bluuebunny/crossref_metadata_2025_split huggingface dataset:
Presence of Oligoclonal T Cells in Cerebrospinal Fluid of a Child with Multiphasic Disseminated Encephalomyelitis following Hepatitis A Virus Infection
Authors: Oleszak Emilia L. , Lin Wan Lu , Legido Agustin , Melvin Joseph , Hardison Huntley , Hoffman Brad E. , Katsetos Christos D. , Platsoucas Chris D. 
Published: July 2002

ABSTRACTWe have investigated the clonality of β-chain T-cell receptor (TCR) transcripts from the cerebrospinal fluid (CSF) and peripheral blood from a 7-year old child who developed a multiphasic disseminated encephalomyelitis following an infection with hepatitis A virus. We amplified β-chain TCR transcripts by nonpalindromic adaptor (NPA)-PCR–Vβ-specific PCR. TCR transcripts from only five Vβ families (Vβ13, Vβ3, Vβ17, Vβ8, and Vβ20) were detected in CSF. The amplified products were combined, cloned, and sequenced. Sequence analysis revealed in the CSF substantial proportions of identical β-chain of TCR transcripts, demonstrating oligoclonal populations of T cells. Seventeen of 35 (48%) transcripts were 100% identical, demonstrating a major Vβ13.3 Dβ2.1 Jβ1.3 clonal expansion. Six of 35 (17%) transcripts were also 100% identical, revealing a second Vβ13 clonal expansion (Vβ13.1 Dβ2.1 Jβ1.2). Clonal expansions were also found within the Vβ3 family (transcript Vβ3.1 Dβ2.1 Jβ1.5 accounted for 5 of 35 transcripts [14%]) and within the Vβ20 family (transcript Vβ20.1 Dβ1.1 Jβ2.4 accounted for 3 of 35 transcripts [8%]). These results demonstrate the presence of T-cell oligoclonal expansions in the CSF of this patient following infection with hepatitis A virus. Analysis of the CDR3 motifs revealed that two of the clonally expanded T-cell clones exhibited substantial homology to myelin basic protein-reactive T-cell clones. In contrast, all Vβ TCR families were expressed in peripheral blood lymphocytes. Oligoclonal expansions of T cells were not detected in the peripheral blood of this patient. It remains to be determined whether these clonally expanded T cells are specific for hepatitis A viral antigen(s) or host central nervous system antigen(s) and whether molecular mimicry between hepatitis A viral protein and a host protein is responsible for demyelinating disease in this patient.
Calcein-Acetyoxymethyl Cytotoxicity Assay: Standardization of a Method Allowing Additional Analyses on Recovered Effector Cells and Supernatants
Authors: Neri Simona , Mariani Erminia , Meneghetti Alessandra , Cattini Luca , Facchini Andrea 
Published: July 2002

ABSTRACTCytotoxicity assays provide an in vitro evaluation of the lytic activity of NK and T cells against tumors or transformed cells. However, none of these methods allow the recovery of cells or supernatants after the assay. We standardized a microcytotoxicity test using calcein-acetoxymethyl (calcein-AM) dye that requires very small quantities of cells while maintaining the same sensitivity as the traditional51Cr assay. The assay is applicable to resting as well as activated human effector cells and uses different targets such as human cell lines that are adherent or growing in suspension and resistant or sensitive. The most important feature of the method is the possibility of recovering cells and supernatants for additional analyses such as phenotyping and evaluation of soluble factors.
Lactoferrin Protects against Development of Hepatitis Caused by Sensitization of Kupffer Cells by Lipopolysaccharide
Authors: Yamaguchi Makoto , Matsuura Motoi , Kobayashi Kiyoshi , Sasaki Hajime , Yajima Takaji , Kuwata Tamotsu 
Published: July 2002

