DS is not superior to that of [99mTc]HYNIC-IL-8 in this animal model.

Topics: Animals; Drug Evaluation, Preclinical; Escherichia coli Infections; Glycosaminoglycans; Interleukin-8; Metabolic Clearance Rate; Organ Specificity; Organotechnetium Compounds; Rabbits; Radionuclide Imaging; Radiopharmaceuticals; Tissue Distribution

Chorioamniotic membranes infection is a pathologic condition in which an abnormal secretion of proinflammatory cytokines halts fetal immune tolerance. The aim of the present study was to evaluate the functional response of human chorioamniotic membranes, as well as the individual contribution of the amnion and choriodecidua after stimulation with Escherichia coli, a pathogen associated with preterm labor.. Explants of chorioamniotic membranes from 10 women (37-40 weeks of gestation) were mounted and cultured in a Transwell system, which allowed us to test the amnion and choriodecidua compartments independently. Escherichia coli (1 x 10 6 CFU/mL) was added to either the amniotic or the choriodecidual regions or both; after a 24-h incubation, the secretion of IL-1beta, IL-6, TNFalpha, IL-8, and IL-10 in both compartments was measured using a specific ELISA. Data were analyzed by Kruskal-Wallis one-way analysis of variance.. After stimulation with Escherichia coli, the choriodecidua compartment showed an increase in the secretion of IL-1beta (21-fold), IL-6 (2-fold), IL-8 (6-fold), and IL-10 (37-fold), regardless of which side of the membrane was stimulated; TNFalpha secretion augmented (22-fold) also but only when the stimulus was applied simultaneously to both sides. When the amnion was stimulated directly, the level of IL-1beta (13-fold) rose significantly; however, the increase in IL-8 secretion was larger (20-fold), regardless of the primary site of infection. TNFalpha secretion in the amnion compartment rose markedly only when Escherichia coli was simultaneously applied to both sides.. Selective stimulation of fetal membranes with Escherichia coli results in a differential production of IL-1beta, IL-6, TNFalpha, IL-8, and IL-10. These tissues were less responsive when the amnion side was stimulated. This is in agreement with the hypothesis that the choriodecidua may play a primary role during an ascending intrauterine infection, being the main barrier to progression of the infection into the amniotic cavity. Therefore, the tissue-specific capacities for the secretion of these immune modulators could be a determining factor for the degree of severity of the inflammation process in fetal membranes.

Topics: Adult; Escherichia coli; Escherichia coli Infections; Extraembryonic Membranes; Female; Humans; Interleukin-10; Interleukin-1beta; Interleukin-6; Interleukin-8; Models, Biological; Pregnancy; Tissue Culture Techniques; Tumor Necrosis Factor-alpha

Acute endotoxemia is characterized by an enhanced inflammatory response. Pentoxifylline (PTX), a phosphodiesterase inhibitor, has been shown to decrease TNF-alpha levels and to down-regulate neutrophil activation, likely because of increases in intracellular cyclic AMP. Its effects on lipopolysaccharide (LPS) induced lung injury, more specifically on tissue neutrophil infiltration and degranulation, adhesion molecule expression, and transcriptional factor activation, have not been fully investigated. We postulated that PTX treatment in acute endotoxemia downregulates the inflammatory response and may decrease lung injury.. Male Sprague-Dawley rats were randomized into three groups: Sham (saline i.v.), LPS (5 mg/kg i.v.), and PTX + LPS (25 mg/kg and 5 mg/kg i.v., respectively; concomitant injection). After 4 hours, bronchoalveolar lavage fluid (BAL), plasma, and lungs were sampled. BAL IL-8 (ELISA), BAL MMP-2, plasma MMP-9, and BAL MMP-9 (Zymography) were measured. Lung histology (H&E), in addition to lung MPO, ICAM-1, and NF-kappaB expression evaluated by immunohistochemistry were analyzed. Lung NF-kappaB DNA binding was evaluated by electrophoretic mobility shift assay.. PTX treatment decreased BAL IL-8 levels, BAL MMP-2, and plasma MMP-9 activity. Lung neutrophil infiltration (MPO), ICAM-1 expression and NF-kappaB activation were decreased by PTX. In addition, PTX treatment caused a marked attenuation of LPS-induced lung injury.. Phosphodiesterase inhibition by PTX attenuates LPS-induced end-organ injury. In addition, proinflammatory cytokine production is also downregulated, likely because of the marked attenuation of NF-kappaB DNA binding and activation.

Topics: Animals; Disease Models, Animal; Endotoxemia; Escherichia coli Infections; Intercellular Adhesion Molecule-1; Interleukin-8; Lipopolysaccharides; Lung; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; NF-kappa B; Pentoxifylline; Peroxidase; Phosphodiesterase Inhibitors; Rats; Rats, Sprague-Dawley; Respiratory Distress Syndrome

Probiotic bacteria may provide protection against intestinal damage induced by pathogens, but the underlying mechanisms are still largely unknown. We investigated whether Bifidobacterium animalis MB5 and Lactobacillus rhamnosus GG (LGG) protected intestinal Caco-2 cells from the inflammation-associated response induced by enterotoxigenic Escherichia coli (ETEC) K88, by inhibiting pathogen attachment to the cells, which is the first step of ETEC pathogenicity, and regulating neutrophil recruitment, a crucial component of inflammation. A partial reduction of ETEC adhesion was exerted by probiotics and their culture supernatant fractions either undigested or digested with proteases. ETEC viability was unaffected by the presence of B. animalis, LGG or their supernatant fractions in the culture medium, indicating an absence of probiotic bactericidal activity. Probiotics and their supernatant fractions, either undigested or digested with proteases, strongly inhibited the neutrophil transmigration caused by ETEC. Both B. animalis and LGG counteracted the pathogen-induced up regulation of IL-8, growth-related oncogene-alpha and epithelial neutrophil-activating peptide-78 gene expression, which are chemokines essential for neutrophil migration. Moreover, the probiotics prevented the ETEC-induced increased expression of IL-1beta and TNF-alpha and decrease of transforming growth factor-alpha, which are regulators of chemokine expression. These results indicate that B. animalis MB5 and LGG protect intestinal cells from the inflammation-associated response caused by ETEC K88 by partly reducing pathogen adhesion and by counteracting neutrophil migration, probably through the regulation of chemokine and cytokine expression.

Topics: Bacterial Adhesion; Bifidobacterium; Caco-2 Cells; Cells, Cultured; Chemokine CXCL1; Chemokines, CXC; Escherichia coli; Escherichia coli Infections; Gene Expression Regulation; Humans; Intercellular Signaling Peptides and Proteins; Interleukin-1; Interleukin-8; Intestines; Lacticaseibacillus rhamnosus; Neutrophil Infiltration; Neutrophils; Probiotics; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transforming Growth Factor alpha; Tumor Necrosis Factor-alpha

Topics: Adult; Colitis, Ulcerative; Escherichia coli Infections; Female; Humans; Interleukin-8; Pyoderma Gangrenosum

Numerous molecular entities with diverse structures have been radiolabeled and investigated as potential infection and inflammation detection agents. However, none of these molecules have gained the acceptance of gallium citrate or radiolabeled autologous white blood cells. We have radioiodinated interleukin-8 using two different methods and tested the biological behavior of the products in mice. As expected, the direct radioiodinated material displayed extensive in vivo deiodination. The use of pyridine-based prosthetic label yielded a product with better kinetics than the direct radioiodination method and showed a better target to non-target ratio. Nonetheless, this method is not suited for labeling of bioactive peptides such as the title peptide because of the very high specific activity required to prevent cytotoxic effects in a human application.

Topics: Animals; Cells, Cultured; Escherichia coli Infections; Interleukin-8; Iodine Radioisotopes; Isotope Labeling; Metabolic Clearance Rate; Mice; Mice, Inbred CBA; Neutrophils; Organ Specificity; Radionuclide Imaging; Radiopharmaceuticals; Tissue Distribution

Nitric oxide (NO) is an antibacterial factor that is produced by the enzyme inducible NO synthase (iNOS). Uroepithelial cells express iNOS in experimental models of urinary tract infection but the stimulatory and regulatory mechanisms are still unclear. We investigated the influence of uropathogenic Escherichia coli strains with different fimbrial expression and the effect of proinflammatory cytokines on the host iNOS response.. A498 human kidney epithelial cells were stimulated with different uropathogenic E. coli strains, namely the P and type 1-fimbriated clinical isolate AD110, the recombinant P-fimbriated strain E. coli HB101(pPIL110-75) and the recombinant type 1-fimbriated strain E. coli AAEC191A(pPKL4). NO production was determined as nitrite production in cell culture medium. Studies of nuclear factor-kappaB (NF-kappaB) binding to the iNOS promoter and reverse transcriptase-polymerase chain reaction of iNOS mRNA were performed to investigate iNOS gene activation in response to uropathogenic E. coli. The effect of interleukin (IL)-6, IL-8 and transforming growth factor-beta on NO production was also examined.. E. coli per se failed to induce NO production and iNOS mRNA in A498 cells. However, in combination with interferon-gamma AD110 and the type 1-fimbriated strain caused a small increase in NO production and iNOS mRNA. AD110 stimulated A498 cells demonstrated weak binding of NF-kappaB to a human iNOS promoter sequence. IL-6, IL-8 and transforming growth factor-a did not affect NO production in A498 cells.. Uropathogenic bacteria are weak inducers of human uroepithelial iNOS, which may be related to insufficient binding of NF-kappaB to iNOS promoter. The uroepithelial iNOS response did not appear to be regulated by proinflammatory cytokines.

Topics: Cells, Cultured; Epithelial Cells; Escherichia coli Infections; Humans; Interleukin-6; Interleukin-8; Kidney; Nitric Oxide; Nitric Oxide Synthase; Transforming Growth Factor beta; Urinary Tract Infections; Urothelium

Septic acute lung injury (ALI) causes high morbidity and mortality in intensive care service as a result of biotrauma and dysfunction in the lungs and other organ systems. We hypothesized that surfactant and/or inhaled nitric oxide (iNO) may have different effects in modulation of inflammatory injury in septic ALI. Twenty-four healthy, 6-9 kg piglets were anesthetized, and intraperitoneally injected with Escherichia coli, followed by a low tidal volume ventilation until sepsis and ALI developed within 4-6 h. They were then randomly treated in groups (n=6 each) as: control (C), inhaled NO at 10 ppm (NO), surfactant at 100mg/kg (Surf), or both surfactant and iNO (SNO). A normal control group (N) was sham-injected and similarly ventilated. Over the 24 h of treatment period, both Surf, and SNO groups had significantly improved PaO2/FiO2, dynamic compliance and resistance of respiratory system. At 24h, the best alveolar aeration and least protein leakage, the lowest wet-to-dry lung weight ratio and lung injury score were found in SNO. Activity of nuclear factor kappa B (NF-kappaB) and myeloperoxidase, interleukin 8 mRNA expression and melondialdehyde were significantly increased, and IL-10 mRNA decreased, in lung tissue of the C group, but were significantly altered in the SNO group, and moderately altered in either NO or Surf group. We conclude that the effects of lung protection by surfactant and/or iNO in this model may be different in modulation of inflammatory cytokine mRNA expression and activity of NF-kappaB, and iNO did not have adverse effects.

Topics: Acute Disease; Administration, Inhalation; Analysis of Variance; Animals; Bronchoalveolar Lavage Fluid; Electrophoretic Mobility Shift Assay; Escherichia coli Infections; Fibroblast Growth Factor 7; Interleukin-10; Interleukin-8; Lung; Male; Malondialdehyde; Methemoglobin; NF-kappa B; Nitrates; Nitric Oxide; Nitrites; Oligonucleotide Probes; Peroxidase; Pneumonia; Protein Binding; Pulmonary Surfactants; Pulmonary Ventilation; Respiratory Insufficiency; RNA, Messenger; Swine

Infection of the bovine mammary gland, in addition to causing animal distress, is a major economic burden of the dairy industry. Greater understanding of the initial host response to infection may lead to more accurate selection of resistant animals or to novel prophylactic or therapeutic intervention strategies. The epithelial cell plays a role in the host response by alerting the immune system to the infection and providing a signal as to where the infection is located. To understand this process better, a cDNA microarray approach was used to search for potential signals produced by mammary epithelial cells in response to exposure to Escherichia coli lipopolysaccharide (LPS). Total RNA from separate cultures of epithelial cells from 4 Holstein cows was harvested 6 h after LPS challenge or control conditions. For each cow, RNA from control or LPS-exposed cells was transcribed to cDNA and labeled with Cy3 or Cy5, then pooled and applied to a bovine total leukocyte (BOTL) microarray slide containing 1278 unique transcripts. Dye reversal was used so that RNA from two of the control cultures was labeled with Cy3 while RNA from the other two control cultures was labeled with Cy5. From the resulting microarray data we selected 4 of the 9 genes significantly (P < 0.02) induced (>1.25-fold) in response to LPS exposure for more detailed analysis. The array signal intensity for 3 of these genes, RANTES/CCL5, IL-6 and T-PA, was relatively low, but quantitative real-time RT-PCR (Q-RT-PCR) analysis revealed that they were induced 208-fold, 10-fold and 3-fold, respectively. The gene that showed the greatest fold induction by microarray analysis (2.5-fold) was CXCL5. This gene had a relatively strong signal intensity on the array and was easily detected by northern blot analysis, which indicated a 10-fold induction. This cell culture model system provides evidence for an important role of the mammary epithelial cell in initiating the innate response to infection.

