bluuebunny/crossref_metadata_2025_split huggingface dataset:
Essential Role of Transcription Factor Nuclear Factor-κB in Regulation of Interleukin-8 Gene Expression by Nitrite Reductase fromPseudomonas aeruginosain Respiratory Epithelial Cells
Authors: Mori Naoki , Oishi Kazunori , Sar Borann , Mukaida Naofumi , Nagatake Tsuyoshi , Matsushima Kouji , Yamamoto Naoki 
Published: December 2019

ABSTRACTPersistent infection withPseudomonas aeruginosaincreases interleukin-8 (IL-8) levels and causes dense neutrophil infiltrations in the airways of patients with chronic airway diseases. Recently, we have reported that nitrite reductase fromP. aeruginosainduces the production of IL-8 in respiratory cells, including bronchial epithelial cells. To determine the molecular mechanism(s) of nitrite reductase-induced IL-8 expression in respiratory cells, A549 epithelial cells were transfected with plasmids containing serial deletions of the 5′-flanking region of the IL-8 gene and then exposed to nitrite reductase. Nitrite reductase significantly enhanced IL-8 gene promoter-driven reporter activity. This increased IL-8 gene expression was inhibited by mutating the nuclear factor-κB (NF-κB) binding element. Nitrite reductase enhanced nuclear localization of the NF-κB binding complex. Furthermore, nitrite reductase induced the degradation of IκBα, the major cytoplasmic inhibitor of NF-κB, and the expression of IκBα mRNA. These data support the critical role of the activation of NF-κB in nitrite reductase-induced IL-8 gene expression in airway epithelium.
Characterization of a
Haemophilus ducreyi
Mutant Deficient in Expression of Cytolethal Distending Toxin
Authors: Stevens Marla K. , Latimer Jo L. , Lumbley Sheryl R. , Ward Christine K. , Cope Leslie D. , Lagergard Teresa , Hansen Eric J. 
Published: December 2019

ABSTRACT

Haemophilus ducreyi
expresses a soluble cytolethal distending toxin (CDT) that kills HeLa, HEp-2, and other human epithelial cells in vitro.
H. ducreyi
CDT activity is encoded by a three-gene cluster (
cdtABC
), and antibody to the
cdtC
gene product can neutralize CDT activity in vitro (L. D. Cope, S. R. Lumbley, J. L. Latimer, J. Klesney-Tait, M. K. Stevens, L. S. Johnson, M. Purven, R. S. Munson, Jr., T. Lagergard, J. D. Radolf, and E. J. Hansen, Proc. Natl. Acad. Sci. USA 94:4056–4061, 1997). Culture supernatant fluid from a recombinant
Escherichia coli
strain containing the
H. ducreyi cdtABC
gene cluster readily killed both HeLa cells and HaCaT keratinocytes and had a modest inhibitory effect on the growth of human foreskin fibroblasts. Insertional inactivation of the
cdtC
gene in this recombinant
E. coli
strain eliminated the ability of this strain to kill HeLa cells and HaCaT keratinocytes. This mutated
H. ducreyi cdtABC
gene cluster was used to construct an isogenic
H. ducreyi cdtC
mutant. Monoclonal antibodies against the
H. ducreyi
CdtA, CdtB, and CdtC proteins were used to characterize protein expression by this
cdtC
mutant. Culture supernatant fluid from this
H. ducreyi cdtC
mutant did not detectably affect any of the human cells used in this study. The presence of the wild-type
H. ducreyi cdtC
gene in
trans
in this
H. ducreyi
mutant restored its ability to express a CDT that killed both HeLa cells and HaCaT keratinocytes. The isogenic
H. ducreyi cdtC
mutant was shown to be as virulent as its wild-type parent strain in the temperature-dependent rabbit model for experimental chancroid. Lack of expression of the
H. ducreyi
CdtC protein also did not affect the ability of this
H. ducreyi
mutant to survive in the skin of rabbits.
Early Emergence of CD8+T Cells Primed for Production of Type 1 Cytokines in the Lungs ofMycobacterium tuberculosis-Infected Mice
Authors: Serbina Natalya V. , Flynn JoAnne L. 
Published: December 2019