ABSTRACTBALB/c mice were intravenously injected with lipopolysaccharide (LPS) (0.05 μg/g of body weight) 7 days after being primed with zymosan. Recombinant human lactoferrin (250 μg/g of body weight), intravenously administered 1 day before the injection of LPS, significantly lessened the severity of hepatitis, as assessed by levels of serum alanine transaminase compared to those seen when casein was administered. The transient rise of serum tumor necrosis factor alpha (TNF-α) after LPS treatment was also significantly lowered by the intravenous administration of lactoferrin, suggesting that the effect of lactoferrin was due to the suppression of TNF-α production. The following results indicate that the sites of action of lactoferrin for the suppression of the development of this type of hepatitis are Kupffer cells. Gadolinium chloride, a substance known to eliminate Kupffer cells, administered 1 day before LPS, inhibited the transient rise of TNF-α and protected against the development of hepatitis. Kupffer cells isolated from mice intraperitoneally injected with recombinant human lactoferrin became refractory to LPS. The specific interaction of recombinant human lactoferrin with the Kupffer cells was shown by a binding assay, which revealed two types of binding sites on mouse Kupffer cells. Of the two dissociation constants determined in this way, the lower dissociation constant, 0.47 × 10−6M, was within the range of the 50% effective doses for the suppression of TNF-α production. These results suggest that recombinant human lactoferrin administered to mice suppresses the production of TNF-α by Kupffer cells by directly associating with the binding sites on these cells.
Grape Seed Extract Activates Th1 Cells In Vitro
Authors: Nair Narayanan , Mahajan Supriya , Chawda Ram , Kandaswami Chithan , Shanahan Thomas C. , Schwartz Stanley A. 
Published: July 2002

ABSTRACT
Although flavonoids manifest a diverse range of biological activities, including antitumor and antiviral effects, the molecular mechanisms underlying these activities await elucidation. We hypothesize that the flavonoid constituents of a proprietary grape seed extract (GSE) that contains procyandins exert significant antiviral and antitumor effects, by inducing production of the Th1-derived cytokine gamma interferon (IFN-γ) by peripheral blood mononuclear cells) from healthy donors. Our results show that GSE significantly induced the transcription of IFN-γ mRNA as demonstrated by reverse transcription-PCR but had no effect on the Th2-derived cytokine interleukin-6. The enhancing effect of GSE on IFN-γ expression was further supported by a concomitant increase in the number of cells with intracytoplasmic IFN-γ as well as the synthesis and secretion of IFN-γ. Our results demonstrate that the potentially beneficial immunostimulatory effects of GSE may be mediated through the induction of IFN-γ.
Epidemiology and Clinical Features ofCryptosporidiumInfection in Immunocompromised Patients
Authors: Hunter Paul R. , Nichols Gordon 
Published: July 2002

SUMMARYCryptosporidium spp. are a major cause of diarrheal disease in both immunocompetent and immunodeficient individuals. They also cause waterborne disease in both the United States and United Kingdom. Studies on the mechanisms of immunity to cryptosporidiosis indicate the importance of the T-cell response. The spectrum and severity of disease in immunocompromised individuals with cryptosporidiosis reflect this importance since the most severe disease is seen in individuals with defects in the T-cell response. The most commonly studied group is that of patients with AIDS. These patients suffer from more severe and prolonged gastrointestinal disease that can be fatal; in addition, body systems other than the gastrointestinal tract may be affected. The widespread use of antiretroviral therapy does appear to be having a beneficial effect on recovery from cryptosporidiosis and on the frequency of infection in human immunodeficiency virus-positive patients. Other diseases that are associated with increased risk of severe cryptosporidiosis, such as primary immunodeficiencies, most notably severe combined immunodeficiency syndrome, are also predominantly associated with T-cell defects. Of the remaining groups, children with acute leukemia seem to be most at risk from cryptosporidiosis. There is less evidence of severe complications in patients with other malignant diseases or in those receiving immunosuppressive chemotherapy.
Integrated Approach to Malaria Control
Authors: Shiff Clive 
Published: July 2002

SUMMARYMalaria draws global attention in a cyclic manner, with interest and associated financing waxing and waning according to political and humanitarian concerns. Currently we are on an upswing, which should be carefully developed. Malaria parasites have been eliminated from Europe and North America through the use of residual insecticides and manipulation of environmental and ecological characteristics; however, in many tropical and some temperate areas the incidence of disease is increasing dramatically. Much of this increase results from a breakdown of effective control methods developed and implemented in the 1960s, but it has also occurred because of a lack of trained scientists and control specialists who live and work in the areas of endemic infection. Add to this the widespread resistance to the most effective antimalarial drug, chloroquine, developing resistance to other first-line drugs such as sulfadoxine-pyrimethamine, and resistance of certain vector species of mosquito to some of the previously effective insecticides and we have a crisis situation. Vaccine research has proceeded for over 30 years, but as yet there is no effective product, although research continues in many promising areas. A global strategy for malaria control has been accepted, but there are critics who suggest that the single strategy cannot confront the wide range of conditions in which malaria exists and that reliance on chemotherapy without proper control of drug usage and diagnosis will select for drug resistant parasites, thus exacerbating the problem. An integrated approach to control using vector control strategies based on the biology of the mosquito, the epidemiology of the parasite, and human behavior patterns is needed to prevent continued upsurge in malaria in the endemic areas.
Identification of 11 pH-Regulated Genes in
Borrelia burgdorferi
Localizing to Linear Plasmids
Authors: Carroll James A. , Cordova Rebecca M. , Garon Claude F. 
Published: July 2002

