interleukin-8 and Klebsiella-Infections

Outer membrane vesicles (OMVs) derived from bacteria are known to play a crucial role in the interactions between bacteria and their environment, as well as bacteria-bacteria and bacteria-host interactions.Specifically, OMVs derived from Klebsiella pneumoniae have been implicated in contributing to the pathogenesis of this bacterium.Hypervirulent Klebsiella pneumoniae (hvKp) has emerged as a global pathogen of great concern due to its heightened virulence compared to classical K. pneumoniae (cKp), and its ability to cause community-acquired infections, even in healthy individuals.The objective of this study was to investigate potential differences between hvKp-derived OMVs and cKp-derived OMVs in their interactions with microorganisms and host cells.. Four strains of K. pneumoniae were used to produce OMVs: hvKp strain NTUH-K2044 (K1, ST23), hvKp clinical strain AP8555, and two cKP clinical strains C19 and C250. To examine the morphology and size of the bacterial OMVs, transmission electron microscopy (TEM) was utilized. Additionally, dynamic light scattering (DLS) was used to analyze the size characterization of the OMVs.The normal pulmonary bronchial cell line HBE was exposed to OMVs derived from hvKp and cKP. Interleukin 8 (IL-8) messenger RNA (mRNA) expression was assessed using reverse transcription-polymerase chain reaction (RT-PCR), while IL-8 secretion was analyzed using enzyme-linked immunosorbent assay (ELISA).Furthermore, the activation of nuclear factor kappa B (NF-κB) was evaluated using both Western blotting and confocal microscopy.. After purification, OMVs appeared as electron-dense particles with a uniform spherical morphology when observed through TEM.DLS analysis indicated that hvKp-derived OMVs from K2044 and AP8555 measured an average size of 116.87 ± 4.95 nm and 96.23 ± 2.16 nm, respectively, while cKP-derived OMVs from C19 and C250 measured an average size of 297.67 ± 26.3 nm and 325 ± 6.06 nm, respectively. The average diameter of hvKp-derived OMVs was smaller than that of cKP-derived OMVs.A total vesicular protein amount of 47.35 mg, 41.90 mg, 16.44 mg, and 12.65 mg was generated by hvKp-K2044, hvKp-AP8555, cKP-C19, and cKP-C250, respectively, obtained from 750 mL of culture supernatant. Both hvKp-derived OMVs and cKP-derived OMVs induced similar expression levels of IL-8 mRNA and protein. However, IL-8 expression was reduced when cells were exposed to BAY11-7028, an inhibitor of the NF-κB pathway.Western blotting and confocal microscopy revealed increased phosphorylation of p65 in cells exposed to OMVs.. Klebsiella pneumoniae produces outer membrane vesicles (OMVs) that play a key role in microorganism-host interactions. HvKp, a hypervirulent strain of K. pneumoniae, generates more OMVs than cKP.The average size of OMVs derived from hvKp is smaller than that of cKP-derived OMVs.Despite these differences, both hvKp-derived and cKP-derived OMVs induce a similar level of expression of IL-8 mRNA and protein.OMVs secreted by K. pneumoniae stimulate the secretion of interleukin 8 by activating the nuclear factor NF-κB.

Topics: Anti-Bacterial Agents; Humans; Interleukin-8; Klebsiella Infections; Klebsiella pneumoniae; Lung; NF-kappa B; Virulence

The aim of this study was to prepare cefquinome-loaded polylactic acid microspheres and to evaluate their in vitro and in vivo characteristics and pharmacodynamics for the therapy of pneumonia in a rat model. Microspheres were prepared using a 0.7 mm two-fluid nozzle spray drier in one step resulting in spherical and smooth microspheres of uniform size (9.8 ± 3.6 μm). The encapsulation efficiency and drug loading of cefquinome were 91.6 ± 2.6% and 18.7 ± 1.2%, respectively. In vitro release of cefquinome from the microspheres was sustained for 36 h. Cefquinome-loaded polylactic acid microspheres as a drug delivery system was successful for clearing experimental Klebsiella pneumonia lung infections. A decrease in inflammatory cells and an inhibition of inflammatory cytokines TNF-α, IL-1β and IL-8 after microspheres treatment was found. Changes in cytokine levels and types are secondary manifestations of drug bactericidal effects. Rats were considered to be microbiologically cured because the bacterial load was less than 100 CFU/g. These results also indicated that the spray-drying method of loading therapeutic drug into polylactic acid microspheres is a straightforward and safe method for lung-targeting therapy in animals.