ABSTRACTSeveral lines of evidence suggest that CD8 T cells are important in protection against tuberculosis. To understand the function of this cell population in the immune response againstMycobacterium tuberculosis, T cells from lungs ofM. tuberculosis-infected mice were examined by flow cytometry. The kinetics of the appearance of CD8 T cells in lungs of infected mice closely paralleled that of CD4 T cells. Both CD4+and CD8+T cells displaying an activated phenotype were found in the lungs as early as 1 week postinfection. By 2 weeks, total cell numbers in the lungs had tripled and percentages of T cells were increased two- to threefold; the percentages of CD4+T cells were ca. twofold higher than those of CD8+T cells. Short-term stimulation withM. tuberculosis-infected antigen-presenting cells induced cytokine production by primed CD4+and CD8+T cells. Intracellular cytokine staining revealed that 30% ± 5% of CD4+and 23% ± 4% of CD8+T cells were primed for production of gamma interferon (IFN-γ). However, a difference in in vivo IFN-γ production by T cells was observed with ∼12% of CD4+T cells and ∼5% of CD8+T cells secreting cytokine in the lungs at any given time during infection. The data presented indicate that although early in infection the majority of IFN-γ is produced by CD4+T cells, cytokine-producing CD8+T cells are readily available when triggered by the appropriate stimuli.
Interaction ofBrucella abortusLipopolysaccharide with Major Histocompatibility Complex Class II Molecules in B Lymphocytes
Authors: Forestier Claire , Moreno Edgardo , Méresse Stéphane , Phalipon Armelle , Olive Daniel , Sansonetti Philippe , Gorvel Jean-Pierre 
Published: December 2019

ABSTRACTLipopolysaccharide (LPS), a major amphiphilic molecule located at the outer membrane of gram-negative bacteria, is a potent antigen known to induce specific humoral immune responses in infected mammals. LPS has been described as a polyclonal activator of B lymphocytes, triggering the secretion of antibodies directed against distinct sugar epitopes of the LPS chain. But, how LPS is handled by B cells remains to be fully understood. This task appears to be essential for a better knowledge of the anti-LPS humoral immune response. In this study, we examine the internalization of LPS and its interaction with antigen-presenting major histocompatibility complex (MHC) class II molecules in murine and human B-cell lines. By use of immunofluorescence, we observe that structurally different LPSs fromBrucellaandShigellastrains accumulate in an intracellular compartment enriched in MHC class II molecules. By use of immunoprecipitation, we illustrate that onlyBrucella abortusLPS associates with MHC class II molecules in a haplotype-independent manner. Taken together, these results raise the possibility thatB. abortusLPS may play a role in T-cell activation.
Opacity-Associated Protein A Contributes to the Binding of
Haemophilus influenzae
to Chang Epithelial Cells
Authors: Prasadarao Nemani V. , Lysenko Elena , Wass Carol A. , Kim Kwang Sik , Weiser Jeffrey N. 
Published: December 2019

ABSTRACT

Opacity-associated protein A (OapA), which is responsible for the transparent-colony phenotype of
Haemophilus influenzae
, has been implicated in the colonization of the nasopharynx in an infant rat model of carriage. In this report, we show that OapA mediates attachment to Chang epithelial cells examined by using genetically defined type b and nontypeable
H. influenzae
strains with or without OapA. We also showed that OapA was conserved among
H. influenzae
strains by comparing deduced amino acid sequences. Both recombinant OapA and polyclonal anti-OapA antiserum blocked the binding of
H. influenzae
to Chang epithelial cells, suggesting that the interaction of
H. influenzae
is specific to OapA. Moreover, the binding of recombinant OapA to epithelial cells further provided evidence that OapA can promote attachment of
H. influenzae
. Expression of
oapA
gene in a nonadherent
Escherichia coli
strain significantly increased the binding to Chang epithelial cells, and disruption of the
oapA
gene with kanamycin resistance cassette insertion resulted in a significant loss of binding. These findings demonstrate that OapA plays a role in
H. influenzae
binding to human conjunctival epithelial cells.
Campylobacter jejuni
81-176 Associates with Microtubules and Dynein during Invasion of Human Intestinal Cells
Authors: Hu Lan , Kopecko Dennis J. 
Published: December 2019