ABSTRACT

When
Borrelia burgdorferi
is transmitted from the tick vector to the mammalian host, the bacterium experiences alterations in its environment, such as changes in temperature and pH. Previously, we observed numerous alterations in the membrane protein profile when
B. burgdorferi
B31 was grown at pH 7.0 compared to pH 8.0. Here we identify 11 genes localizing to linear plasmids that are up-regulated at pH 7.0 relative to pH 8.0 in vitro. Seven genes (
bba03, bba24, bba64, bba66, bbe31, bbj41/bbi39
[encoding products that are 99% identical], and
bbk01
) were indirectly identified by proteomic analysis of membrane proteins. Another gene,
bba36
, was identified by screening a
B. burgdorferi
B31 genomic library with cross-adsorbed hyperimmune rabbit serum. Two additional genes,
bba65
and
bba73
, were identified by Northern blot analysis. Genes
bba64, bba65, bba66, bbj41/bbi39
, and
bba73
are members of paralogous gene family 54, and
bbe31
is a member of the closely related paralogous gene family 60. Gene
bba24
is part of a bicistronic operon with
bba25
that encodes the well-characterized decorin binding proteins A and B. All 11 genes were transcriptionally regulated, yet the degree of pH regulation varied, with some genes more tightly regulated than others. The regions upstream of these pH-regulated genes appeared to be unrelated, yet many contained dyad repeats ranging from 12 to 25 nucleotides in length that may be involved in the regulation of these genes.
Immunopathologic Effects of Tumor Necrosis Factor Alpha in Murine Mycobacterial Infection Are Dose Dependent
Authors: Bekker Linda-Gail , Moreira Andre L. , Bergtold Amy , Freeman Sherry , Ryffel Bernard , Kaplan Gilla 
Published: July 2002

ABSTRACTIn experimental mycobacterial infection, tumor necrosis factor alpha (TNF-α) is required for control of bacillary growth and the protective granulomatous response, but may cause immunopathology. To directly examine the positive and detrimental effects of this cytokine, a murine model was used in which different amounts of TNF-α were delivered to the site of infection. Mice with a disruption in the TNF-α gene (TNF-KO) or wild-type mice were infected with low or high doses of recombinantMycobacterium bovisBCG that secreted murine TNF-α (BCG-TNF). Infection of TNF-KO mice with BCG containing the vector (BCG-vector) at a low dose led to increased bacillary load in all organs and an extensive granulomatous response in the lungs and spleen. The mice succumbed to the infection by ∼40 days. However, when TNF-KO mice were infected with low doses of BCG-TNF, bacillary growth was controlled, granulomas were small and well differentiated, the spleen was not enlarged, and the mice survived. Infection with high inocula of BCG-TNF resulted in bacterial clearance, but was accompanied by severe inflammation in the lungs and spleen and earlier death compared to the results from the mice infected with high inocula of BCG-vector. Wild-type mice controlled infection with either recombinant strain, but showed decreased survival following high-dose BCG-TNF infection. The effects of TNF-α required signaling through an intact receptor, since the differential effects were not observed when TNF-α receptor-deficient mice were infected. The results suggest that the relative amount of TNF-α at the site of infection determines whether the cytokine is protective or destructive.
Mycobacterium bovis-Infected Cervine Alveolar Macrophages Secrete Lymphoreactive Lipid Antigens
Authors: Aldwell Frank E. , Dicker Bridget L. , Da Silva Tatley Fernanda M. , Cross Martin F. , Liggett Simon , Mackintosh Colin G. , Griffin J. Frank T. 
Published: July 2002