Topics: Animals; Cattle; Cell Culture Techniques; Chemokine CXCL5; Chemokines, CXC; Epithelial Cells; Escherichia coli Infections; Female; Gene Expression Profiling; Gene Expression Regulation; Interleukin-8; Leukocytes; Lipopolysaccharides; Mammary Glands, Animal; Mastitis, Bovine; Oligonucleotide Array Sequence Analysis; Reverse Transcriptase Polymerase Chain Reaction; Staphylococcal Infections

Escherichia coli is associated with inflammation in the brain. To investigate whether astrocytes are involved in E. coil-induced inflammation, we assessed the levels of expression of proinflammatory mediators produced by E. coli-infected astrocytes. E. coli infection in primary human astrocytes and cell lines increased expression of the CXC chemokine IL-8/GRO-alpha, the CC chemokine MCP-1, TNF-alpha, and iNOS. E. coli infection activated p65/p50 heterodimeric NF-kappaB and concurrently decreased the signals of IkappaBalpha. Blocking the NF-kappaB signals by IkappaBalpha-superrepressor-containing retrovirus or antisense p50 oligonucleotide transfection resulted in down-regulation of expression of the proinflammatory mediators. Furthermore, superrepressors of IkappaBalpha, IkappaB kinase (IKK) or NF-kappaB inducing kinase (NIK) inhibited the up-regulated expression of the downstream target genes of NF-kappaB such as IL-8 and MCP-1, and superrepressors of TNF receptor-associated factor (TRAF)2 and TRAF5 also inhibited expression of the E. coli-induced target genes of NF-kappaB. These results indicate that proinflammatory mediators such as the CXC chemokine IL-8/GRO-alpha, the CC chemokine MCP-1, TNF-alpha, and iNOS can be expressed in E. coli-infected astrocytes via an NF-kappaB pathway, suggesting that these mediators may contribute to inflammation in the brain, including infiltration of inflammatory cells.

Topics: Astrocytes; Cell Line, Tumor; Chemokine CCL2; Chemokine CXCL1; Chemokines; Chemokines, CXC; Escherichia coli Infections; Humans; I-kappa B Kinase; Intercellular Signaling Peptides and Proteins; Interleukin-8; NF-kappa B; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Phosphorylation; Protein Serine-Threonine Kinases; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; TNF Receptor-Associated Factor 2; TNF Receptor-Associated Factor 5; Tumor Necrosis Factor-alpha

Acute Escherichia coli mastitis is one of the major sources of economic loss in the dairy industry due to reduced milk production, treatment costs, discarded milk, and occasional fatal disease. Nonsteroidal anti-inflammatory drugs (NSAIDs) are frequently used as adjunctive therapy to antibiotics. The objective of the current study was to evaluate the effect of carprofen treatment following infusion of Escherichia coli into the mammary glands of primiparous cows during the periparturient period. Severity of mastitis was scored based on the average milk production in the uninfected quarters on d +2 postinoculation and a clinical severity score. Carprofen was administered intravenously at 9 h postchallenge, when clinical signs of mastitis appeared. In previous work, efficacy of NSAIDs was mainly evaluated using clinical symptoms. In the present study, the effect of carprofen on innate immune response was also assessed by quantification of inflammatory mediators. All primiparous cows reacted as moderate responders throughout the experimental period. Primiparous cows were intramammarily inoculated with 1 x 10(4) cfu of E. coli P4:O32 in 2 left quarters. Analysis of blood and milk parameters, including IL-8, complement component C5a, lipopolysaccharide-binding protein (LBP), soluble CD14, prostaglandin E2, and thromboxane B2 was performed from d 0 to d +6 relative to intramammary inoculation. Rectal temperature in carprofen-treated animals was lower than in control animals at 3 and 6 h posttreatment. Treatment also restored the decreased reticulorumen motility that occurs during E. coli mastitis to preinfection levels faster than in control animals. Carprofen treatment resulted in an earlier normalization of the clinical severity score. Eicosanoid (prostaglandin E2 and thromboxane B2) production in milk tended to be inhibited by carprofen. No significant differences in the kinetic patterns of somatic cell count, IL-8, complement component C5a, LBP, and soluble CD14 were observed. In conclusion, carprofen treatment improved general clinical condition by effective antipyrexia and restoration of reticulorumen motility but did not significantly inhibit eicosanoid production. Carprofen treatment did not result in a significant decrease of chemotactic inflammatory mediators, IL-8 and C5a, and early innate immune molecules, sCD14 and LBP. Therefore, major modulatory effects from NSAID administration were not observed in this mastitis model, although a larger study mig

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Carbazoles; Cattle; Cell Count; Colony Count, Microbial; Complement C5a; Dinoprostone; Escherichia coli Infections; Female; Hematocrit; Interleukin-8; Lactation; Leukocyte Count; Lipopolysaccharide Receptors; Mastitis, Bovine; Milk; Parity; Pregnancy; Thromboxane B2

Studies in vitro reveal that endothelin-1 (ET-1) activates the alpha isoform of protein kinase C (PKC-alpha) in cultures of endothelial cells, thereby deranging cellular integrity. Sepsis and endotoxemia are associated with increased plasma concentrations of ET-1 that induce acute lung injury (ALI). We recently reported that non-selective ET-1 receptor blockade attenuates ALI in sheep by reducing the endotoxin-induced increase in extravascular lung water index (EVLWI). The aim of this study was to find out whether this attenuation is associated with reduced translocation of PKC-alpha from the cytosolic to the membrane fraction of lung tissue homogenate.. Seventeen awake, instrumented sheep were randomly assigned to a sham-operated group (n = 3), a lipopolysaccharide (LPS) group (n = 7) receiving an intravenous infusion of Escherichia coli 15 ng/kg per min for 24 hours, and a tezosentan group (n = 7) subjected to LPS and, from 4 hours, an intravenous injection of tezosentan 3 mg/kg followed by infusion at 1 mg/kg per hour for the reminder of the experiment. Pulmonary micro-occlusion pressure (Pmo), EVLWI, plasma concentrations of ET-1, tumor necrosis factor-a (TNF-a), and interleukin-8 (IL-8) were determined every 4 hours. Western blotting was used to assess PKC-alpha.. In non-treated sheep a positive correlation was found between the plasma concentration of ET-1 and Pmo in the late phase of endotoxemia (12 to 24 hours). A positive correlation was also noticed between Pmo and EVLWI in the LPS and the LPS plus tezosentan groups, although the latter was significantly reduced in comparison with LPS alone. In both endotoxemic groups, plasma concentrations of ET-1, TNF-alpha, and IL-8 increased. In the LPS group, the cytosolic fraction of PKC-alpha decreased by 75% whereas the membrane fraction increased by 40% in comparison with the sham-operated animals. Tezosentan completely prevented the changes in PKC-alpha in both the cytosolic and the membrane fractions, concomitantly causing a further increase in the plasma concentrations of ET-1, TNF-alpha, and IL-8.. In endotoxemic sheep, ET-1 receptor blockade alleviates lung injury as assessed by a decrease in EVLWI paralleled by a reduction in Pmo and the prevention of activation of PKC-alpha.

Topics: Animals; Endothelin A Receptor Antagonists; Endothelin-1; Endotoxemia; Enzyme Activation; Escherichia coli Infections; Interleukin-8; Protein Kinase C; Pyridines; Respiratory Distress Syndrome; Sheep; Tetrazoles; Tumor Necrosis Factor-alpha; Vasodilator Agents

Enterohemorrhagic Escherichia coli O157:H7 (EHEC) is an enteric pathogen that causes potentially fatal symptoms after intimate adhesion, modulation of intestinal epithelial signal transduction, and alteration of epithelial function (eg, barrier disruption). Although the epithelial barrier is critical to gut homeostasis, only a few agents, such as transforming growth factor (TGF)-beta, can enhance or protect epithelial barrier function. Our aims were to delineate the mechanism(s) behind TGF-beta-induced barrier enhancement and to determine whether TGF-beta could prevent EHEC-induced barrier disruption. Using monolayers of the human T84 colonic epithelial cell line, we found that TGF-beta induced a significant increase in transepithelial electrical resistance (a measure of paracellular permeability) through activation of ERK MAPK and SMAD signaling pathways and up-regulation of the tight junction protein claudin-1. Additionally, TGF-beta pretreatment of epithelia blocked the decrease in transepithelial electrical resistance and the increase in transepithelial passage of [(3)H]-mannitol caused by EHEC infection. EHEC infection was associated with reduced expression of zonula occludens-1, occludin, and claudin-2 (but not claudin-1 or claudin-4); TGF-beta pretreatment prevented these changes. These studies provide insight into EHEC pathogenesis by illustrating the mechanisms underlying TGF-beta-induced epithelial barrier enhancement and identifying TGF-beta as an agent capable of blocking EHEC-induced increases in epithelial permeability via maintenance of claudin-2, occludin, and zonula occludens-1 levels.

Topics: Cell Line, Tumor; Claudin-1; Enzyme-Linked Immunosorbent Assay; Epithelial Cells; Escherichia coli Infections; Escherichia coli O157; Flavonoids; Gene Expression Regulation; Humans; Interleukin-8; MAP Kinase Signaling System; Membrane Proteins; Tight Junctions; Transforming Growth Factor beta

The evolution and the relationship between inflammatory and renal-injury markers in women with acute uncomplicated pyelonephritis under antimicrobial therapy were investigated in a prospective study. Markers were measured before and 6 and 24 h after the intravenous administration of 1 g of ceftriaxone. Before treatment, the median levels of all markers except the serum creatinine levels were high. Twenty-four hours after the onset of antibiotic treatment, the C-reactive protein (CRP) level continued to be high, while the serum interleukin-6 (IL-6) levels and the urine IL-6, IL-8, albumin, and immunoglobulin G (IgG) levels decreased significantly. In contrast, serum creatinine and tumor necrosis factor alpha levels and urine N-acetyl-beta-glucosaminidase, alpha1-microglobulin, and beta2-microglobulin levels did not change over time. There was a significant correlation between IL-6 and IL-8 levels and urine albumin and IgG levels (urine albumin and IgG levels are glomerular and urinary tract-injury markers) as well as between serum CRP levels and the levels of the tubular-injury markers. In women with acute pyelonephritis, appropriate antibiotic treatment rapidly decreases serum IL-6 levels and urine IL-6 and IL-8 levels, which correlate well with urine albumin and IgG levels.

Topics: Acute Disease; Adult; Aged; Anti-Bacterial Agents; Biomarkers; C-Reactive Protein; Ceftriaxone; Escherichia coli Infections; Female; Humans; Immunoglobulin G; Inflammation Mediators; Interleukin-6; Interleukin-8; Kidney; Middle Aged; Prospective Studies; Pyelonephritis

The objective of the current study was to evaluate the dynamics of infection and the immunological response to varying numbers of Escherichia coli injected into the mammary glands of primiparous cows during the periparturient period. Primiparous cows have been shown to be more resistant to intramammary E. coli challenge, and an increase of the inoculum dose by 2 log10 units induced a more rapid clinical response and clearance of the organisms. Recognition of lipopolysaccharide (LPS) is a key event in the innate immunity response to gram-negative infection and is mediated by the accessory molecules CD14 and LPS-binding protein (LBP). Primiparous cows were inoculated with 1 x 10(4) (Group A; n=8) or 1 x 10(6) (Group B; n=8) cfu E. coli P4:O32 in their 2 left quarters during the periparturient period. Clinical examination and analysis of blood and milk parameters, including IL-8, complement fragment 5a (C5a), LBP, and soluble CD14 (sCD14), were performed from d -4 to d +3 relative to infection. Primiparous cows in Group B initiated a more rapid clinical response following intramammary infection (IMI), resulting in typical clinical signs and changes in blood and milk parameters approximately 3 h earlier compared with primiparous cows in Group A. Based on average milk production in the noninfected quarters on d +2 postinoculation, all heifers reacted as moderate responders. Distinct differences in the kinetic patterns of rectal temperature, somatic cell count (SCC), IL-8, C5a, LBP, and sCD14 were observed between both groups during the early phase of inflammation. Both C5a and IL-8 increased before cellular influx into the infected glands, followed by increases in sCD14 and LBP. In conclusion, primiparous cows are able to clear an intramammary E. coli infection efficiently. Moreover, increasing the inoculum dose induces a more rapid inflammatory reaction, mainly because of early activation of the innate host immune response.

Topics: Animals; Cattle; Cell Count; Colony Count, Microbial; Complement C5a; Escherichia coli; Escherichia coli Infections; Female; Immunity, Innate; Interleukin-8; Lipopolysaccharide Receptors; Mastitis, Bovine; Milk; Parity; Random Allocation

Interleukin (IL)-8 is a chemotactic cytokine that binds with high affinity to receptors on neutrophils. Previously we showed that (99m)Tc-labeled IL-8 is highly suitable for scintigraphic imaging in rabbit models of IM infection and of colitis.. (99m)Tc-labeled IL-8 was tested for its potential to image pulmonary infection in three experimental rabbit models: aspergillosis in immunocompromised rabbits, pneumococcal (Gram-positive) pneumonia, and Escherichia coli-induced (Gram-negative) pneumonia in immunocompetent rabbits (four rabbits in each group). A derivative of hydrazinonicotinamide was used as bifunctional coupling agent to label IL-8 with (99m)Tc. Biodistribution of (99m)Tc IL-8 was determined both by gamma-camera imaging and by counting dissected tissues at 6 h after injection.. (99m)Tc IL-8 enabled early (within 2 h after injection) and excellent visualization of localization and extent of pulmonary infection in each of the three experimental models of pulmonary infection. Uptake of (99m)Tc IL-8 in the infected lung and the contralateral lung was (in percentage of the injected dose per gram of tissue +/- SEM) at 6 h after injection 0.63 +/- 0.12 and 0.12 +/- 0.02 (aspergillosis), 0.89 +/- 0.04 and 0.44 +/- 0.04 (pneumococcal pneumonia), and 1.53 +/- 0.12 and 0.36 +/- 0.06 (E coli pneumonia), respectively. In the E coli model, uptake of (99m)Tc IL-8 in the focus of infection even exceeded uptake in the kidneys, the main clearing organs.. (99m)Tc IL-8 offers many advantages over the conventionally used radiopharmaceuticals to image pulmonary infection, (67)Ga citrate and radiolabeled leukocytes, ie, rapid and easy preparation, short time span between injection and imaging, low radiation burden and, most importantly, clear delineation of the infectious foci.