Topics: Animals; Anti-Bacterial Agents; Bacterial Load; Cephalosporins; Disease Models, Animal; Drug Carriers; Drug Compounding; Drug Liberation; Host-Pathogen Interactions; Inflammation Mediators; Interleukin-1beta; Interleukin-8; Klebsiella Infections; Klebsiella pneumoniae; Lung; Male; Microspheres; Particle Size; Pneumonia, Bacterial; Polyesters; Rats, Wistar; Surface Properties; Technology, Pharmaceutical; Time Factors; Tumor Necrosis Factor-alpha

Klebsiella pneumoniae (K. pneumoniae) is a hospital-acquired infectious agent that causes a range of diseases. Herein we have developed a novel CXCL8-IP10 hybrid protein and evaluated its efficacy in an animal model of K. pneumoniae infection. Neutrophil chemotaxis data revealed that CXCL8-IP10 could inhibit human neutrophil chemotactic responses induced by the ELR-CXC chemokine CXCL8. To evaluate the effect of CXCL8-IP10 on K. pneumoniae infection, C57BL/6 mice were challenged with K. pneumoniae followed by treatment with CXCL8-IP10 (500 μg/kg, i.p.), or dexamethasone (0.8 mg/kg, s.c.), or ceftazidime (200 mg/kg, s.c.) individually. CXCL8-IP10, dexamethasone or ceftazidime markedly inhibit Klebsiella-induced pulmonary inflammation as assessed by gross examination and histopathology. Moreover, the chemotactic responses of neutrophils was blocked by CXCL8-IP10 rather than dexamethasone or ceftazidime. Furthermore, the levels of inflammatory factors IL-1β, IFN-γ and TNF-α were decreased after CXCL8-IP10, dexamethasone or ceftazidime treatment. Together, these results suggest that CXCL8-IP10 may provide a novel strategy for treating K. pneumoniae infection.

Topics: Animals; Chemokine CXCL10; Chemotaxis, Leukocyte; Disease Models, Animal; Female; Interleukin-8; Klebsiella Infections; Klebsiella pneumoniae; Mice, Inbred C57BL; Neutrophils; Pneumonia, Bacterial; Recombinant Fusion Proteins

Klebsiella pneumoniae is a common cause of nosocomial pneumonia, especially in children. Toll-like receptors plays an important role in defense against this pathogen. The impact of human TLR6 polymorphisms on susceptibility to K. pneumoniae infection is poorly understood. The aim of the present work was to determine whether single nucleotide polymorphisms in TLR6 are associated with altered immune responses to K. pneumoniae.. The TLR6 coding region was sequenced in 126 K. pneumoniae culture-positive patients and 142 hospitalized K. pneumoniae culture-negative controls.. The frequency of V327M polymorphism was found to be significantly higher in patients than that in controls (16.7% vs. 7.7%). In vitro studies showed that V327M polymorphism did not impair TLR6 expression in transfected HEK 293T cells. Further studies demonstrated that V327M polymorphism was associated with increased IL-8 mRNA expression in transfected HEK 293T cells when stimulated with K. pneumoniae and the specific ligand for TLR2/TLR6 heterodimers known as Pam2CSK4. The present data showed V327M polymorphism to be associated with increased apoptosis of HEK 293T cells when challenged with K. pneumoniae.. Taken together, these data indicated that TLR6 V327M may be involved in mediating deleterious inflammatory responses and modulating host susceptibility to K. pneumoniae. These results provide new insight into the pathophysiologic role of TLR6 V327M in the innate immune response to bacterial infection in human.