ABSTRACTTuberculosis is caused by intracellular bacteria belonging to the genusMycobacterium, includingM. tuberculosisandM. bovis. Alveolar macrophages (AMs) are the primary host cell for inhaled mycobacteria. However, little is known about the mechanisms by which infected AMs can process and present mycobacterial antigens to primed lymphocytes and how these responses may affect ensuing protection in the host. In the present study, we sought to determine whether AMs from a naturally susceptible host forMycobacterium bovis(red deer) could produce and secrete soluble immunoreactive antigens following mycobacterial infection in vitro. Confluent monolayers of deer AMs were infected with either heat-killed or live virulentM. bovisorM. bovisBCG at a multiplicity of infection of 5:1 and cultured for 48 h. Culture supernatants were collected, concentrated, and tested for the presence of mycobacterial antigens in a lymphocyte proliferation assay by using peripheral blood mononuclear cells fromM. bovis-sensitized or naive deer. Supernatants derived from macrophages which had been infected with live bacilli stimulated the proliferation of antigen-sensitized, but not naive, lymphocytes. Supernatants derived from uninoculated AMs or AMs inoculated with heat-killed bacilli failed to stimulate lymphocyte proliferation. The lymphoproliferative activity was retained following lipid extraction of the supernatants, which were free of amino groups as determined by thin-layer chromatography. These results demonstrate that mycobacteria which are actively growing within AMs produce lipids which are secreted into the extracellular milieu and that these lipids are recognized by lymphocytes from mycobacterium-primed hosts. We suggest that mycobacterial lipids are released from AMs following aerosol infection in vivo and that they play an important role in the early immune response to tuberculosis.
A Peptide Permease Mutant of
Mycobacterium bovis
BCG Resistant to the Toxic Peptides Glutathione and
S
-Nitrosoglutathione
Authors: Green Renee M. , Seth Anjali , Connell Nancy D. 
Published: July 2002

ABSTRACT

Oligopeptides play important roles in bacterial nutrition and signaling. Using sequences from the available genome database for
Mycobacterium tuberculosis
H37Rv, the oligopeptide permease operon (
oppBCDA
) of
Mycobacterium bovis
BCG was cloned from a cosmid library. An
opp
mutant strain was constructed by homologous recombination with an allele of
oppD
interrupted by kanamycin and streptomycin resistance markers. The deletion was complemented with a wild-type copy of the
opp
operon. Two approaches were taken to characterize the peptide transporter defect in this mutant strain. First, growth of wild-type and mutant strains was monitored in media containing a wide variety of peptides as sole source of carbon and/or nitrogen. Among 25 peptides ranging from two to six amino acids in length, none was capable of supporting measurable growth as the sole carbon source in either wild-type or mutant strains. The second approach exploited the resistance of permease mutants to toxic substrates. The tripeptide glutathione (γ-glutamyl-
l
-cyteinylglycine [GSH]) is toxic to wild-type BCG and was used successfully to characterize peptide uptake in the
opp
mutant. In 2 mM GSH, growth of the wild-type strain is inhibited, whereas the
opp
mutant is resistant to concentrations as high as 10 mM. Similar results were found with the tripeptide
S
-nitrosoglutathione (GSNO), thought to be a donor of NO in mammalian cells. Using incorporation of [
3
H]uracil to monitor the effects of GSH and GSNO on macromolecular synthesis in growing cells, it was demonstrated that the
opp
mutant is resistant, whereas the wild type and the mutant complemented with a wild-type copy of the operon are sensitive to both tripeptides. In uptake measurements, incorporation of [
3
H]GSH is reduced in the mutant compared with wild type and the complemented mutant. Finally, growth of the three strains in the tripeptides suggests that GSH is bacteriostatic, whereas GSNO is bacteriocidal.
GAA Trinucleotide Repeat Region Regulates M9/pMGA Gene Expression in
Mycoplasma gallisepticum
Authors: Liu Li , Dybvig Kevin , Panangala Victor S. , van Santen Vicky L. , French Christopher T. 
Published: July 2002