Topics: Animals; Aspergillosis; Escherichia coli Infections; Immunocompromised Host; Interleukin-8; Lung; Lung Diseases, Fungal; Organotechnetium Compounds; Pneumonia, Bacterial; Pneumonia, Pneumococcal; Rabbits; Radionuclide Imaging; Radiopharmaceuticals

This study investigated the role of P fimbriae in colonization of Escherichia coli, host response, and bacterial persistence in humans. Human volunteers were inoculated intravesically with the nonadherent ABU isolate E. coli 83972 and with P fimbriated transformants of the same strain. During the following 24 h all urine samples, and thereafter daily samples, were collected for urine culture, analysis of neutrophil numbers, and cytokine concentrations (IL-6 and IL-8). The P fimbriated transformants showed enhanced bacterial colonization in comparison to E. coli 83972 and lowered the bacterial numbers needed for persistent bacteriuria. The P fimbriated transformants also lowered the bacterial numbers needed for a significant neutrophil and cytokine host response. We conclude that P fimbriae enhance bacterial colonization and trigger the host response in the human urinary tract.

Topics: Adult; Ampicillin Resistance; Bacterial Adhesion; Child, Preschool; Colony Count, Microbial; Escherichia coli; Escherichia coli Infections; Female; Fimbriae, Bacterial; Genotype; Humans; Interleukin-6; Interleukin-8; Kanamycin Resistance; Leukocyte Count; Male; Neutrophils; Pyuria; Receptors, Immunologic; Recombination, Genetic; Urinary Tract Infections; Virulence

Blockade of tissue factor before lethal sepsis prevents acute lung injury and renal failure in baboons, indicating that activation of coagulation by tissue factor is an early event in the pathogenesis of acute lung injury and organ dysfunction. We hypothesized that blockade of tissue factor would also attenuate these injuries in established sepsis by prevention of further fibrin deposition and inflammation. Twelve male baboons received heat-killed Escherichia coli intravenously followed 12 hours later by live E. coli infusion. Six animals were treated 2 hours after the live bacteria with site-inactivated Factor VIIa, a competitive tissue factor inhibitor, and six animals were vehicle-treated sepsis control subjects. Animals were ventilated and monitored for 48 hours. Physiologic and hematologic parameters were measured every 6 hours, and pathologic evaluation was performed after 48 hours. Animals treated with site inactivated Factor VIIa had less severe lung injury, with preserved gas exchange, better lung compliance and histology scores, and decreased lung wet/dry weight. In treated animals, urine output was higher, metabolic acidosis was attenuated, and renal tubular architecture was protected. Coagulopathy was attenuated, and plasma interleukin-6, interleukin-8, and soluble tumor necrosis factor receptor-1 levels were significantly lower in the treated animals. These results show that blockade of coagulation attenuates acute lung and renal injury in established Gram-negative sepsis accompanied by antiinflammatory effects of therapy.

Topics: Acute Kidney Injury; Animals; Antigens, CD; Cytokines; Disease Models, Animal; Drug Monitoring; Escherichia coli Infections; Factor VIIa; Hemodynamics; Inflammation; Interleukin-6; Interleukin-8; Lung Compliance; Male; Papio; Pulmonary Gas Exchange; Receptors, Tumor Necrosis Factor; Receptors, Tumor Necrosis Factor, Type I; Respiratory Distress Syndrome; Sepsis; Severity of Illness Index; Thromboplastin

Bacterial protein synthesis starts with a formylated methionine residue, and this residue is sequentially cleaved away by a unique peptide deformylase (PDF) and a methionine aminopeptidase to generate mature proteins. The formylation-deformylation of proteins is a unique hallmark of bacterial metabolism and has recently become an attractive target for the development of antimicrobial agents. The innate immune system uses the formylation of bacterial proteins as a target, and professional phagocytes, e.g., neutrophils, express specific receptors for bacterium-derived formylated peptides. Activation of formyl peptide receptors (FPR) mediates neutrophil migration and the release of oxygen radicals and other antimicrobial substances from these cells. We hypothesize that the use of a PDF inhibitor would increase the production of proinflammatory peptides from the bacteria and thus trigger a more pronounced innate immune response. We tested this hypothesis by exposing Escherichia coli to subinhibitory doses of the PDF inhibitor actinonin and show that actinonin indeed increases the production and secretion of neutrophil-activating peptides that activate human neutrophils through FPR. These findings could be potentially used as a new approach to antibacterial chemotherapy.

Topics: Amidohydrolases; Aminopeptidases; Anti-Bacterial Agents; Bacteria; Chemotactic Factors; Chemotaxis, Leukocyte; Enzyme Inhibitors; Escherichia coli; Escherichia coli Infections; Humans; Hydroxamic Acids; In Vitro Techniques; Interleukin-8; NADPH Oxidases; Neutrophils

Enteroaggregative Escherichia coli (EAEC) infection can be identified in 26% of travelers with diarrhea and is associated with fecal interleukin (IL)-8 production. We hypothesized that single-nucleotide polymorphisms (SNPs) in the IL-8 gene are associated with EAEC-related symptoms. Fecal IL-8 production and IL-8 SNPs at 5 loci were identified in 69 US students who remained in Mexico for 5 weeks; 23 subjects had EAEC-associated diarrhea, 7 were asymptomatic EAEC carriers, 22 had nonspecific diarrhea, and 17 were asymptomatic without an enteropathogen. The chances of having EAEC-associated diarrhea were significantly increased among those with the AA genotype at the -251 position (odds ratio [OR], 208.51; 95% confidence interval [CI], 28.5-1525.36) and among those with AT genotype (OR, 14.3; 95% CI, 1.98-105.74), compared with those with the TT genotype at the -251 position. Among subjects with EAEC-associated diarrhea, the AA genotype at the -251 position produced greater concentrations of fecal IL-8 than those with the AT or TT genotype (P=.0053). In the present study, the AA genotype at the -251 position was associated with the occurrence of EAEC-associated diarrhea and increased levels of fecal IL-8.

Topics: Diarrhea; Escherichia coli; Escherichia coli Infections; Feces; Genetic Predisposition to Disease; Genotype; Humans; Interleukin-8; Mexico; Polymorphism, Single Nucleotide; Students; Travel; United States

There is increasing evidence that by facilitating translocation of Shiga toxin (Stx) across the intestinal epithelium and by transporting bound toxin to remote sites such as the renal endothelium, polymorphonuclear leukocytes (PMNs) play a key role in the pathogenesis of Shiga-toxigenic Escherichia coli (STEC) disease. Plasma levels of PMN-attracting CXC chemokines such as interleukin-8 (IL-8) also appear to correlate in humans with the severity of disease. Thus, the capacity of STEC strains to elicit CXC chemokine responses in intestinal epithelial cells may be a crucial step in pathogenesis. Accordingly, we attempted to determine which STEC factors are responsible for CXC chemokine induction in human colonic epithelial cells. Infection of Hct-8 cells with locus for enterocyte effacement (LEE)-negative STEC strains isolated from patients with severe STEC disease resulted in up-regulation of IL-8, macrophage inflammatory protein 2alpha (MIP-2alpha), MIP-2beta, and ENA-78 mRNA significantly higher and earlier than that elicited by several LEE-positive STEC strains, including the O157:H7 strain EDL933. Similarly, levels of IL-8 protein in LEE-negative STEC-infected Hct-8 culture supernatants were significantly higher than in LEE-positive STEC-infected culture supernatants. The difference in responses could not be attributed to the expression or nonexpression of LEE genes, the presence or absence of an STEC megaplasmid, or differences in O serogroups or in the type or amount of Stx produced. Interestingly, however, several of the LEE-negative STEC strains eliciting the strongest chemokine responses belonged to flagellar serotype H21. Incubation of Hct-8 cells with isolated H21 flagellin elicited IL-8 and MIP-2alpha responses similar to those seen in the presence of the most potent LEE-negative STEC strains. Deletion of the fliC gene, but not the stx(2) gene, largely abolished the capacity of O113:H21 LEE-negative STEC strain 98NK2 to elicit IL-8 and MIP-2alpha responses in Hct-8 cells. Taken together, these data suggest that although Stx is capable of inducing CXC chemokine responses, the elevated responses seen in cells infected with certain STEC strains are largely attributable to the production of flagellin.

Topics: Base Sequence; Cell Line; Chemokines, CXC; Colon; DNA; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Flagellin; Gastroenteritis; Genes, Bacterial; Humans; Interleukin-8; Mutation; Phosphoproteins; RNA, Messenger; Shiga Toxin 2; Up-Regulation; Virulence

Leukocytes are implicated in the pathogenesis of diarrhea-associated hemolytic uremic syndrome (D(+) HUS). We hypothesized that increased circulating levels of granulocyte colony-stimulating factor (G-CSF), and the chemokines epithelial cell-derived neutrophil-activating protein-78 (ENA-78), growth related oncogen-alpha (GRO-alpha), macrophage inflammatory protein-1beta (MIP-1beta), and monocyte chemotactic protein-1 (MCP-1) are related to the severity of illness in Escherichia coli O157:H7 infections. We compared the circulating concentrations of these mediators in the course of E. coli O157:H7 enteritis, hemorrhagic colitis, and HUS. Our data show that, on admission, children with HUS presented 10-fold abnormally increased levels of G-CSF (p < 0.007), 3-fold increased MIP-1beta concentrations (p < 0.001), and 2-fold lower values of ENA-78 (p < 0.0001). One week later, a further 4-fold decrease in ENA-78 concentration was noted (p < 0.0001) whereas MIP-1beta levels returned to normal. HUS patients requiring peritoneal dialysis showed 6-fold increased G-CSF (p < 0.001) and 5-fold decreased ENA-78 (p < 0.001) levels. On admission, children with uncomplicated O157:H7 hemorrhagic colitis (HC) presented 3-fold abnormally increased concentrations of G-CSF (p < 0.001) and MIP-1beta (p < 0.0001). Those with O157:H7 enteritis but no bloody stools showed higher rates of abnormal GRO-alpha, MIP-1beta, and MCP-1 measurements than children with O157:H7 HC or HUS: GRO-alpha (50% enteritis, 36% HC, 17% HUS; p < 0.06), MIP-1beta (40% enteritis, 22% HC, 11% HUS; p < 0.02), MCP-1 (77% enteritis, 20% HC, 18% HUS; p < 0.0001). The data indicates that GRO-alpha, MIP-1beta, and MCP-1 are produced during E. coli O157:H7 enteritis, whether or not HC or HUS develops. Our data suggest that children with O157:H7 associated HUS may present abnormally increased circulating levels of G-CSF and decreased ENA-78 concentrations. The mechanisms responsible for leukocytes recruitment in O157:H7 infections are unclear and await further studies.

Topics: Adolescent; Case-Control Studies; Chemokine CCL2; Chemokine CCL4; Chemokine CXCL1; Chemokine CXCL5; Chemokines; Chemokines, CC; Chemokines, CXC; Chemotactic Factors; Child; Child, Preschool; Colitis; Enteritis; Escherichia coli Infections; Escherichia coli O157; Female; Granulocyte Colony-Stimulating Factor; Hemolytic-Uremic Syndrome; Humans; Intercellular Signaling Peptides and Proteins; Interleukin-8; Macrophage Inflammatory Proteins; Male; Peritoneal Dialysis

Bacterial adherence to mucosal cells is a key virulence trait of pathogenic bacteria. The type 1 fimbriae and the P-fimbriae of Escherichia coli have both been described to be important for the establishment of urinary tract infections. While P-fimbriae recognize kidney glycosphingolipids carrying the Galalpha4Gal determinant, type 1 fimbriae bind to the urothelial mannosylated glycoproteins uroplakin Ia and Ib. The F1C fimbriae are one additional type of fimbria correlated with uropathogenicity. Although it was identified 20 years ago its receptor has remained unidentified. Here we report that F1C-fimbriated bacteria selectively interact with two minor glycosphingolipids isolated from rat, canine, and human urinary tract. Binding-active compounds were isolated and characterized as galactosylceramide, and globotriaosylceramide, both with phytosphingosine and hydroxy fatty acids. Comparison with reference glycosphingolipids revealed that the receptor specificity is dependent on the ceramide composition. Galactosylceramide was present in the bladder, urethers, and kidney while globotriaosylceramide was present only in the kidney. Using a functional assay, we demonstrate that binding of F1C-fimbriated Escherichia coli to renal cells induces interleukin-8 production, thus suggesting a role for F1C-mediated attachment in mucosal defense against bacterial infections.

Topics: Animals; Bacterial Adhesion; Chromatography, Thin Layer; Dogs; Escherichia coli; Escherichia coli Infections; Galactosylceramides; Glycosphingolipids; Humans; Interleukin-8; Magnetic Resonance Spectroscopy; Mucous Membrane; Rats; Receptors, Immunologic; Sphingosine; Trihexosylceramides; Urinary Tract; Urinary Tract Infections

Standard therapies including administration of potent antibiotics, aggressive fluid resuscitation and metabolic support have not been successful in relieving symptoms and reducing mortality associated with acute coliform mastitis. It is important to understand the pathophysiological response of the mammary gland to coliform infections when designing preventive or therapeutic regimens for controlling coliform mastitis. Our laboratory has previously shown that macrophages and polymorphonuclear neutrophils in milk express CD14 on their cell surface. In this study, we found that soluble CD14 (sCD14) is present in milk whey as a 46kDa protein reacted with anti-ovine CD14 antibody. Additional functional studies found that: (1) under serum-free condition, complexes of LPS-recombinant bovine soluble CD14 (rbosCD14) induced activation of mammary ductal epithelial cells (as measured by changes in interleukin-8 (IL-8) mRNA level by competitive RT-PCR) at low concentrations of LPS after 6 or 24h incubation (1-1000ng/ml), whereas LPS alone did not induce activation of mammary ductal epithelial cells at the same concentrations, and (2) intramammary injection of low concentrations of LPS did not increase concentration of leukocytes in milk. In contrast, LPS-rbosCD14 complex containing the same concentration of LPS increased the concentration of leukocytes in the injected mammary gland at 12 and 24h post-injection. These results indicate that rbosCD14 sensitizes mammary epithelial cells to low concentrations of LPS in vitro and in vivo. Endogenous sCD14 in milk may be important in initiating host responses to Gram-negative bacterial infections.