Topics: Apoptosis; Case-Control Studies; Coculture Techniques; Genetic Predisposition to Disease; HEK293 Cells; Humans; Interleukin-8; Klebsiella Infections; Klebsiella pneumoniae; Lipopeptides; NF-kappa B; Polymorphism, Single Nucleotide; Risk Factors; RNA, Messenger; Signal Transduction; Toll-Like Receptor 2; Toll-Like Receptor 6; Transfection

Klebsiella pneumoniae-associated pathology is largely mediated by neutrophilic inflammation. In this study, we administered Klebsiella pneumoniae to experimental guinea pig groups and the ELR-CXC chemokine antagonist CXCL8(3-72), ceftazidime, and dexamethasone to different groups, respectively. After 24 h, we assessed the animal's pulmonary inflammatory levels, including gross histopathology, airway neutrophilia, lung myeloperoxidase levels, expressions of CXCL8 and TNF, and airway bacterial loads. Compared with ceftazidime and dexamethasone treatments, the administration of the ELR-CXC chemokine antagonist CXCL8(3-72) alone was more effective than other methods, although it did not markedly attenuate the bacterial load. These results suggest new methods for the treatment of Klebsiella pneumoniae pathology.

Topics: Animals; Ceftazidime; Dexamethasone; Guinea Pigs; Humans; Interleukin-8; Klebsiella Infections; Klebsiella pneumoniae; Lung; Pneumonia; Receptors, Interleukin-8A; Receptors, Interleukin-8B

Outer membrane protein A (OmpA) is a class of proteins highly conserved among the Enterobacteriaceae family and throughout evolution. Klebsiella pneumoniae is a capsulated gram-negative pathogen. It is an important cause of community-acquired and nosocomial pneumonia. Evidence indicates that K. pneumoniae infections are characterized by a lack of an early inflammatory response. Data from our laboratory indicate that K. pneumoniae CPS helps to suppress the host inflammatory response. However, it is unknown whether K. pneumoniae employs additional factors to modulate host inflammatory responses. Here, we report that K. pneumoniae OmpA is important for immune evasion in vitro and in vivo. Infection of A549 and normal human bronchial cells with 52OmpA2, an ompA mutant, increased the levels of IL-8. 52145-Δwca(K2)ompA, which does not express CPS and ompA, induced the highest levels of IL-8. Both mutants could be complemented. In vivo, 52OmpA2 induced higher levels of tnfα, kc, and il6 than the wild type. ompA mutants activated NF-κB, and the phosphorylation of p38, p44/42, and JNK MAPKs and IL-8 induction was via NF-κB-dependent and p38- and p44/42-dependent pathways. 52OmpA2 engaged TLR2 and -4 to activate NF-κB, whereas 52145-Δwca(K2)ompA activated not only TLR2 and TLR4 but also NOD1. Finally, we demonstrate that the ompA mutant is attenuated in the pneumonia mouse model. The results of this study indicate that K. pneumoniae OmpA contributes to attenuate airway cell responses. This may facilitate pathogen survival in the hostile environment of the lung.

Topics: Animals; Bacterial Outer Membrane Proteins; Female; HEK293 Cells; Humans; Immune Evasion; Inflammation; Interleukin-8; Klebsiella Infections; Klebsiella pneumoniae; Mice; Mitogen-Activated Protein Kinase Kinases; Mutation; NF-kappa B; Nod1 Signaling Adaptor Protein; Pneumonia, Bacterial; Respiratory Mucosa; Toll-Like Receptors

We previously demonstrated that exposure to febrile-range hyperthermia (FRH) accelerates pathogen clearance and increases survival in murine experimental Klebsiella pneumoniae peritonitis. However, FRH accelerates lethal lung injury in a mouse model of pulmonary oxygen toxicity, suggesting that the lung may be particularly susceptible to injurious effects of FRH. In the present study, we tested the hypothesis that, in contrast with the salutary effect of FRH in Gram-negative peritonitis, FRH would be detrimental in multilobar Gram-negative pneumonia. Using a conscious, temperature-clamped mouse model and intratracheal inoculation with K. pneumoniae Caroli strain, we showed that FRH tended to reduce survival despite reducing the 3 day-postinoculation pulmonary pathogen burden by 400-fold. We showed that antibiotic treatment rescued the euthermic mice, but did not reduce lethality in the FRH mice. Using an intratracheal bacterial endotoxin LPS challenge model, we found that the reduced survival in FRH-treated mice was accompanied by increased pulmonary vascular endothelial injury, enhanced pulmonary accumulation of neutrophils, increased levels of IL-1beta, MIP-2/CXCL213, GM-CSF, and KC/CXCL1 in the bronchoalveolar lavage fluid, and bronchiolar epithelial necrosis. These results suggest that FRH enhances innate host defense against infection, in part, by augmenting polymorphonuclear cell delivery to the site of infection. The ultimate effect of FRH is determined by the balance between accelerated pathogen clearance and collateral tissue injury, which is determined, in part, by the site of infection.