ABSTRACT

Campylobacter jejuni
uptake into cultured INT407 cells was analyzed kinetically over a wide range of starting multiplicities of infection (MOI; from 0.02 to 20,000 bacteria/epithelial cell). The efficiency of internalization was the highest at MOI of 0.02 and decreased steadily at higher MOIs, presumably due to reported
C. jejuni
autoagglutination at higher densities. Total internalized
Campylobacter
CFU increased gradually from an MOI of 0.02 to a peak at an MOI of 200 (reaching an average of two bacteria internalized per epithelial cell) and decreased at higher MOIs. The invasion process was apparently saturated within 2 h at an MOI of 200, indicating stringent host cell limitations on this entry process. Furthermore, whereas control
Salmonella typhi
invaded all monolayer cells within 1 h, only two-thirds of monolayer cells were infected after 2 h with
C. jejuni
at MOIs of 200 to 2,000. The percentage of
Campylobacter
-infected host cells gradually increased to 85% after 7 h of infection, suggesting that
C. jejuni
entry may be host cell cycle dependent. Direct evidence of the involvement of microtubules in
C. jejuni
internalization, suggested previously by biochemical inhibitor studies, was obtained by time course immunofluorescence microscopic analyses. Bacteria initially bound to the tips of host cell membrane extensions containing microtubules, then aligned in parallel with microtubules during entry, colocalized specifically with microtubules and dynein but not with microfilaments, and moved over 4 h, presumably via microtubules to the perinuclear region of host cells. Orthovanadate, which inhibits dynein activity, specifically reduced
C. jejuni
81-176 entry, suggesting that this molecular motor is involved in entry and endosome trafficking during this novel bacterial internalization process. Collectively, these data suggest that
C. jejuni
enters host cells in a targeted and tightly controlled process leading to uptake into an endosomal vacuole which apparently moves intracellularly along microtubules via the molecular motor, dynein, to the perinuclear region.
Plasma Membrane Expression of Heat Shock Protein 60 In Vivo in Response to Infection
Authors: Belles Cindy , Kuhl Alicia , Nosheny Rachel , Carding Simon R. 
Published: December 2019

ABSTRACTHeat shock protein 60 (hsp60) is constitutively expressed in the mitochondria of eukaryotic cells. However, it has been identified in other subcellular compartments in several disease states and in transformed cells, and it is an immunogenic molecule in various infectious and autoimmune diseases. To better understand the factors that influence expression of hsp60 in normal cells in vivo, we analyzed its cellular and subcellular distribution in mice infected with the intracellular bacteriumListeria monocytogenes. Western blotting of subcellular fractionated spleen cells showed that although endogenous hsp60 was restricted to the mitochondria in noninfected animals, it was associated with the plasma membrane as a result of infection. The low levels of plasma membrane-associated hsp60 seen in the livers in noninfected animals subsequently increased during infection. Plasma membrane hsp60 expression did not correlate with bacterial growth, being most evident during or after bacterial clearance and persisting at 3 weeks postinfection. Using flow cytometry, we determined that Mac-1+, T-cell receptor γδ+, and B220+cells represented the major Hsp60+populations in spleens of infected mice. By contrast, B220+cells were the predominant hsp60+population in livers of infected mice. Of the immune cells analyzed, the kinetic profile of the γδ T-cell response most closely matched that of hsp60 expression in both the spleen and liver. Collectively, these findings show that during infection hsp60 can be localized to the plasma membrane of viable cells, particularly antigen-presenting cells, providing a means by which hsp60-reactive lymphocytes seen in various infectious disease and autoimmune disorders may be generated and maintained.
Intranasal Immunization of Mice with Influenza Vaccine in Combination with the Adjuvant LT-R72 Induces Potent Mucosal and Serum Immunity Which Is Stronger than That with Traditional Intramuscular Immunization
Authors: Barackman J. D. , Ott G. , O’Hagan D. T. 
Published: December 2019

ABSTRACTImmunization of mice by the intranasal route with influenza virus hemagglutinin in combination with the mutantEscherichia coliheat-labile enterotoxin R72 (LT-R72) induced significantly enhanced serum and mucosal antibodies, surpassing, in most cases, responses achieved by traditional intramuscular immunization using inactivated split influenza vaccine. Furthermore, intranasal immunization with LT-R72 induced a potent serum immunoglobulin G2a response, indicating that this adjuvant has Th1 character.
Identification of Amino Acid Residues of the T-Cell Epitope ofMycobacterium tuberculosisα Antigen Critical for Vβ11+Th1 Cells
Authors: Kariyone Ai , Higuchi Kazue , Yamamoto Shigeki , Nagasaka-Kametaka Ai , Harada Mamoru , Takahashi Akio , Harada Nobuyuki , Ogasawara Kazumasa , Takatsu Kiyoshi 
Published: December 2019