Topics: Animals; Antibodies, Monoclonal; Cattle; Epithelial Cells; Escherichia coli Infections; Female; Immunization; Interleukin-8; Lipopolysaccharide Receptors; Lipopolysaccharides; Mammary Glands, Animal; Mastitis, Bovine; Milk; Recombinant Proteins; Sheep; Solubility; Transcription, Genetic

Enterohemorrhagic Escherichia coli (EHEC) infections are associated with hemorrhagic colitis and the hemolytic-uremic syndrome (HUS). In vivo, elevated plasma levels of the proinflammatory cytokine interleukin-8 (IL-8) in EHEC-infected children are correlated with a high risk of developing HUS. As IL-8 gene transcription is regulated by the transcription factors NF-kappaB and AP-1, we analyzed the role of these factors in the regulation of IL-8 production after infection of the epithelial intestinal T84 cell line by EHEC. By 6 h of infection, EHEC had induced significant secretion of IL-8 (35.84 +/- 6.76 ng/ml versus 0.44 +/- 0.04 ng/ml in control cells). EHEC induced AP-1 and NF-kappaB activation by 3 h of infection. Moreover, the three mitogen-activated protein kinases (MAPK) (ERK1/2, p38, and JNK) were phosphorylated in EHEC-infected T84 cells concomitant with induction of AP-1 DNA binding activity, and IkappaB-alpha was phosphorylated and then degraded concomitant with induction of NF-kappaB DNA binding activity. Pretreatment of cells with the highly specific MEK1/2 inhibitor U0126, the p38 inhibitor SB203580, and/or the proteasome inhibitor ALLN led to inhibition of the IL-8 secretion induced in EHEC-infected T84 cells. These findings demonstrate that (i) EHEC can induce in vitro a potent proinflammatory response by secretion of IL-8 and (ii) the secretion of IL-8 is due to the involvement of MAPK, AP-1, and NF-kappaB signaling pathways.

Topics: Cell Line; Enzyme Activation; Escherichia coli Infections; Escherichia coli O157; Humans; Interleukin-8; Lipopolysaccharides; Mitogen-Activated Protein Kinases; NF-kappa B; Phosphorylation; Transcription Factor AP-1

Although various inflammatory mediators have been previously shown to be released into the peritoneal cavity during peritonitis in peritoneal dialysis patients, those that are involved in governing changes in peritoneal permeability to small solutes and protein remain incompletely defined.. We determined the importance of prostanoid production in the enhanced protein loss observed during acute peritonitis by inhibition experiments using indomethacin, an inhibitor of cyclooxygenase activity. The association between changes in peritoneal permeability and the generation of inflammatory mediators after adding Escherichia coli to peritoneal dialysate was first examined in series 1 experiments. Series 2 experiments then determined the effect of intraperitoneal administration of indomethacin (75 microg/mL) on changes in peritoneal permeability after adding E. coli to peritoneal dialysate. All experiments were performed in male New Zealand White rabbits (2.6 to 3.4 kg body weight) using an eight-hour dwell of dialysate containing 2.5% glucose. Peritoneal permeability to creatinine and protein was assessed by time-dependent changes in the dialysate to plasma concentration ratios of these solutes.. Series 1 experiments showed enhanced leukocyte migration into the peritoneal cavity and increased peritoneal permeability to protein during bacterial challenge that was accompanied by an increase in the dialysate concentrations of prostaglandin E2 (PGE2), 6-keto-PGF1alpha, and interleukin-8, but not nitrate + nitrite (a measure of local nitric oxide production). Inhibition of prostanoid production by intraperitoneal administration of indomethacin in series 2 experiments resulted in lower dialysate concentrations of PGE2 and 6-keto-PGF1alpha and in lower peritoneal permeability to protein, both to control levels. No effect of indomethacin on transperitoneal migration of leukocytes or the generation of interleukin-8 was observed.. Enhanced production of prostanoids likely plays an important role in governing the increase in peritoneal permeability to protein during acute, bacterial peritonitis in the rabbit.

Topics: 6-Ketoprostaglandin F1 alpha; Animals; Cell Movement; Cyclooxygenase Inhibitors; Dialysis Solutions; Dinoprostone; Escherichia coli Infections; Indomethacin; Injections, Intraperitoneal; Interleukin-8; Leukocytes; Male; Peritoneum; Peritonitis; Prostaglandins; Proteins; Rabbits

The purpose of this study was to investigate whether anisodamine could inhibit Shiga toxin-1 (Stx1)-induced cytokine production and increase the survival of Stx1-treated mice. Human monocytic cells were stimulated by Stx1 (1 to 100 ng/mL) with or without anisodamine addition (1 to 400 microg/mL). For in vivo evaluations, C57BL/6 mice were given a single intraperitoneal injection of anisodamine (1 mg) or saline solution after intraperitoneal injection of Stx1 (2.75 microg/kg). The results showed that anisodamine significantly suppressed Stx1-induced tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and IL-8 production. Reverse transcriptase-polymerase chain reaction (RT-PCR) showed that anisodamine suppressed Stx1-mediated TNF-alpha mRNA expression. Further study showed that this TNF-alpha inhibitory effect was via a prostaglandin E2-dependent mechanism. Anisodamine treatment prolonged the survival time of mice and decreased the lethality of Stx1 (94.5% to 44%). Because cytokines, in particular TNF-alpha, contribute to the pathologic process in Stx-producing Escherichia coli (STEC) infection, this study suggested that anisodamine could be a potential drug for treatment of STEC infection.

Topics: Animals; Cell Line; Cell Survival; Cytokines; Dinoprostone; Escherichia coli Infections; Gene Expression; Humans; Injections, Intraperitoneal; Interleukin-1; Interleukin-8; Male; Mice; Mice, Inbred C57BL; Monocytes; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Shiga Toxin 1; Solanaceous Alkaloids; Survival Rate; Tumor Necrosis Factor-alpha

Interleukin-8 (IL-8) is a chemotactic cytokine involved in activation and recruitment of neutrophils to areas of infection. In our previous studies in rabbits we tested 123I-labeled IL-8 for its potential to image infections and showed that IL-8 rapidly and efficiently accumulated in infectious foci. However, labeling of IL-8 with 123I is costly and laborious and the specific activity of the preparation was low. In this study IL-8 was labeled with 99mTc through the hydrazinonicotinamide (HYNIC) chelator.. The leukocyte receptor-binding capacity of the preparation was determined in vitro. Rabbits with Escherichia coli abscesses were injected intravenously with 7 MBq 99mTc-HYNIC-IL-8. Biodistribution of the radiolabel was determined by gamma camera imaging and tissue counting at 8 h after injection. 99mTc-HYNIC-lysozyme was used as a size-matched control.. The leukocyte receptor-binding capacity of the 99mTc-HYNIC-IL-8 preparation was preserved as determined in vitro, but labeling efficiency was modest with a specific activity of 3 MBq/microg. 99mTc-HYNIC-IL-8 accumulated rapidly in the abscess up to 0.33 +/- 0.06 percentage injected dose per gram (%ID/g) at 8 h after injection (vs. 0.025 +/- 0.003 %lD/g for 99mTc-HYNIC-lysozyme). Total uptake in the abscess was 4.9 +/- 0.7 %ID (vs. 0.44 +/- 0.05 %ID for 99mTc-HYNIC-lysozyme). Abscess-to-contralateral muscle ratios increased up to 127 +/- 23 (compared with 6.7 +/- 1.1 for 99mTc-HYNIC-lysozyme) and abscess-to-blood ratios increased to 11.9 +/- 2.2 (0.24 +/- 0.03 for 99mTc-HYNIC-lysozyme). The radiolabel was excreted renally, with a retention in the kidneys of 28 %ID. Gamma camera imaging rapidly visualized the abscess from 1 h after injection onward, with abscess-to-background ratios improving with time up to 22 at 8 h after injection (vs. 2.7 for 99mTc-HYNIC-lysozyme), as determined by quantitative analysis of the images. Most important, only a transient (30 min) moderate drop of leukocyte counts and no leukocytosis were observed after injection of an imaging dose of 99mTc-HYNIC-IL-8.. IL-8 can be labeled with 99mTc using HYNIC as a chelator. By this method the leukocyte receptor-binding capacity is preserved. The preparation allows rapid visualization of infection in a rabbit model with high target-to-background ratios. The mild transient drop of leukocyte counts and the absence of leukocytosis suggest that 99mTc-HYNIC-IL-8 may be used as an imaging agent with only mild and transient side effects.

Topics: Abscess; Animals; Escherichia coli Infections; Female; Humans; Hydrazines; Interleukin-8; Isotope Labeling; Ligands; Nicotinic Acids; Rabbits; Radioimmunodetection; Radiopharmaceuticals; Technetium; Tissue Distribution

Urinary tract epithelial cells (T 24/83) are able to express interleukin (IL)-6, IL-8, platelet-derived growth factor (PDGF) and tumour necrosis factor-alpha, but not IL-1 beta, IL-2, IL-4 and IL-10 in response to an infection with uropathogenic bacteria. The process of cytokine secretion is time dependent, with a significant increase in the cytokine activity after 60 min. The expression of virulence factors of the bacteria does not seem to play a role. The interaction between bacterial products (e.g. lipopolysaccharide) and/or bacterial adhesion mediated by adhesins and specific receptor molecules of cell surfaces may be responsible for the activity of mediator protein expression in the epithelial cells. The release of PDGF and IL-8 was found to be higher when due to Escherichia coli HB 101 (rough form) than that caused by other bacterial strains. Citrobacter CB 3009 provoked the highest level of IL-6. The PDGF level correlated significantly with IL-6 and IL-8 values (P<0.001). There was a significant correlation between the time-dependent release of IL-6 and IL-8 (P<0.05). In epithelial cytokine response to bacterial infection, the reaction of the epithelial cells may modify themselves (e.g. internalization of bacteria) and the immuno-regulatory processes that are caused by infection and responsible for parenchymal injury.

Topics: Citrobacter; Cytokines; Enterobacteriaceae Infections; Enzyme-Linked Immunosorbent Assay; Escherichia coli; Escherichia coli Infections; Humans; Interleukin-6; Interleukin-8; Platelet-Derived Growth Factor; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha; Urinary Bladder Neoplasms; Urinary Tract Infections; Urogenital System; Urothelium; Virulence

Neutrophil migration to infected mucosal sites involves a series of complex interactions with molecules in the lamina propria and at the epithelial barrier. Much attention has focussed on the vascular compartment and endothelial cells, but less is known about the molecular determinants of neutrophil behavior in the periphery. We have studied urinary tract infections (UTIs) to determine the events that initiate neutrophil recruitment and interactions of the recruited neutrophils with the mucosal barrier. Bacteria activate a chemokine response in uroepithelial cells, and the chemokine repertoire depends on the bacterial virulence factors and on the specific signaling pathways that they activate. In addition, epithelial chemokine receptor expression is enhanced. Interleukin (IL)-8 and CXCR1 direct neutrophil migration across the epithelial barrier into the lumen. Indeed, mIL-8Rh knockout mice showed impaired transepithelial neutrophil migration, with tissue accumulation of neutrophils, and these mice developed renal scarring. They had a defective antibacterial defense and developed acute pyelonephritis with bacteremia. Low CXCR1 expression was also detected in children with acute pyelonephritis. These results demonstrate that chemokines and chemokine receptors are essential to orchestrate a functional antimicrobial defense of the urinary tract mucosa. Mutational inactivation of the IL-8R caused both acute disease and chronic tissue damage.

Topics: Animals; Bacterial Adhesion; Bacteriuria; Chemotaxis, Leukocyte; Child; Disaccharides; Drosophila Proteins; Escherichia coli; Escherichia coli Infections; Fimbriae, Bacterial; Genetic Predisposition to Disease; Glycosphingolipids; Humans; Immunity, Innate; Interleukin-8; Macrophages; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Knockout; Mucous Membrane; Neutrophils; Pyelonephritis; Receptors, Cell Surface; Receptors, Chemokine; Receptors, Interleukin-8A; Recurrence; Toll-Like Receptors; Urinary Tract Infections; Urothelium; Virulence

Enteropathogenic Escherichia coli (EPEC) is an extracellular bacterial pathogen that infects the human intestinal epithelium and is a major cause of infantile diarrhea in developing countries. EPEC belongs to the group of attaching and effacing (A/E) pathogens. It uses a type III secretion system to deliver proteins into the host cell that mediate signal transduction events in host cells. We used gene array technology to study epithelial cell responses to EPEC infection at the level of gene expression. We found that EPEC induces the expression of several genes in infected HeLa cells by a lipopolysaccharide (LPS)-independent mechanism, including cytokines and early growth response factor 1 (Egr-1). The transcription factor Egr-1 is an immediate-early-induced gene that is activated in most cell types in response to stress. EPEC-induced upregulation of egr-1 is mediated by the activation of the MEK/extracellular signal-regulated kinase signal transduction pathway and is dependent on the type III secretion system. egr-1 is also induced during infection of mice by the A/E pathogen Citrobacter rodentium, suggesting that both Egr-1 and the activation of this mitogen-activated protein kinase signal transduction pathway may play a role in disease.

