bluuebunny/crossref_metadata_2025 huggingface dataset:
Angiotensin II Increases Host Resistance to Peritonitis
Authors: Rodgers Kathleen , Xiong Shiquan , Espinoza Theresa , Roda Norma , Maldonado Sonia , diZerega Gere S.
Published: July 2002

Studies by other laboratories have shown that angiotensin II (AII) can affect the function of cells which comprise the immune system. In the present study, the effect of AII on the function of peritoneal macrophages and peripheral blood monocytes was assessed. In vitro exposure (4 h prior to assay) of peritoneal macrophages from mice and rats to AII increased the percentage of cells that phagocytosed opsonized yeast and the number of yeast per macrophage. Furthermore, AII increased the respiratory burst capacity of peritoneal macrophages from mice and rats and peripheral blood mononuclear cells from humans. Because of these observations, the effect of AII on host resistance to bacterial infection was assessed. Intraperitoneal administration of AII was shown to increase host resistance (reduced abscess formation) in an animal model of bacterial peritonitis. Studies were then conducted to assess whether parenteral administration of AII, a clinically relevant route, could affect peritoneal host resistance in a manner similar to that observed after peritoneal administration. These studies showed that subcutaneous administration of AII throughout the postinfection interval increased the level of host resistance to bacterial peritonitis. Furthermore, in a study which compared AII and Neupogen, an agent approved for use for the reduction of febrile neutropenia after myeloablative therapy, daily subcutaneous administration of AII reduced abscess size and incidence, whereas Neupogen did not have any therapeutic benefit in this model. These data suggest that AII may be of therapeutic benefit as an immunomodulatory agent.
Subtilisin Increases Macromolecular Efflux from the Oral Mucosa
Authors: Rubinstein Israel
Published: July 2002

The purpose of this study was to determine whether subtilisin, a potent serine proteinase derived fromBacillusspecies contaminating smokeless tobacco, increases macromolecular efflux from the oral mucosa and, if so, whether local elaboration of bradykinin mediates this response. Using intravital microscopy, I found that suffusion of subtilisin elicits significant, concentration-dependent leaky site formation and an increase in the clearance of fluorescein isothiocyanate-labeled dextran (molecular mass, 70 kDa) from the in situ hamster cheek pouch (P< 0.05). Heat-inactivated subtilisin had no significant effects on macromolecular efflux. Subtilisin-induced responses were significantly attenuated by Hoe 140 and NPC 17647, two structurally distinct selective bradykinin B2receptor antagonists, but not by des-Arg9-[Leu8]bradykinin, a selective bradykinin B1receptor antagonist, or CP-96,345, a selective neurokinin-1 receptor antagonist. Aprotinin, but not leupeptin, significantly attenuated subtilisin-induced increase in macromolecular efflux. Indomethacin had no significant effects on subtilisin-induced responses. Collectively, these data indicate that subtilisin increases the macromolecular efflux from the in situ hamster cheek pouch in a catalytic-site-dependent fashion through local elaboration of bradykinin. This response does not involve the stimulation of local afferent nerves or the production of prostaglandins.
CD8α-Deficient Mice Are Highly Susceptible to 5-Fluorouracil-Induced Lethality
Authors: Itoh Naoto , Nishimura Hitoshi , Matsuguchi Tetsuya , Yajima Toshiki , Mokuno Yasuji , Hiromatsu Takashi , Nimura Yuji , Yoshikai Yasunobu
Published: July 2002

Intestinal intraepithelial lymphocytes (i-IEL) expressing CD8α are located in the intestine and may confer protection against invasion of intestinal microflora. We found that mice rendered deficient in CD8α molecules by homologous recombination were susceptible to 5-fluorouracil (5-FU)-induced lethality accompanied by translocation of members of the enterobacteria. The number of i-IEL was greatly reduced on day 6 after 5-FU administration in both CD8α+/−mice and CD8α−/−mice, whereas the recovery of the level of i-IEL thereafter was significantly impaired in CD8α−/−mice compared with that in CD8α+/−mice. The ability of i-IEL to produce gamma interferon in response to immobilized T-cell receptor (TCR) αβ or TCR γδ monoclonal antibodies was significantly lower in CD8α−/−mice than in CD8α+/−mice. Transfer of CD8+i-IEL conferred significant protection against 5-FU-induced lethality in CD8α−/−mice. The results suggest that CD8+i-IEL play an important role in protection against 5-FU-induced lethality with translocation ofEnterobacteriaceae.
Treatment with Megestrol Acetate Improves Human Immunodeficiency Virus-Associated Immune Thrombocytopenia
Authors: Gomez Francisco , Ruiz Pedro , Lopez Rafaela , Rivera Consuelo
Published: July 2002