ABSTRACTStimulation ofMycobacterium tuberculosis-primed lymph node cells from C57BL/6 mice with α antigen (also known as antigen 85B and MPT59) induced cell proliferation, production of interleukin 2 and gamma interferon, and expansion of Vβ11+CD4+T cells in conjunction with antigen-presenting cells in an I-Ab-restricted manner. Using a series of 15-amino-acid peptides that overlapped each other by 5 amino acids and spanned the mature α antigen, we identified the antigenic epitope for α antigen-specific Vβ11+Th1 cells. That peptide (peptide-25), which corresponds to amino acid residues 240 to 254 of α antigen, contains a motif that is conserved in I-Aband requires processing by antigen-presenting cells. Using peptide-25-reactive Vβ11+T-cell clones and substituted peptide-25 mutants, we determined which amino acid residues within peptide-25 were critical for T-cell receptor (TCR) recognition. Our results showed that the amino acid residues at positions 245, 246, 248, 250, and 251 are important for recognition of TCRVβ11 and that residues at positions 244, 247, 249, and 252 are I-Abcontact residues. We also observed that active immunization of C57BL/6 mice with peptide-25 can lead to decreased bacterial load in the lungs ofM. tuberculosisH37Rv-infected mice. These results should provide us with a useful tool for delineating the regulation of Vβ11+Th1-cell development duringM. tuberculosisinfection and for developing a vaccine inducing a Th1-dominant immune response.
Detection of Intrastrain Antigenic Variation of
Bacteroides fragilis
Surface Polysaccharides by Monoclonal Antibody Labelling
Authors: Patrick Sheila , Gilpin Deirdre , Stevenson Leanne 
Published: December 2019

ABSTRACT

Bacteroides fragilis
is a constituent of the normal resident microbiota of the human intestine and is the gram-negative obligately anaerobic bacterium most frequently isolated from clinical infection. Surface polysaccharides are implicated as potential virulence determinants. We present evidence of within strain immunochemical variation of surface polysaccharides in populations that are noncapsulate by light microscopy as determined by monoclonal antibody labelling. Expression of individual epitopes can be enriched from a population of an individual strain by use of immunomagnetic beads. Also, individual colonies in which either >94% or <7% of the bacteria carry an individual epitope retain this level of expression when subcultured into broth. In broth cultures where >94% of the bacteria carry a given epitope, there is no enrichment for other epitopes recognized by different polysaccharide-specific monoclonal antibodies. This intrastrain variation has important implications for the development of potential vaccines or immunodiagnostic tests.
Truncated Surface Protective Antigen (SpaA) of
Erysipelothrix rhusiopathiae
Serotype 1a Elicits Protection against Challenge with Serotypes 1a and 2b in Pigs
Authors: Imada Yumiko , Goji Noriko , Ishikawa Hitoshi , Kishima Masato , Sekizaki Tsutomu 
Published: December 2019

ABSTRACT

Erysipelothrix rhusiopathiae
is a causal agent of swine erysipelas, which is of economic importance in the swine industry by virtue of causing acute septicemia, chronic arthritis, and endocarditis. However, little is known about the genetic properties of its protective antigens. Recently, a surface protective antigen (SpaA) gene was identified from serotype 2 in a mouse model. We cloned
spaA
from virulent strain Fujisawa (serotype 1a) and determined that the N-terminal 342 amino acids without C-terminal repeats of 20 amino acids have the ability to elicit protection in mice. Fusions of 342 amino acids of Fujisawa SpaA and histidine hexamer (HisSpa1.0) protected pigs against challenge with both serotype 1 and serotype 2, the most important serotypes in the swine industry. Pigs immunized with HisSpa1.0 reacted well with both HisSpa1.0 and intact SpaA by enzyme-linked immunosorbent assay and immunoblotting. Serum collected at the time of challenge from a pig immunized with HisSpa1.0 markedly enhanced the in vitro phagocytic and killing activity of pig neutrophils against the bacteria. DNA sequences of protective regions of
spaA
genes from five strains of serotypes 1 and 2 were almost identical. The full DNA sequences also seemed to be conserved among strains of all 12 serotype reference strains harboring the
spaA
gene by restriction fragment length polymorphism analysis of PCR products. These results indicates that SpaA is a common protective antigen of serotypes 1 and 2 of
E. rhusiopathiae
in swine and will be a useful tool for development of new types of vaccines and diagnostic tools for effective control of the disease.
Down-Regulation of Th2 Responses byBrucella abortus, a Strong Th1 Stimulus, Correlates with Alterations in the B7.2-CD28 Pathway
Authors: Agranovich Inna , Scott Dorothy E. , Terle Douglas , Lee Katherine , Golding Basil 
Published: December 2019