Topics: Animals; Blotting, Northern; Citrobacter freundii; DNA-Binding Proteins; Early Growth Response Protein 1; Enterobacteriaceae Infections; Enzyme Activation; Epithelial Cells; Escherichia coli Infections; Gene Expression; HeLa Cells; Humans; Immediate-Early Proteins; Interleukin-8; Lipopolysaccharides; MAP Kinase Signaling System; Mice; Mice, Inbred C57BL; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Phosphorylation; Reverse Transcriptase Polymerase Chain Reaction; Transcription Factors

The Yucatan micropig has been used to develop an experimental model of chronic bacteremia. This animal exhibits clinical and biological characteristics that are close to those in humans, and the pharmacokinetic behaviours of many classes of drugs in this model are similar to those in man. Six adult female were intravenously inoculated with a mean Escherichia coli inoculum of 5.1 x 10(9) bacteria. During five days of spontaneous evolution, the medical follow-up includes biological, clinical and bacteriological parameters. A systemic inflammatory syndrome, a sepsis, an organ insufficiency and positive blood cultures mimic the human disease. In all animals there is an adynamia, a lack of motor coordination, an anorexia, a tachypnea, a fever, a leuconeutropenia followed by an hyperleucocytosis, an anemia, a thrombopenia, an acute tubulonephritis and an elevated sedimentation rate. In some cases, there is an increase of the C reactive protein, in others, an increase of IL-6 and IL-8. At day five, all animals are alive, and five micropigs have positive blood cultures. This chronic, reproducible model is thus suitable for further antibacterial treatments evaluations.

Topics: Acute Kidney Injury; Acute-Phase Reaction; Animals; Anorexia; Ataxia; Bacteremia; Chronic Disease; Disease Progression; Escherichia coli Infections; Fever; Hematologic Diseases; Interleukin-6; Interleukin-8; Models, Animal; Multiple Organ Failure; Nephritis, Interstitial; Reproducibility of Results; Swine, Miniature; Systemic Inflammatory Response Syndrome

Postoperative cholangitis is one of the most common complications after bile duct reconstruction. The pathogenesis and early consequences of ascending cholangitis still are unidentified.. Male Sprague-Dawley rats were divided into 5 treatment groups: control (n = 4), blood sampling and liver biopsy only; group I, [BDL/Eschericha coli; n = 6], ligation of common bile duct (BDL) for a week, followed by Roux-en-Y choledochojejunostomy (RYCJ) and injection of E coli (ATCC 25922) into Roux limb after 24 hours; group II, [BDL/NS; n = 5], same procedures as in group I, with injection of normal saline (NS) into Roux limb; group III, [SBDL/E coli; n = 6], primary RYCJ was constructed 1 week after sham ligation of common bile duct (SBDL) followed by the same treatment as group I; Group IV, [SBDL/N.S; n = 6], same procedures as in group III, but injecting NS into Roux limb. All animals were killed after 24 hours of treatment. Blood was sampled for culture and serum cytokine levels. The liver was harvested for quantitative bacterial culture, as well as for MCP-1, interleukin (IL)-8 (CINC in the rat) and transforming growth factor beta1 mRNA expression by reverse transcriptase polymerase chain reaction (RT-PCR) and for immunohistochemistry. The choledochojejunostomy was resected for culture. Serum cytokine levels were detected by ELISA kits.. A significant increase of E coli ATCC 25922, occurred in the livers of group I rats, compared with group IV (P =.037). MCP-1 expression increased in all groups, compared with control (P =.000). The IL-8 mRNA expression was significantly higher in group I than in control (P =.021). The expression of TGF-beta1 mRNA was similar among the groups (P =.361), consistent with the immunohistochemistry results. The serum MCP-1 and IL-8 levels were higher in the 4 groups than in the control (P =.000) and were significantly higher in group I than in group IV (P =.001).. This study found that a significant colonization of E coli of the same strain was present in the cholestatic rat liver injected into the Roux limb, which was associated with a higher expression of liver MCP-1 and IL-8 mRNA, a significant increase of serum MCP-1 and IL-8, and a more evident inflammatory cell infiltration into the porta hepatis.

Topics: Anastomosis, Roux-en-Y; Animals; Chemokine CCL2; Cholangitis; Choledochostomy; Cholestasis, Intrahepatic; Common Bile Duct; Escherichia coli; Escherichia coli Infections; Interleukin-8; Ligation; Liver Cirrhosis; Male; Postoperative Complications; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta

In upper urinary tract infections, tubular epithelial cells (TEC) may play a pivotal role in the initiation of the renal inflammatory response. They exert crucial immunological functions such as processing and presentation of foreign antigen, secretion of proinflammatory cytokines (interleukin-6 [IL-6] and tumor necrosis factor alpha) and chemokines (IL-8, MCP-1, ENA-78, and RANTES). Since monolayer cultures are a limited model for polarized tubular epithelial cells, we studied the side-dependent IL-8 secretion of TEC by using cell culture inserts as a basement membrane imitation. Primary cultures of proximal TEC were stimulated with differently fimbriated mutants of Escherichia coli, E. coli LPS, S-fimbria isolates, and IL-1alpha. IL-8 protein was measured by enzyme-linked immunosorbent assay, and IL-8-like biological activity was tested by measuring elastase release from polymorphonuclear cells in supernatants of the upper and lower compartments. IL-8 mRNA was compared by competitive PCR. IL-8 secretion by TEC into the basolateral environment was significantly higher than secretion into the apical compartment, representing the tubular lumen. However, stimulation of IL-8 secretion by TEC was restricted to IL-1alpha and was not inducible by E. coli mutants, S fimbriae, or lipopolysaccharide. With this in vitro model of polarized TEC, we show that luminal contact of TEC with uropathogenic E. coli does not result in enhanced IL-8 secretion. The basolaterally directed production of the neutrophil chemotactic factor IL-8 by TEC after stimulation with IL-1alpha might play an important role in the initiation of inflammatory cell influx into the renal parenchyma.

Topics: Base Sequence; Cell Polarity; Cells, Cultured; DNA Primers; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Fimbriae, Bacterial; Gene Expression; Humans; Immunohistochemistry; Interleukin-1; Interleukin-8; Kidney Cortex; Kidney Tubules; Lipopolysaccharides; Microscopy, Electron; Mutation; Pyelonephritis; RNA, Messenger

Plasma-derived antithrombin III (ATIII) prevents the lethal effects of Escherichia coli infusion in baboons, but the mechanisms behind this effect are not clear. In the present study, we evaluated the effects of recombinant human ATIII (rhATIII) on the clinical course and the inflammatory cytokine and coagulation responses in baboons challenged with lethal dose of E coli. Animals in the treatment group (n = 5) received high doses of rhATIII starting 1 hour before an E coli challenge. Those in the control group were administered saline. Survival was significantly improved in the treatment group (P =.002). Both groups had similar hemodynamic responses to E coli challenge but different coagulation and inflammatory responses. The rhATIII group had an accelerated increase of thrombin-ATIII complexes and significantly less fibrinogen consumption compared to controls. In addition, the rhATIII group had much less severe thrombotic pathology on autopsy and virtually no fibrinolytic response to E coli challenge. Furthermore, the rhATIII group had a significantly attenuated inflammatory response as evidenced by marked reduction of the release of various cytokines. We conclude that the early administration of high doses of rhATIII improves the outcome in baboons lethally challenged with E coli, probably due to the combined anticoagulation and anti-inflammatory effects of this therapy. (Blood. 2000;95:1117-1123)

Topics: Animals; Antithrombin III; Blood Cell Count; Blood Coagulation; Blood Pressure; Drug Administration Schedule; Escherichia coli Infections; Female; Fibrinogen; Heart Rate; Humans; Inflammation; Interleukin-10; Interleukin-6; Interleukin-8; Male; Papio; Peptide Hydrolases; Recombinant Proteins; Survival; Time Factors; Tumor Necrosis Factor-alpha

The prompt recruitment of neutrophils to the site of infection is essential for the defense of the bovine mammary gland against invading pathogens and is determinant for the outcome of the infection. Escherichia coli is known to induce clinical mastitis, characterized by an intense neutrophil recruitment leading to the eradication of the bacteria, whereas Staphylococcus aureus induces subclinical mastitis accompanied by a moderate neutrophil recruitment and the establishment of chronic mastitis. To elicit the neutrophil recruitment into the udder, inflammatory mediators must be produced after recognition of the invading pathogen. To our knowledge, those mediators have never been studied during S. aureus mastitis, although understanding of the neutrophil recruitment mechanisms could allow a better understanding of the differences in the pathogeneses elicited by E. coli and S. aureus. Therefore, we studied, at several time points, the accumulation of neutrophils and the presence of the chemoattractant complement fragment C5a and of the cytokines interleukin-1beta (IL-1beta), tumor necrosis factor alpha, and IL-8 in milk after inoculation of E. coli or S. aureus in lactating bovine udders. The low levels of C5a and the absence of cytokines in milk from S. aureus-infected cows, compared to the high levels found in milk from E. coli-infected animals, mirror the differences in the severities of the two inflammatory reactions. The cytokine deficit in milk after S. aureus inoculation in the lactating bovine mammary gland could contribute to the establishment of chronic mastitis. This result could help in the design of preventive or curative strategies against chronic mastitis.

Topics: Animals; Cattle; Complement C5a; Escherichia coli Infections; Female; Haptoglobins; Interleukin-1; Interleukin-8; Mastitis; Serum Albumin, Bovine; Staphylococcal Infections; Staphylococcus aureus; Tumor Necrosis Factor-alpha

Several small receptor-binding agents have been tested for imaging of infection and inflammation. The potential of chemotactic peptides and of interleukins is promising and superior to that of conventional agents. In this study, we investigated the potential of interleukin-8 (IL-8) to image infection in rabbits.. IL-8 was labeled with 123I using the Bolton-Hunter method. Twenty-fours hours after induction of Escherichia coli abscesses in the left thigh muscle, rabbits were injected intravenously with 18.5 MBq 123I-IL-8. Gamma camera images were obtained at 5 min and at 1, 4, and 8 h after injection. Biodistribution was determined 8 h after injection.. 123I-IL-8 rapidly cleared from the blood. Accumulation of 123I-IL-8 in the abscess was visible as early as 1 h after injection. The highest abscess uptake was obtained 4 h after injection (2.6+/-0.2 percentage injected dose [%ID]), whereas 123I-IL-8 rapidly cleared from all other tissues. This resulted in increases in abscess-to-background ratios to 13.0+/-0.7 (8 h after injection), as determined by quantification of the images. In tissue biodistribution (8 h after injection), the abscess uptake was 0.057+/-0.011 %ID/g with abscess-to-contralateral muscle ratios of 114.7+/-23.0. The radioiodination method clearly affected the in vivo biodistribution of IL-8 because IL-8 iodinated using the lodo-Gen method cleared significantly slower from the blood and most other organs, resulting in poor visualization of the abscess.. The superior characteristics of IL-8 radioiodinated using the Bolton-Hunter method--i.e., high abscess uptake and rapid background clearance within a few hours--make IL-8 a promising agent to image infection and inflammation.

Topics: Abscess; Animals; Escherichia coli Infections; Humans; Interleukin-8; Iodine Radioisotopes; Rabbits; Radioimmunodetection; Time Factors; Tissue Distribution

To investigate the effects of lidocaine administration on hemodynamics and cytokine concentrations in Escherichia coli endotoxemia in rabbits.. Randomized, prospective laboratory study.. University laboratory.. Thirty-two Japanese rabbits anesthetized with urethane and ventilated mechanically.. Animals were randomly assigned to one of four groups: endotoxemic controls (n = 8), receiving intravenous E. coli endotoxin (0.5 mg/kg bolus) via the mesenteric vein; laparotomy controls (n = 8), treated identically to the endotoxemic controls except for the substitution of 0.9% saline for endotoxin; lidocaine controls (n = 8), treated identically to the laparotomy controls with the addition of intravenous lidocaine (3 mg/kg bolus followed by infusion at 2 mg/kg/hr) administered immediately after the injection of 0.9% saline; and lidocaine-treated rabbits (n = 8), treated identically to the endotoxemic controls with the addition of intravenous lidocaine (3 mg/kg bolus followed by infusion at 2 mg/kg/hr) administered immediately after the injection of endotoxin.. We compared the cardiac output, systemic vascular resistance, blood gases, and plasma cytokine concentrations (tumor necrosis factor, interleukin [IL]-6, and IL-8) for each group. After endotoxin injection, the mean arterial pressure, cardiac output, and systemic vascular resistance decreased progressively in the endotoxemic controls. At 4 hrs after injection, all of the variables except the heart rate and central venous pressure were lower in the endotoxemic controls than in the other groups. At 4 hrs after endotoxin injection, both IL-6 and IL-8 concentrations increased in all groups. However, the mean concentrations of IL-6 and IL-8 in the endotoxemic controls significantly exceeded those in the other groups. No significant differences existed between the laparotomy controls and lidocaine-treated rabbits.. Lidocaine had a profound inhibitory effect on the hemodynamic and cytokine responses to endotoxemia when it was administered immediately after exposure to endotoxin. Our results demonstrate the potential usefulness of lidocaine as an anti-inflammatory agent in endotoxemia.