ABSTRACT

Mycoplasma gallisepticum
, the cause of chronic respiratory infections in the avian host, possesses a family of M9/pMGA genes encoding an adhesin(s) associated with hemagglutination. Nucleotide sequences of M9/pMGA gene family members indicate extensive sequence similarity in the promoter regions of both the transcribed and silent genes. The mechanism that regulates M9/pMGA gene expression is unknown, but studies have revealed an apparent correlation between gene expression and the number of tandem GAA repeat motifs located upstream of the putative promoter. In this study, transposon Tn
4001
was used as a vector with the
Escherichia coli lacZ
gene as the reporter system to examine the role of the GAA repeats in M9/pMGA gene expression in
M. gallisepticum
. A 336-bp M9 gene fragment (containing the GAA repeat region, the promoter, and the translation start codon) was amplified by PCR, ligated with a
lacZ
gene from
E. coli
, and inserted into the Tn
4001
-containing plasmid pISM2062. This construct was transformed into
M. gallisepticum
PG31. Transformants were filter cloned on agar supplemented with 5-bromo-4-chloro-3-indolyl-β-
d
-galactopyranoside (X-Gal) to monitor
lacZ
gene expression on the basis of blue/white color selection. Several cycles of filter cloning resulted in cell lineages in which
lacZ
gene expression alternated between the On and Off states in successive generations of progeny clones. The promoter regions of the M9-
lacZ
hybrid genes of individual progeny clones were amplified by PCR and sequenced. The only differences between the promoter regions of the blue and white colonies were in the number of GAA repeats. Clones that expressed
lacZ
had exactly 12 tandem copies of the GAA repeat. Clones that did not express
lacZ
invariably had either more than 12 (14 to 16) or fewer than 12 (5 to 11) GAA repeats. Southern analysis of
M. gallisepticum
chromosomal DNA confirmed that the phase-variable expression of the
lacZ
reporter gene was not caused by Tn
4001
transposition. These data strongly indicate that changes in the length of the GAA repeat region are responsible for regulating M9/pMGA gene expression.
Differential Regulation of CD4 Lymphocyte Recruitment between the Upper and Lower Regions of the Genital Tract duringChlamydia trachomatisInfection
Authors: Kelly Kathleen A. , Walker Jennifer C. , Jameel Shimul H. , Gray Heather L. , Rank Roger G. 
Published: July 2002

ABSTRACTGenital infection withChlamydia trachomatisresults in both the local recruitment of protective immune responses and an inflammatory infiltrate that may also participate in tubal pathology. As a beginning to understanding the etiology of immune system-mediated tubal pathology, we evaluated the regional recruitment of lymphocyte subsets to different areas of the female genital tract (GT) over the course of a murine infection with the mouse pneumonitis agent ofChlamydia trachomatis(MoPn). Using flow cytometric techniques we found that the CD4 lymphocyte subset was preferentially recruited to the upper GT (oviduct and uterine horn) over the lower GT (cervical-vaginal region) throughout the course of MoPn infection. The influx of CD4 cells also correlated with the expression of endothelial cell adhesion molecules (ECAMs) and in vitro lymphocyte adherence in the upper GT. Interestingly, the expression of ECAMs in the lower GT was not maintained longer than 7 days after infection, even in the presence of viable chlamydiae. Taken together, these data suggest that regulatory mechanisms of lymphocyte recruitment differ between the upper and lower regions of the GT and may influence the clearance of chlamydiae and the development of tubal pathology.
pH-Induced Conformational Changes in
Clostridium difficile
Toxin B
Authors: Qa'Dan Maen , Spyres Lea M. , Ballard Jimmy D. 
Published: July 2002

ABSTRACT

Toxin B from
Clostridium difficile
is a monoglucosylating toxin that targets substrates within the cytosol of mammalian cells. In this study, we investigated the impact of acidic pH on cytosolic entry and structural changes within toxin B. Bafilomycin A1 was used to block endosomal acidification and subsequent toxin B translocation. Cytopathic effects could be completely blocked by addition of bafilomycin A1 up to 20 min following toxin treatment. Furthermore, providing a low extracellular pH could circumvent the effect of bafilomycin A1 and other lysosomotropic agents. Acid pH-induced structural changes were monitored by using the fluorescent probe 2-(
p
-toluidinyl) naphthalene-6-sulfonic acid, sodium salt (TNS), inherent tryptophan fluorescence, and relative susceptibility to a specific protease. As the toxin was exposed to lower pH there was an increase in TNS fluorescence, suggesting the exposure of hydrophobic domains by toxin B. The change in hydrophobicity appeared to be reversible, since returning the pH to neutrality abrogated TNS fluorescence. Furthermore, tryptophan fluorescence was quenched at the acidic pH, indicating that domains may have been moving into more aqueous environments. Toxin B also demonstrated variable susceptibility to
Staphylococcus aureus
V8 protease at neutral and acidic pH, further suggesting pH-induced structural changes in this protein.