Topics: Animals; Cell Line; Epithelial Cells; Fever; Humans; Interleukin-1; Interleukin-8; Klebsiella Infections; Klebsiella pneumoniae; Lipopolysaccharides; Lung; Lung Injury; Male; Mice; Neutrophils; Pneumonia, Bacterial; Recombinant Proteins; Tumor Necrosis Factor-alpha

Gram-negative bacteria are responsible for almost one-half of the clinical cases of mastitis that occur annually. Of those gram-negative bacteria that induce mastitis, Klebsiella pneumoniae remains one of the most prevalent. Detection of infectious pathogens and the induction of a proinflammatory response are critical components of host innate immunity. The objective of the current study was to characterize several elements of the bovine innate immune response to intramammary infection with Klebsiella pneumoniae. The inflammatory cytokine response and changes in the levels of soluble CD14 (sCD14) and lipopolysaccharide (LPS)-binding protein (LBP), 2 proteins that contribute to host recognition of gram-negative bacteria, were studied. The contralateral quarters of 7 late-lactating Holstein cows were challenged with either saline or K. pneumoniae, and milk and blood samples were collected. Initial increases in the chemoattractants C5a and IL-8, as well as TNF-alpha, were evident in infected quarters within 16 h of challenge and were temporally coincident with increases in milk somatic cells. Augmented levels of TNF-alpha and IL-8 were observed in infected quarters until >48 h postchallenge, respectively. Elevated levels of IL-12, IFN-gamma, and the antiinflammatory cytokine, IL-10, which were first detected between 12 and 20 h postinfection, persisted in infected quarters throughout the study (>96 h). Initial increases in milk LBP and sCD14 were detected 16 and 20 h, respectively, after challenge. Together, these data demonstrate that intramammary infection with K. pneumoniae elicits a host response characterized by the induction of proinflammatory cytokines and elevation of accessory molecules involved in LPS recognition.

Topics: Acute-Phase Proteins; Animals; Carrier Proteins; Cattle; Female; Interferon-gamma; Interleukin-10; Interleukin-12; Interleukin-8; Kinetics; Klebsiella Infections; Klebsiella pneumoniae; Lipopolysaccharide Receptors; Mastitis, Bovine; Membrane Glycoproteins; Milk; Solubility; Tumor Necrosis Factor-alpha

CXCL5 (epithelial-cell-derived neutrophil-activating protein (ENA-)78) is a CXC-chemokine that specifically acts on neutrophils. To obtain insight into the extent of local presence and action of CXCL5 during bacterial meningitis, we measured its concentrations in cerebrospinal fluid (CSF) of patients with culture-proven bacterial meningitis (n=14), aseptic meningitis (n=6), and controls (n=32) and compared these results with levels of other CXC-chemokines, CXCL8- (interleukin-8) and CXCL1-related oncogene (growth-related oncogene (GRO)-alpha). Patients with bacterial meningitis had profoundly elevated CSF concentrations of all three chemokines. CXCL5 was not detectable in patients with aseptic meningitis or control subjects. CSF from patients with bacterial meningitis exerted chemotactic activity towards neutrophils, which was partially inhibited by neutralizing antibodies against CXCL5 and CXCL8, but not CXCL1. CSF from controls exerted minor chemotactic activity, which could be strongly enhanced by the addition of recombinant CXCL5, CXCL8 or CXCL1. During bacterial meningitis, CXCL5 is elevated in CSF, where it is involved in the recruitment of neutrophils to the central nervous system.