Topics: Animals; Blood Gas Analysis; Cytokines; Endotoxemia; Escherichia coli Infections; Hemodynamics; Interleukin-6; Interleukin-8; Lidocaine; Prospective Studies; Rabbits; Random Allocation; Time Factors

Shiga toxin, produced by Escherichia coli O157:H7, is important for the pathogenicity of the epidemic form of hemolytic uremic syndrome (HUS). This toxin has recently been found to stimulate endothelin-1 synthesis in cultured endothelial cells in vitro.. We investigated endothelin and cytokine levels in sera during a large outbreak of E. coli O157:H7 infection in Osaka, Japan, in 1996. Eleven patients with HUS and 9 patients with hemorrhagic colitis at the onset of E. coli O157:H7 infection were studied.. Serum IL-6 (p < 0.01), IL-8 (p < 0.05), IL-10 (p < 0.001) and endothelin (p < 0.001) levels were significantly increased in patients with HUS compared to those with colitis only. The serum thrombomodulin level, a molecular marker of endothelial damage, also showed a significant positive correlation with serum IL-6 (p < 0.01), IL-8 (p < 0.01), IL-10 (p < 0.01) and endothelin (p < 0.001) levels. In a HUS patient, the increase in serum IL-10 and endothelin levels reached a plateau prior to the peak of serum creatinine levels.. Increased serum endothelin synthesis by Shiga toxin in vivo was proven in HUS secondary to E. coli O157:H7 infection. Increased serum endothelin and IL-10 levels were speculated to be associated with the development of HUS through vascular endothelial damage caused by E. coli O157:H7 infection.

Topics: Case-Control Studies; Colitis; Creatinine; Disease Outbreaks; Endothelin-1; Escherichia coli Infections; Escherichia coli O157; Female; Gastrointestinal Hemorrhage; Hemolytic-Uremic Syndrome; Humans; Interleukin-10; Interleukin-6; Interleukin-8; Japan; Male; Thrombomodulin; Time Factors

CD14, a pattern recognition receptor found on myeloid cells, is a critical component of the innate immune system that mediates local and systemic host responses to Gram-negative and Gram-positive bacterial products. Previous studies in normal animals have tested the effect of CD14 blockade on the systemic response to i.v. LPS. The goals of the study were to determine whether CD14 blockade protected against the deleterious systemic response associated with Escherichia coli pneumonia and to determine whether this strategy affected the pulmonary response to tissue infection. Rabbits were pretreated with either anti-CD14 mAb or isotype control mAb at 2.5 mg/kg. E. coli (1 x 109 CFU) was inoculated into the lungs, and the animals were observed for either 4 or 24 h. The blockade of CD14 improved the mean arterial blood pressure (p = 0.001) and decreased the i.v. fluid requirements (p = 0.01). Although this therapy protected the vascular compartment, rabbits treated with anti-CD14 mAb had increased bacterial burdens in the bronchoalveolar lavage fluid recovered from the instilled lung (p = 0.005) and widened alveolar-arterial oxygen difference. Blockade of CD14 prevents the deleterious systemic responses that occur in sepsis; however, other measures are necessary to control bacterial proliferation at the primary site of infection.

Topics: Animals; Antibodies, Monoclonal; Blood Pressure; Bronchoalveolar Lavage Fluid; Chemokine CCL2; Chemotactic Factors; Escherichia coli Infections; Growth Substances; Inflammation; Interleukin-8; Lipopolysaccharide Receptors; Lung; Pneumonia, Bacterial; Pulmonary Gas Exchange; Rabbits; Sepsis; Tumor Necrosis Factor-alpha

Cytokine mediators and leukocyte-endothelial cell adhesion molecules are critical and interdependent components of the acute inflammatory response in sepsis. We hypothesized that the administration of monoclonal antibodies to intercellular adhesion molecule-1 (CD54) or E- and L-selectin (CD62E/L) would decrease serum levels of the proinflammatory cytokines interleukin-1beta (IL-1), IL-6, and IL-8 and tumor necrosis factor receptor (TNFR-1) in baboons during sepsis. Adult male baboons received infusions of 1 x 10(9) colony forming units (CFU)/kg heat-killed Escherichia coli (E. coli) followed 12 h later by live E. coli (1 x 10(10) CFU/kg). At the time of live bacterial infusion, six septic animals were treated with a monoclonal antibody to CD54 and six with an antibody to CD62E and L (1 mg/kg). Eight untreated septic animals served as controls. Sequentially drawn serum samples were assayed for IL-1, IL-6, IL-8, and TNFR-1 using enzyme-linked immunoassay (ELISA). Data were compared using Mann-Whitney U tests and Chi-square analyses. Median survival was decreased in both treatment groups compared to controls (P < 0.05). Peak IL-1 level was higher than controls in septic animals treated with anti-CD54 but not anti-CD62E/L (P < 0.05, P = NS, respectively). Elevations in IL-6, IL-8, and TNFR-1 were increased and prolonged in both antibody treated groups compared to controls (P < 0.05). These results provide the first in vivo evidence that leukocyte-endothelial adhesion molecules CD54 and CD62E/L regulate cytokine production in sepsis.

Topics: Animals; Antibodies, Monoclonal; Antigens, CD; Cytokines; E-Selectin; Escherichia coli; Escherichia coli Infections; Inflammation Mediators; Intercellular Adhesion Molecule-1; Interleukin-1; Interleukin-6; Interleukin-8; L-Selectin; Male; Papio; Receptors, Tumor Necrosis Factor; Receptors, Tumor Necrosis Factor, Type I; Sepsis

Pyelonephritis is one of the most common febrile diseases in children. If not treated appropriately, it causes irreversible renal damage and accounts for a large proportion of end stage renal failures. Renal scarring can occur in the absence of inflammatory cells, indicating that bacteria may have a direct signalling effect on renal cells. Intracellular calcium ([Ca2+]i) oscillations can protect cells from the cytotoxic effects of prolonged increases in intracellular calcium. However, no pathophysiologically relevant protein that induces such oscillations has been identified. Here we show that infection by uropathogenic Escherichia coli induces a constant, low-frequency oscillatory [Ca2+]i response in target primary rat renal epithelial cells induced by the secreted RTX (repeats-in-toxin) toxin alpha-haemolysin. The response depends on calcium influx through L-type calcium channels as well as from internal stores gated by inositol triphosphate. Internal calcium oscillations induced by alpha-haemolysin in a renal epithelial cell line stimulated production of cytokines interleukin (IL)-6 and IL-8. Our findings indicate a novel role for alpha-haemolysin in pyelonephritis: as an inducer of an oscillating second messenger response in target cells, which fine-tunes gene expression during the inflammatory response.

Topics: Animals; Bacterial Proteins; Calcium; Calcium Channel Blockers; Calcium Channels, L-Type; Cell Line; Epithelial Cells; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Estrenes; Female; Hemolysin Proteins; Humans; Interleukin-6; Interleukin-8; Kidney; Nifedipine; Pyelonephritis; Pyrrolidinones; Rats; Rats, Sprague-Dawley

Neutrophil migration across infected mucosal surfaces is chemokine dependent, but the role of chemokine receptors has not been investigated. In this study, chemokine receptors were shown to be expressed by epithelial cells lining the urinary tract, and to play an essential role for neutrophil migration across the mucosal barrier. Uroepithelial CXCR1 and CXCR2 expression was detected in human urinary tract biopsies, and in vitro infection of human uroepithelial cell lines caused a dramatic increase in both receptors. As a consequence, there was higher binding of IL-8 to the cells and the IL-8-dependent neutrophil migration across the infected epithelial cell layers was enhanced. Abs to IL-8 or to the CXCR1 receptor inhibited this increase by 60% (p<0.004), but anti-CXCR2 Abs had no effect, suggesting that CXCR1 was the more essential receptor in this process. Similar observations were made in the mouse urinary tract, where experimental infection stimulated epithelial expression of the murine IL-8 receptor, followed by a rapid flux of neutrophils into the lumen. IL-8 receptor knockout mice, in contrast, failed to express the receptor, their neutrophils were unable to cross the epithelial barrier, and accumulated in massive numbers in the tissues. These results demonstrate that epithelial cells express CXC receptors and that infection increases receptor expression. Furthermore, we show that CXCR1 is required for neutrophil migration across infected epithelial cell layers in vitro, and that the murine IL-8 receptor is needed for neutrophils to cross the infected mucosa of the urinary tract in vivo.

Topics: Animals; Cell Communication; Cell Line; Cell Movement; Epithelial Cells; Escherichia coli Infections; Female; Genetic Predisposition to Disease; Humans; Interleukin-8; Mice; Mice, Inbred BALB C; Mice, Knockout; Neutrophils; Protein Binding; Receptors, Interleukin-8A; Receptors, Interleukin-8B; Urinary Tract Infections; Urothelium

To investigate bacterial growth and inflammatory mediator release in the early stage of the immune response, a unilateral acute ascending pyelonephritis was induced in rats by intrabladder inoculation of Escherichia coli. The infected left kidney showed a significant bacterial proliferation, local production of interleukin (IL)-6 and IL-8 as detected by immunocytochemistry, and extensive destruction of renal parenchyma associated with impressive leukocyte recruitment. Inducible and constitutive nitric oxide synthases (NOS) were locally expressed, and a time-dependent increase in urinary secretion of nitric oxide (NO) was seen that could be blocked by NG-monomethyl-L-arginine. However, there was a discrepancy between the NO profile in the kidney and urine. The results demonstrate that in the early stage of acute pyelonephritis kidney tubules participate actively in the local host response by producing important inflammatory mediators and that urinary NO levels are not suitable for predicting renal NOS activity.

Topics: Acute Disease; Animals; Colony Count, Microbial; Disease Models, Animal; Escherichia coli; Escherichia coli Infections; Female; Immunohistochemistry; Inflammation Mediators; Interleukin-6; Interleukin-8; Kidney; Macrophages; Monocytes; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Pyelonephritis; Rats; Rats, Sprague-Dawley

The interactions between leukocytes and cytokines during the acute response to intramammary infections in the dry mammary gland of sheep were studied. Dry ewes were experimentally infected in one udder half with either Staphylococcus aureus or Escherichia coli, or infused with saline as control. Udder secretion samples, blood samples and udder tissue samples were collected before and 4, 8 and 24 h after infections/infusions. Total and differential leukocyte counts were calculated in both blood and mammary secretions, and flow cytometry was used to detect the presence of CD4+, CD8+, WC1+, IL-2R+, CD18+ or L-selectin + lymphocytes, CD18+ or L-selectin + neutrophils, and CD14+ leukocytes. Moreover, the concentrations of interleukin-1 beta (IL-1 beta), interleukin-8 (IL-8) and granulocyte-macrophage colony stimulating factor (GM-CSF) in udder secretions were measured using ELISA, and RT-PCR was used to detect the presence of corresponding cytokine mRNA in udder tissue biopsies. The results suggest an association between the concentrations of IL-1 beta, IL-8 and the intensity of neutrophil infiltration of the infected gland. Immunologically relevant changes in proportions of lymphocyte subpopulations might also occur in the acute phase of the inflammatory reaction of the udder. Greater cellular and cytokine responses to E. coli infection may have contributed to the milder clinical picture and more rapid resolution of infection than that seen for S. aureus. Enhancing the production of pro-inflammatory cytokines may improve defence against bacterial mastitis.

Topics: Animals; Biopsy; DNA; Enzyme-Linked Immunosorbent Assay; Escherichia coli; Escherichia coli Infections; Female; Flow Cytometry; Granulocyte-Macrophage Colony-Stimulating Factor; Interleukin-1; Interleukin-8; Leukocyte Count; Mammary Glands, Animal; Mastitis; Neutrophils; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sheep; Sheep Diseases; Staphylococcal Infections; Staphylococcus aureus

Enteroaggregative E. coli (EAggEC) are emerging as an important cause of persistent diarrhea, especially in children in the developing world, yet the pathogenesis of EAggEC infection is poorly understood. In an ongoing prospective study of childhood diarrhea in an urban Brazilian slum, EAggEC are the leading cause of persistent diarrhea. Children from this study with EAggEC and persistent diarrhea had significant elevations in fecal lactoferrin, interleukin (IL)-8, and IL-1beta. Moreover, children with EAggEC without diarrhea had elevated fecal lactoferrin and IL-1beta concentrations. The children with EAggEC in their stool had significant growth impairment after their positive culture, regardless of the presence or absence of diarrhea. Finally, 2 EAggEC strains were shown to cause IL-8 release from Caco-2 cells, apparently via a novel heat-stable, high-molecular-weight protein. These findings suggest that EAggEC may contribute to childhood malnutrition, trigger intestinal inflammation in vivo, and induce IL-8 secretion in vitro.

Topics: Brazil; Caco-2 Cells; Case-Control Studies; Cells, Cultured; Child, Preschool; Diarrhea; Escherichia coli; Escherichia coli Infections; Feces; Growth Disorders; Humans; Infant; Infant, Newborn; Inflammation; Interleukin-1; Interleukin-8; Intestines; Lactoferrin; Polymerase Chain Reaction; Prospective Studies; RNA, Messenger

Antitissue factor antibody attenuated the coagulopathic and lethal responses to LD100 Escherichia coli, whereas active site inhibited factor Xa inhibited only the coagulopathic response. In this study, we wished to determine: (1) whether active site inhibited factor VIIa blocks the coagulopathic and/or attenuates the lethal effects of LD100 E coli and (2) whether these effects are accompanied by attenuation of the inflammatory cytokine response to LD100 E coli. Eight baboons infused for 2 hours with LD100 E coli also were given five bolus infusions of DEGR VIIa of 280 microg/kg at T = -10 minutes, +2, 4, 6, and 8 hours and observed for changes in vital signs, and the concentrations of hemostatic components (fibrinogen, platelets, fibrin degradation products) and inflammatory mediators (tumor necrosis factor [TNF], interleukin-6 [IL-6], IL-8) at T = 0, 1, 2, 4, 6, and 8 hours. Eight control baboons were also infused with LD100 E coli alone and followed as described above. Four of the eight baboons treated with DEGR VIIa were permanent 7-day survivors versus none in the control group. The mean survival times for the treated and control groups were 116 +/- 22 and 26 +/- 8 hours, respectively. These values differed significantly from each other, (P = .0008). The decrease in platelet and fibrinogen concentrations and the increase in fibrin degradation products observed in the control group were significantly attenuated in the treated group, as was thrombosis of renal glomerular capillaries. Treatment with DEGR VIIa showed no effect on the peak TNF response to LD100 E coli at T = 2 hours (170 +/- 32 v 120 +/- 35 ng/mL). DEGR VIIa, however, did attenuate the IL-6 and IL-8 responses at T = 8 hours (ie, the IL-6 concentrations were 81 +/- 10 for treated and 1,256 +/- 236 for the control groups and the IL-8 concentrations were 28 +/- 3.9 for the treated and 60 +/- 8.2 for the control group). These values for IL-6 and IL-8 differed significantly from each other between the treated and control groups (P = .0001 and .0074, respectively). It should be noted that the initial responses of IL-6 and IL-8 up to T = 4 hours were not attenuated. We concluded that DEGR VIIa treatment attenuates inflammatory, as well as hemostatic system responses to LD100 E coli. We hypothesize that this occurs through interference with the assembly and/or interactions of tissue factor/VIIa complexes.