ABSTRACTDown-regulation of the Th2-like response induced by ovalbumin-alum (OVA/alum) immunization by heat-killedBrucella abortuswas not reversed by anti-IL-12 antibody treatment or in gamma interferon (IFN-γ) knockout mice, suggesting that induction of Th1 cytokines was not the only mechanism involved in theB. abortus-mediated inhibition of the Th2 response to OVA/alum. The focus of this study was to determine whether an alternative pathway involves alteration in expression of costimulatory molecules. First we show that the Th2-like response to OVA/alum is dependent on B7.2 interaction with ligand since it can be abrogated by anti-B7.2 treatment. Expression of costimulatory molecules was then studied in mice immunized with OVA/alum in the absence or presence ofB. abortus. B7.2, but not B7.1, was up-regulated on mouse non-T and T cells following immunization withB. abortus. Surprisingly,B. abortusinduced down-regulation of CD28 and up-regulation of B7.2 on murine CD4+and CD8+T cells. These effects on T cells were maximal for CD28 and B7.2 at 40 to 48 h and were not dependent on interleukin-12 (IL-12) or IFN-γ. On the basis of these results, we propose that the IL-12/IFN-γ-independent inhibition of Th2 responses to OVA/alum is secondary to the effects ofB. abortuson expression of costimulatory molecules on T cells. We suggest that down-regulation of CD28 following activation inhibits subsequent differentiation of Th0 into Th2 cells. In addition, decreased expression of CD28 and increased expression of B7.2 on T cells would favor B7.2 interaction with CTLA-4 on T cells, and this could provide a negative signal to developing Th2 cells.
Sulfated Polysaccharide-Directed Recruitment of Mammalian Host Proteins: a Novel Strategy in Microbial Pathogenesis
Authors: Duensing Thomas D. , Wing Jenny S. , van Putten Jos P. M. 
Published: December 2019

ABSTRACTFundamental to the virulence of microbial pathogens is their capacity for adaptation and survival within variable, and often hostile, environments encountered in the host. We describe a novel, extragenomic mechanism of surface modulation which may amplify the adaptive and pathogenic potential of numerous bacterial species, includingStaphylococcus,Yersinia, and pathogenicNeisseriaspecies, as well asHelicobacter pyloriandStreptococcus pyogenes. The mechanism involves specific bacterial recruitment of heparin, glycosaminoglycans, or related sulfated polysaccharides, which in turn serve as universal binding sites for a diverse array of mammalian heparin binding proteins, including adhesive glycoproteins (vitronectin and fibronectin), inflammatory (MCP-3, PF-4, and MIP-1α) and immunomodulatory (gamma interferon) intermediates, and fibroblast growth factor. This strategy impacts key aspects of microbial pathogenicity as exemplified by increased bacterial invasion of epithelial cells and inhibition of chemokine-induced chemotaxis. Our findings illustrate a previously unrecognized form of parasitism that complements classical virulence strategies encoded within the microbial genome.
Stress-Induced Membrane Association of the
Streptococcus mutans
GTP-Binding Protein, an Essential G Protein, and Investigation of Its Physiological Role by Utilizing an Antisense RNA Strategy
Authors: Baev Didi , England Reg , Kuramitsu Howard K. 
Published: December 2019

ABSTRACT

SGP (for
Streptococcus
GTP-binding protein) is a
Streptococcus mutans
essential GTPase which has significant sequence identity to the previously identified
Escherichia coli
Era protein and to numerous other prokaryotic GTPase proteins of unknown function. Recent studies in our laboratory have addressed the possible role of SGP in the stress response of the oral pathogen
S. mutans
. Here we report that during growth in the early stationary phase, and in response to elevated temperatures or acidic pH, the distribution of SGP between the cytoplasm and the membranes of
S. mutans
cells varies. Immunoblot analysis of soluble and membrane protein fractions collected from the mid-log and early stationary growth phases of bacterial populations grown at normal temperature (37°C) and at the elevated temperature of 43°C, or at acidic pH, demonstrated that the total amount of SGP increased with the age of the bacterial culture, elevated temperature, or acidic pH. Furthermore, it was established that a substantial amount of SGP is associated with the membrane fraction under stress conditions. In order to investigate the physiological role of SGP, we constructed an
S. mutans
strain capable of chromosomal
sgp
antisense RNA expression, which interferes with the normal information processing of the
sgp
gene. Utilizing this strain, we determined conditions whereby the streptococcal cells can be depleted of SGP, thus avoiding the problem of constructing a conditional lethal system. From the results of measurements of the nucleotide pools extracted from the antisense strain and its isogenic counterpart, we propose that one of the physiological roles of SGP is regulation and modulation of the GTP/GDP ratio under different growth conditions. Moreover, we observed that in SGP-depleted cells the levels of glucan-binding protein A (GbpA) substantially increased, suggesting that GbpA may have stress response-related physiological functions. Finally, the potential applications of the antisense RNA approach that we employed are discussed.