Topics: Adolescent; Chemokine CXCL1; Chemokine CXCL5; Chemokines, CXC; Chemotaxis, Leukocyte; Child; Humans; Intercellular Signaling Peptides and Proteins; Interleukin-8; Klebsiella Infections; Meningitis, Aseptic; Meningitis, Bacterial; Meningitis, Meningococcal; Meningitis, Pneumococcal; Neutrophil Activation

This study was designed to assess monoclonal antibodies (MAbs) directed against tumor necrosis factor-alpha (TNF-alpha) (anti-TNF) or interleukin-8 (anti-IL-8) as radioactive agents for the detection of Staphylococcus aureus-or Klebsiella pneumoniae-infected thighs in mice. At 5 min (acute infection) or 20 h (established) post-infection, 20 micrograms of the 99mTc-labeled MAbs were injected. At various time intervals, the accumulation of the radiotracer in the infected thighs was assessed and expressed as a target-to-nontarget (T/NT) ratio. The binding of 99mTc-labeled MAbs to circulating mononuclear cells and granulocytes was quantitated 20 h after injection. The pharmacokinetics of the MAbs, in relation to the control agents 99mTc-labeled polyclonal human immunoglobulin (IgG) and a 99mTc-labeled nonspecific IgG1 MAb, were also studied. In acute infections, 99mTc-anti-TNF accumulated to a higher extent (p < 0.05) in S. aureus-infected thighs in mice until 4 h after the injection than 99mTc-IgG and was higher at 0.25 h in K. pneumoniae-infected mice (p < 0.03) compared with 99mTc-IgG. In established S. aureus and K. pneumoniae infections, 99mTc-anti-IL-8 detected the infection more intensely than 99mTc-IgG until 1 h after injection. In both S. aureus and K. pneumoniae infections, localization of sites of infection correlates (p < 0.05) with increased binding of the 99mTc-labeled MAbs to granulocytes and mononuclear cells in both acute and established infections. It was concluded that 99mTc-labeled MAbs, directed against TNF-alpha and IL-8, accumulate in bacterial infections in mice to a higher extent than does 99mTc-IgG after infection and is related to the binding of the antibodies to blood leukocytes. With these 99mTc-labeled MAbs, information might be gained about the development of an infection.

Topics: Animals; Antibodies, Monoclonal; Humans; Immunoglobulin G; Interleukin-8; Isotope Labeling; Klebsiella Infections; Klebsiella pneumoniae; Leukocytes; Male; Mice; Muscle, Skeletal; Radionuclide Imaging; Staphylococcal Infections; Technetium; Tissue Distribution; Tumor Necrosis Factor-alpha

Topics: Animals; Blood Bactericidal Activity; CD18 Antigens; Chemokine CXCL2; Cytokines; Gene Transfer Techniques; Genetic Therapy; Humans; Interleukin-12; Interleukin-8; Klebsiella Infections; Klebsiella pneumoniae; Lung; Mice; Mice, Inbred Strains; Monokines; Neutrophils; Phagocytosis; Pneumonia, Bacterial

The effect of treatment with interleukin-8 (IL-8), a neutrophil-activating cytokine, was investigated in normal and neutropenic mice infected with a lethal dose of Pseudomonas aeruginosa, Klebsiella pneumoniae, or Plasmodium berghei. Intraperitoneal (i.p.) IL-8 treatment was associated with accelerated death when IL-8 was administered shortly before i.p. infection with P. aeruginosa or shortly after i.p. infection with P. aeruginosa and K. pneumoniae. Histopathological analyses demonstrated a tendency to more severe organ lesions in IL-8-treated mice. Only nonneutropenic mice that received IL-8 shortly before the infectious challenge and at the site of infection were protected by IL-8. Whether IL-8 is protective of or detrimental to the survival of infection appeared to depend on the presence of bacteria at the injection site and on the presence of neutropenia. IL-8 may be an important participant in the cascade of interacting cytokines that is induced by the lethal infectious challenge.

Topics: Animals; Bacterial Infections; Brain; Female; Interleukin-8; Klebsiella Infections; Klebsiella pneumoniae; Leukocyte Count; Malaria; Mice; Neutropenia; Peritoneal Cavity; Plasmodium berghei; Pseudomonas Infections