Topics: Animals; Binding Sites; Blood Coagulation; Capillaries; Escherichia coli Infections; Factor VIIa; Fibrin Fibrinogen Degradation Products; Fibrinogen; Humans; Interleukin-6; Interleukin-8; Interleukins; Kidney Glomerulus; Kinetics; Papio; Platelet Count; Thrombosis; Tumor Necrosis Factor-alpha

Radiolabelled interleukin-8 (IL-8) is a promising agent for the imaging of infection and inflammation. Several experiments were performed to explore further the imaging potential of radiolabelled IL-8. IL-8 was radioiodinated via the Bolton-Hunter method. Rabbits with focal infection (Escherichia coli, Staphylococcus aureus) or sterile inflammation (zymosan) were injected intravenously with 18.5 MBq (0.5 mCi) of 123I-IL-8. In separate studies, rabbits were injected intravenously with 111In-granulocytes with or without 125I-IL-8. Gamma camera images were obtained at 5 min, 1, 4 and 8 h post-injection (p.i.). Biodistribution was determined at 8 h p.i. In all models, the biodistribution of 123I-IL-8 was characterized by rapid blood clearance and high uptake in infection and sterile inflammation. All foci could be clearly visualized within 4 h p.i. Ex vivo abscess-to-contralateral muscle ratios increased to 114.7+/-23.0 (E. coli), 52.3+/-24.5 (S. aureus) and 49.8+/-8.3 (zymosan) at 8 h p.i. In the circulation, most 123I-IL-8 was bound to erythrocytes. The abscess uptake of 125I-IL-8 reached high levels despite reduced migration of granulocytes towards the site of infection due to the anti-inflammatory activity of intravenously injected IL-8. IL-8 could be injected without induction of neutropenia at a dosage of 2 ng kg(-1). In conclusion, the characteristics of radiolabelled IL-8 for imaging of infection and sterile inflammation are highly encouraging and warrant further optimization for clinical application.

Topics: Animals; Antigens, CD; Escherichia coli Infections; Female; Humans; Inflammation; Interleukin-8; Iodine Radioisotopes; Neutrophils; Rabbits; Radionuclide Imaging; Radiopharmaceuticals; Receptors, Interleukin; Receptors, Interleukin-8A; Recombinant Proteins; Staphylococcal Infections; Staphylococcus aureus; Tissue Distribution; Zymosan

The systemic inflammatory response to Escherichia coli O157 infection was studied from the profiles of circulating inflammatory and anti-inflammatory cytokines. Twelve patients transferred sequentially to our hospital for the intensive care with acute illness due to Escherichia coli O157 infection and the possible form of haemolytic uraemic syndrome were included in this study. Increased circulating concentrations of tumour necrosis factor, interleukin 6, interleukin 8, granulocyte colony-stimulating factor, and interleukin 10 were found in patients with various stages of this infection and haemolytic uraemic syndrome. Especially, the degree of the increase of circulating interleukin 10 in those who had a typical signs of haemolytic uraemic syndrome was higher than those of other inflammatory cytokines. Two groups of E. coli infection could be classified into one with a typical haemolytic uraemic syndrome and the other with atypically bacteremic state over haemolytic uraemic syndrome according to these cytokine levels.

Topics: Adult; Aged; Aged, 80 and over; Anti-Inflammatory Agents; Child; Cytokines; Escherichia coli Infections; Escherichia coli O157; Female; Granulocyte Colony-Stimulating Factor; Hemolytic-Uremic Syndrome; Humans; Interleukin-10; Interleukin-6; Interleukin-8; Male; Middle Aged; Tumor Necrosis Factor-alpha

The activation of complement and the release of TNF-alpha, IL-6 and IL-8 are important pathogenic factors behind organ dysfunction in sepsis. The aim of this study was to determine whether infusion of anti-TNF antibodies alters complement activation and plasma concentrations of pro-inflammatory cytokines at high doses of Escherichia coli. Six baboons received intravenously 2 x 10(9) live E. coli bacteria per kg body weight (group 1), in addition five received pretreatment with 1 mg per kg body weight anti-TNF antibodies (group 2), and seven received 5 x 10(8) live E. coli bacteria per kg body weight (group 3). Two hours after the start of infusion of the bacteria, plasma concentrations of C3 activation products, C5a and the terminal SC5b-9 complement complex were increased in groups 1 and 2 (P < 0.05), but there was no significant difference between the groups. At 2 h the levels of TNF-alpha, IL-6 and IL-8 were lower in group 2 compared with group 1 (P<0.05). In group 2 compared with group 1 the TNF-alpha concentrations were, however, higher at 4, 8 and 24 h. The explanation for this phenomenon is probably that TNF-alpha binds to the anti-TNF antibody complex and is released slowly after it has been bound. The study showed that infusion of anti-TNF antibodies reduced the concentrations of TNF-alpha, IL-6 and IL-8, without any detectable influence on complement activation.

Topics: Animals; Antibodies, Monoclonal; Complement Activation; Complement C3; Complement C5a; Disease Models, Animal; Escherichia coli Infections; Immunoglobulins, Intravenous; Interleukin-6; Interleukin-8; Papio; Sepsis; Tumor Necrosis Factor-alpha

Generation of inflammatory mediators and leukocyte recruitment to infection at an epithelial surface were studied during Escherichia coli-induced mastitis. One uninfected gland of each of eight midlactation cows was challenged with only 30 CFU of E. coli McDonald strain 487, a serum-resistant isolate from a cow with mastitis. Bacteria grew logarithmically during the first 10 to 12 h after challenge, reaching concentrations of more than 10(5) CFU/ml with no detectable host response during this time. An intense inflammatory reaction began approximately 12 h after the challenge and was characterized by a breakdown in the blood-milk permeability barrier followed by pyrexia and a pronounced leukocytic influx. Coincident with the onset of mammary inflammation was the appearance of neutrophil chemotactic activity in the milk from infected glands. Factors able to upregulate CD18 expression on peripheral blood neutrophils also appeared in milk at this time. The lack of appearance of chemotactic and CD18-upregulating activities until 12 h after challenge indicated that delays in neutrophil recruitment resulted from an initial lack of bacterial recognition and inflammatory mediator production. Production of complement fragment C5a, tumor necrosis factor, and interleukin-1 (IL-1) occurred earlier than production of IL-6 or IL-8. The early and intense production of C5a indicates that this chemoattractant may be more important than IL-8 during the initial recruitment and activation of neutrophils to a developing E. coli infection.

Topics: Animals; Cattle; Chemotaxis, Leukocyte; Complement C5a; Cytokines; Escherichia coli Infections; Female; Inflammation Mediators; Interleukin-8; Mammary Glands, Animal; Neutrophils; Tumor Necrosis Factor-alpha

Despite the fact that Helicobacter pylori is known to be non-invasive, mucosal infiltration of inflammatory cells have been observed in the gastric mucosa. The exact pathogenesis of such an inflammatory reaction has not been well defined. We explored the repertoire of cytokine genes expressed in human gastric epithelial cells in response to coculture with H. pylori. After gastric epithelial cells, SNU-5 and KATO III, were infected with H. pylori, expression of several cytokine genes was assessed using quantitative reverse transcription polymerase chain reaction. Interleukin (IL)-8, -1 alpha and -1 beta mRNA were expressed in both gastric epithelial cells throughout the entire infection period. In SNU-5, IL-1 alpha and IL-8 mRNA were expressed at 1 h, reached a peak level at 4 h and then decreased. Interleukin-1 beta mRNA was expressed less frequently than IL-1 alpha, or IL-8 mRNA. In SNU-5 cells, granulocyte-macrophage colony-stimulating factor (GM-CSF), monocyte chemoattractant protein-1 (MCP-1), and tumour necrosis factor-alpha (TNF-alpha) mRNA were expressed at 9 h, but was not expressed in KATO III. Gene expression paralleled the amount of IL-8 protein measured by enzyme-linked immunoabsorbent assay (ELISA). Interleukin-8 mRNA expression was not observed in KATO III cells infected with Campylobacter fetus ssp. fetus, Campylobacter jejuni or Escherichia coli. IL-8 mRNA expression was increased not only in gastric epithelial cells but also in non-gastric cells infected with H. pylori. These results suggest that an inflammatory reaction induced by H. pylori may be initially triggered by an array of pro-inflammatory cytokines expressed by infected gastric epithelial cells.

Topics: Campylobacter Infections; Cells, Cultured; Chemokine CCL2; Coculture Techniques; Cytokines; Epithelium; Escherichia coli Infections; Granulocyte-Macrophage Colony-Stimulating Factor; Helicobacter Infections; Helicobacter pylori; Humans; Interleukin-1; Interleukin-8; Polymerase Chain Reaction; RNA, Messenger; Stomach; Time Factors; Tumor Necrosis Factor-alpha

Interleukin (IL)-12 is thought to be a key factor for the induction of interferon gamma (IFN-gamma), a cytokine essential for the lethal effects of endotoxin. We report here on the release of the nonfunctional subunit of IL-12, p40, as well as biologically active heterodimeric IL-12, p70, after administration of a lethal (n = 5) or sublethal (n = 8) dose of live Escherichia coli to baboons. Remarkably, on lethal challenge, peak levels of p40 were observed at 3 hours that were about twofold lower than those elicited after sublethal challenge (2,813 +/- 515 pg/mL v 4,972 +/- 732 pg/mL, P < .05). This disparity was also observed, although to a lesser extent, for IL-12 p70 antigen, of which maximum levels of 91 +/- 47 pg/mL and 151 +/- 41 pg/mL were measured 6 hours after a lethal or sublethal dose of E coli, respectively. Circulating p70 antigen correlated with IL-12 biologic activity (r = 0.869; P < .001). When comparing lethal to sublethal conditions, lower peak levels of IL-12 on lethal E coli sharply contrasted with higher levels of other proinflammatory cytokines, such as tumor necrosis factor (TNF)-alpha, IL-1beta, IL-6, and IL-8 observed in these animals. Lower IL-12 concentrations in the lethal group may have resulted in part from the enhanced production of IL-10, a known inhibitor of IL-12 synthesis in vitro, as peak levels of this cytokine 3 hours postchallenge inversely correlated with peak levels of IL-12, in particular p40 (r = -0.802; P < .01). Contrary to what might be expected if IFN-gamma were solely induced by IL-12, lethally challenged baboons generated threefold more IFN-gamma at 6 hours than those receiving a sublethal dose (P < .05). Moreover, higher levels of IFN-gamma were associated with lower p40/p70 ratios, suggesting that, in agreement with observations in vitro, IFN-gamma may have preferentially upregulated the release of p70 over p40. These data show that IL-12 is released in experimental septic shock in nonhuman primates and suggest that IL-10 and IFN-gamma are involved in the regulation of this release. Furthermore, this study indicates that the systemic release of IL-12 might be essential, but is not likely sufficient, to promote lethal production of IFN-gamma in sepsis.

Topics: Animals; Bacteremia; Cytokines; Escherichia coli Infections; Gene Expression Regulation; Interferon-gamma; Interleukin-10; Interleukin-12; Interleukin-8; Killer Cells, Natural; Papio; Phagocytes; Shock, Septic; T-Lymphocytes; Tumor Necrosis Factor-alpha

As collections of lower respiratory tract specimens from young children with cystic fibrosis (CF) are difficult, we determined whether oropharyngeal cultures predicted lower airway pathogens. During 1992-1994, 75 of 90 (83%) infants with CF diagnosed by neonatal screening had 150 simultaneous bronchoalveolar lavage (BAL) and oropharyngeal specimens collected for quantitative bacterial culture at a mean age of 17 months (range, 1-52). Ten children undergoing bronchoscopy for stridor served as controls. Total and differential cell counts and interleukin-8 concentrations were measured in BAL fluid. A subset of bacterial pathogens were typed by pulsed field gel electrophoresis. A non-linear relationship with inflammatory markers supported a diagnosis of lower airway infection when > or = 10(5) colony-forming units/ml were detected. This criterion was met in 47 (31%) BAL cultures from 37 (49%) children. Staphylococcus aureus (19%), Pseudomonas aeruginosa (11%), and Hemophilus influenzae (8%) were the major lower airway pathogens. In oropharyngeal cultures, S. aureus (47%), Escherichia coli (23%), H. influenzae (15%), and P. aeruginosa (13%) predominated. The sensitivity, specificity, and positive and negative predictive values of oropharyngeal cultures for pathogens causing lower respiratory infections were 82%, 83%, 41%, and 97%, respectively. When there was agreement between paired oropharyngeal and BAL cultures, genetic fingerprinting showed some strains of the same organism were unrelated. We conclude that oropharyngeal cultures do not reliably predict the presence of bacterial pathogens in the lower airways of young CF children.

Topics: Bacteriological Techniques; Bronchoalveolar Lavage Fluid; Child, Preschool; Cystic Fibrosis; Escherichia coli Infections; Female; Haemophilus Infections; Haemophilus influenzae; Humans; Infant; Interleukin-8; Male; Oropharynx; Pneumonia, Bacterial; Predictive Value of Tests; Pseudomonas Infections; Respiratory Tract Infections; Staphylococcal Infections

Sepsis and septic shock lead to activation of the complement cascade and to the release of pro-inflammatory cytokines.. The effects of E coli infusion and of infusion of anti-TNF antibodies and a xanthine derivative (HWA 138) on complement activation and cytokine release was evaluated in 17 baboons. All animals received 5 x 10(8) live bacteria per kg body weight. Five animals received only bacteria, five received in addition 0.5 mg per kg body weight of anti-TNF-antibody, and seven received an infusion of 6 mg per kg body weight of HWA 138 in addition to the bacteria.. In baboons receiving 5 x 10(8) live E coli per kg body weight increased plasma levels of TCC, TNF-alpha and IL-8 were found. The release of TNF-alpha was lower in the group receiving HWA 138 at 2 h after the infusion. In baboons receiving an infusion of anti-TNF antibody the concentration of IL-8 was lower at 2 and 4 h than in animals receiving just E coli or HWA 138.. Infusion of anti-TNF-antibody before E coli infusion will decrease the formation of IL-8. Infusion of HWA 138 before the E coli infusion will also inhibit the formation of TNF-alpha.

Topics: Animals; Antibodies; Complement Activation; Complement Membrane Attack Complex; Escherichia coli Infections; Hemodynamics; Interleukin-8; Male; Papio; Pentoxifylline; Sepsis; Tumor Necrosis Factor-alpha; Xanthines

To define causes of increased susceptibility to coliform mastitis after parturition.. 12 healthy Holstein cows assigned to 2 groups. Group-1 cows (n = 6) had calved between 6 and 10 days earlier. Group-2 cows (n = 6) were in midlactation.. Cows from each group were paired and challenge exposed with Escherichia coli in 1 mammary gland. Mastitis severity was determined by bacterial concentration in milk, pyrexia, and milk production. Measures of host defense were neutrophil chemotaxis, adhesion molecule expression, leukocyte recruitment, and cytokine production.. After challenge exposure, group-1 cows had more rapid E coli growth, higher peak bacterial concentration, and higher fever. Leukocyte recruitment was poor in 1 group-1 cow that had peracute mastitis. In contrast, leukocyte recruitment in 5 other group-1 cows began sooner than that in group-2 cows. In these group-1 cows, prechallenge-exposure milk somatic cell counts (SCC) were significantly lower than those in group-2 cows. Prechallenge-exposure SCC were correlated to stimulated CD18 expression (R2 = 0.79), and both measures correlated inversely with bacterial growth rate (R2 = -0.75). Values for tumor necrosis factor alpha, interleukin 1, and interleukin 8 in group-1 cows after challenge exposure were greater than or equal to those in group-2 cows.. Weak leukocyte recruitment to the mammary gland is associated with increased severity of coliform mastitis. Impaired production of cytokines measured is not a cause of increased susceptibility to coliform mastitis in early lactation.. Low milk SCC after calving may increase susceptibility to severe coliform mastitis.

Topics: Animals; Cattle; Cytokines; Escherichia coli Infections; Female; Interleukin-1; Interleukin-8; Labor, Obstetric; Lactation; Mastitis, Bovine; Milk; Neutrophils; Pregnancy; Tumor Necrosis Factor-alpha

Tumor necrosis factor (TNF) and interleukin (IL)-1 are two cytokines for which naturally occurring inhibitors have been identified. The present study was undertaken to evaluate the extent to which scavenging of TNF in bacteremia attenuates the plasma levels of IL-1 receptor antagonist (IL-1ra) and soluble TNF receptors (sTNFR). Ten male baboons received 2 x 10(9) colony-forming units/kg live Escherichia coli over 2 h and were subjected to either placebo or anti-TNF antibody (anti-TNF Ab) treatment (1 mg/kg CDP571, Celltech, UK) 2 h before E. coli infusion (observation time: 72h). IL-1ra (range: 50-100 ng/ml) and sTNFR (range: 55kDa, 20-25 ng/ml; 75 kDa, 30-35 ng/ml) release was more sustained than that of IL-1 and TNF and was significantly attenuated by anti-TNF treatment, as were the circulating levels of IL-1, IL-8, and monocyte chemotactic peptide-1 (MCP-1) in the anti-TNF Ab group. We conclude that the increase in circulating natural cytokine modulators observed in nonhuman primate bacteremia is under the partial control of endogenous TNF because it was influenced by anti-TNF pretreatment. This attenuation is comparable to the anti-TNF effect on the chemokine MCP-1.

Topics: Animals; Antibodies, Monoclonal; Bacteremia; Chemokine CCL2; Escherichia coli Infections; Interleukin-1; Interleukin-8; Male; Papio; Receptors, Interleukin-1; Receptors, Tumor Necrosis Factor; Solubility; Tumor Necrosis Factor-alpha

Different immunologic parameters were measured in cord blood to test their usefulness in the early diagnosis of early onset sepsis. Cord blood levels of circulating intercellular adhesion molecule-1 (cICAM-1), interleukin-6 (IL-6) and interleukin-8 (IL-8) were significantly elevated in septic compared to nonseptic neonates. No significant difference between either population was seen for cord blood C3a and elastase-alpha 1-proteinase inhibitor complex (E alpha 1 PI). Measured concentrations of cICAM-1, IL-6 and IL-8 in fetal and maternal blood did not correlate, indicating that the neonate's response to sepsis is clearly different from the mother. Our data suggest that cord blood measurements of cICAM-1, IL-6 and IL-8 might be useful in identifying neonates with early-onset sepsis.

Topics: Escherichia coli Infections; Fetal Blood; Haemophilus Infections; Haemophilus influenzae; Humans; Infant, Newborn; Intercellular Adhesion Molecule-1; Interleukin-6; Interleukin-8; Sepsis

The chemokine monocyte chemotactic protein 1 (MCP-1) is a cytokine with chemotactic activity specific for mononuclear phagocytes. To investigate the possible involvement of MCP-1 in the pathogenesis of sepsis, its course was studied in baboons challenged intravenously with a sublethal or lethal dose of Escherichia coli. Levels of MCP-1 started to increase in both groups of animals 2 h after injection of E. coli, reaching peak levels 4 and 6 h after a sublethal (186 +/- 21 ng/mL) or a lethal (213 +/- 24 ng/mL) dose, respectively. Levels of MCP-1 correlated significantly with plasma levels of another chemokine, interleukin-8 (IL-8; r = .826. P < .001), suggesting that common stimuli mediate the release of both cytokines in this model.

Topics: Animals; Bacteremia; Chemokine CCL2; Chemotactic Factors; Cytokines; Escherichia coli Infections; Interleukin-8; Papio; Time Factors

Urine and serum concentrations of interleukin (IL)-6 and IL-8 were determined in 43 women with acute pyelonephritis caused by Escherichia coli. Urine and serum samples were also collected 2 weeks after the infection and during a subsequent episode of cystitis (n = 8) or asymptomatic bacteriuria (n = 8). Concentrations of IL-6 and IL-8 were related to the expression of 5 virulence markers of E. coli and glomerular filtration rate (GFR) after pyelonephritis. Patients with acute pyelonephritis had elevated urine and serum IL-6 and IL-8 levels as compared to 37 healthy women (IL-6: p < 0.001 in both cases, and IL-8: p < 0.001 in both cases). Patients infected with E. coli producing hemolysin and/or cytotoxic necrotizing factor (CNF) had significantly higher IL-6 levels in serum during acute pyelonephritis as compared to patients infected with strains without the ability to produce these factors (p = 0.0025 and p = 0.0154, respectively). Patients who had high concentrations of IL-8 in urine during acute pyelonephritis had lower GFR at follow-up as compared to patients with lower levels of IL-8 in urine (r = -0.48, p = 0.0123). In conclusion, acute pyelonephritis is accompanied by elevated urinary and serum IL-6 and IL-8 levels. Bacteria producing hemolysin and CNF seem to induce higher concentrations of IL-6 in serum. The secretion of IL-8 from renal cells may participate in the initiation and maintenance of renal inflammation which in turn may influence renal function.

Topics: Acute Disease; Adolescent; Adult; Bacterial Toxins; C-Reactive Protein; Cytotoxins; Escherichia coli; Escherichia coli Infections; Escherichia coli Proteins; Female; Fimbriae, Bacterial; Glomerular Filtration Rate; Hemolysin Proteins; Humans; Interleukin-6; Interleukin-8; Kidney; Middle Aged; Pyelonephritis; Virulence

1. Uropathogenic E. coli adhere to mucosal sites. 2. In the urinary tract, adherence is followed by inflammation, including a mucosal cytokine response. 3. Bacteria activate epithelial cells to secrete IL-6 and IL-8. IL-6 may cause the fever and acute phase response that accompany systemic urinary tract infections. IL-8 may function as a neutrophil chemoattractant. 4. E. coli up-regulate adhesion molecule expression on epithelial cell lines and neutrophil migration through epithelial cell monolayers. This process is inhibited by antibodies to CD18 and ICAM-1. 5. Cytokines released by nonepithelial cells (T cells and monocytes) modify the epithelial cell cytokine response to bacteria.

Topics: Bacterial Adhesion; Carbohydrate Sequence; Cytokines; Escherichia coli; Escherichia coli Infections; Fimbriae, Bacterial; Glycolipids; Humans; Interleukin-6; Interleukin-8; Molecular Sequence Data; Neutrophils; T-Lymphocytes; Urinary Tract Infections

Urinary tract infections activate a mucosal inflammatory response, which includes cytokine secretion and neutrophil influx. The mechanisms involved in the neutrophil influx have not been identified. Interleukin-8, a potent chemoattractant for neutrophils, is produced by urinary tract epithelial cell lines in vitro. This study analyzed the human IL-8 response to deliberate Escherichia coli infection of the urinary tract. Urine and serum samples were obtained before and after intravesical instillation of E. coli. Neutrophil numbers were determined on uncentrifuged urine, and IL-8 levels were measured by ELISA. A urinary IL-8 response was found in all patients after bacterial instillation, but no serum IL-8 was detected. There was a strong correlation between urinary IL-8 levels and urinary neutrophil numbers. The same E. coli strains used to colonize the patients stimulated IL-8 production in urinary tract epithelial cells. The level of IL-8 secreted by epithelial cell lines was influenced by the fimbrial properties of the E. coli. These results demonstrated that E. coli elicit a mucosal IL-8 response in humans, and suggested that IL-8 is involved in the onset of pyuria. Epithelial cells may be an important source of IL-8 during urinary tract infection.

Topics: Cell Line; Enzyme-Linked Immunosorbent Assay; Epithelium; Escherichia coli Infections; Female; Humans; Immunohistochemistry; Interleukin-8; Neutrophils; Urinary Tract; Urinary Tract Infections

The role of tumor necrosis factor (TNF) in interleukin (IL)-8 release in septicemia in the baboon (2-h infusion of live Escherichia coli, 5 x 10(8) cfu/kg) was investigated. Four experiments were done: one control (n = 7) and three with pretreatment to reduce TNF plasma levels. Pretreatment was with anti-TNF antibody (anti-TNF) 15 mg/kg, which neutralized circulating TNF (n = 4); 0.5 mg/kg of anti-TNF, which reduced peak TNF from 6.2 ng/mL (controls) to 0.6 ng/mL (n = 4); and a xanthine derivate (HWA138), which reduced TNF to 1 ng/mL (n = 5). With TNF levels < 1 ng/mL, a significant reduction of circulating IL-8 from 10.4 ng/mL (peak) in controls to 1 ng/mL (peak) in anti-TNF-treated animals was found, but with HWA138 only some decrease in IL-8 was seen. Despite high endotoxin levels (10-30 ng/mL peak), neutralization of TNF resulted in diminished release of IL-8 and significantly lower levels of granulocyte elastase.

Topics: Animals; Escherichia coli Infections; Interleukin-8; Male; Papio; Sepsis; Tumor Necrosis Factor-alpha

A hyperdynamic sepsis model was set up in seven adult baboons to evaluate neutrophil-activating peptide-1/interleukin (IL)-8 (NAP-1/IL-8), IL-1 beta, IL-6, tumor necrosis factor-alpha (TNF alpha), and IFN-gamma in plasma. By continuous intravenous administration of 10(10) cfu/kg live Escherichia coli over 8 h with additional infusion therapy (less than or equal to 50 ml/kg/h), endotoxin plasma levels of 2.7-22.3 ng/ml were observed. In plasma the kinetics of NAP-1/IL-8 and IL-6 were similar to those of IL-1 at the end of the experiment (8 h) (peak median values, 34, 4197, and 230 ng/ml, respectively). Differences were greatest for IL-6. Monocyte activation during sepsis was confirmed by elevated plasma neopterin levels (91-139 mumol/mmol of creatine). Granulocyte activation was evident from both incipient neutropenia and the massive release of neutrophil elastase into the plasma as measured by a new immunoassay (peak level, 374 ng/ml). Thus, in primate bacteremia, early TNF release is followed by a concomitant increase of NAP-1/IL-8 with plasma kinetics similar to those of IL-6 and IL-1 and accompanied by massive activation of neutrophils.

Topics: Animals; Biopterins; Disease Models, Animal; Endotoxins; Escherichia coli Infections; Interferon-gamma; Interleukin-1; Interleukin-6; Interleukin-8; Leukocyte Elastase; Male; Neopterin; Pancreatic Elastase; Papio; Sepsis; Tumor Necrosis Factor-alpha

Topics: Animals; Biopterins; Endotoxins; Escherichia coli Infections; Humans; In Vitro Techniques; Infant, Newborn; Interleukin-1; Interleukin-6; Interleukin-8; Kidney; Leukocytes; Lipopolysaccharides; Lung; Male; Models, Biological; Neopterin; Pancreatic Elastase; Papio; Sepsis; Tumor Necrosis Factor-alpha