Splenic macrophage Fcγ receptors participate in the pathophysiology of immune cytopenias, and in such disorders, the beneficial effects of glucocorticoids are in part mediated by decreased expression of macrophage Fcγ receptors. In the animal model, progesterones, like glucocorticoids, inhibit expression of these receptors. Megestrol acetate (MA) is a progesterone frequently used for treating human immunodeficiency virus (HIV)-associated anorexia-cachexia. Twenty-eight patients with HIV-associated thrombocytopenia with shortened platelet survival and increased platelet-associated immunoglobulin G (IgG) who were being treated with MA for anorexia-cachexia were prospectively studied for a 6-month period to assess the potential role of progesterones in the treatment of immune thrombocytopenia. Treatment with MA for nonconsecutive periods of 2 months and 1 month significantly increased platelet count and platelet survival without significant alteration of platelet-associated immunoglobulin levels. Of the 28 patients studied, 22 presented a complete response, 19 presented a complete response 1 month after finishing the MA treatment regimen, and 12 remained in complete response for a further month. Expression of Fcγ receptors (FcγRI and FcγRII) by peripheral blood monocytes and the in vitro recognition of IgG-sensitized cells by monocytes were significantly decreased by the MA treatment. Decreased expression and functioning of these receptors significantly correlated with platelet counts and survival times, but no relationship was found with platelet-associated immunoglobulin, circulating immune complexes, body mass index, plasma HIV load, or CD4 lymphocyte levels. These results suggest that treatment with progesterones, like MA, may be an alternative therapy for immune cytopenias, with few side effects.
Leishmania majorReaches Distant Cutaneous Sites Where It Persists Transiently while Persisting Durably in the Primary Dermal Site and Its Draining Lymph Node: a Study with Laboratory Mice
Authors: Nicolas Luc , Sidjanski Sacha , Colle Jean-Hervé , Milon Geneviève
Published: July 2002

So far, studies ofLeishmaniapersistence in mice have used injections of parasites administered either intravenously in the tail vein or subcutaneously in the footpad. These routes poorly reflect the natural conditions when the sandfly delivers metacyclic promastigotes intradermally. In this study B10D2 and BALB/c mice were inoculated within the ear dermis with 104Leishmania majormetacyclic promastigotes. The parasite load was monitored by quantitative PCR in different tissues from the dermal inoculation site to distant tissues. The two sites of multiplication and persistence of parasites were the site ofL. majorinoculation and the draining lymph node (DLN), with a different pattern in the two mouse inbred lines. These two organs were the only sites harboring parasites 12 months postinoculation, with the DLN of BALB/c mice harboring around 107parasites, a stable load from months 3 to 12. In these two sites, 8 and 12 months after inoculation, interleukin 4 (IL-4), gamma interferon, and inducible nitric oxide synthase transcripts parallel the parasite load while IL-10 transcript levels remain high. In addition, at early time points until month 3, parasite DNA was also detected in distant tissues such as the contralateral noninoculated ear or the tail skin, indicating that blood was at least transiently disseminating the parasites. In contrast,L. majorDNA in liver, spleen, and femoral bone marrow remained sporadic in mice of both lines. This study is discussed within the framework ofLeishmaniatransmission from the vertebrate host to the sandfly vector, a complex process still poorly understood.
Pyelonephritogenic Diffusely AdheringEscherichia coliEC7372 Harboring Dr-II Adhesin Carries Classical Uropathogenic Virulence Genes and Promotes Cell Lysis and Apoptosis in Polarized Epithelial Caco-2/TC7 Cells
Authors: Guignot Julie , Breard Jacqueline , Bernet-Camard Marie-Françoise , Peiffer Isabelle , Nowicki Bogdan J. , Servin Alain L. , Blanc-Potard Anne-Beatrice
Published: July 2002

Diffusely adhering Escherichia coli (DAEC) strains expressing adhesins of the Afa/Dr family bind to epithelial cells in a diffuse adherence pattern by recognizing a common receptor, the decay-accelerating factor (CD55). Recently, a novel CD55-binding adhesin, named Dr-II, was identified from the pyelonephritogenic strain EC7372. In this report, we show that despite the low level of sequence identity between Dr-II and other members of the Afa/Dr family, EC7372 induces pathophysiological effects similar to those induced by other Afa/Dr DAEC strains on the polarized epithelial cell line Caco-2/TC7. Specifically, the Dr-II adhesin was sufficient to promote CD55 and CD66e clustering around adhering bacteria and apical cytoskeleton rearrangements. Unlike other Afa/Dr DAEC strains, EC7372 expresses a functional hemolysin that promotes a rapid cellular lysis. In addition, cell death by apoptosis or necrosis was observed in EC7372-infected Caco-2/TC7 cells, depending on infection time. Our results indicate that EC7372 harbors a pathogenicity island (PAI) similar to the one described for the pyelonephritogenic strain CFT073, which carries bothhlyandpapoperons. Cumulatively, our findings indicate that strain EC7372 can be considered a prototype of a subclass of Afa/Dr DAEC isolates that have acquired a PAI harboring several classical uropathogenic virulence genes.
Identification of Two Distinct Types of Flagellar Cap Proteins, FliD, in
Pseudomonas aeruginosa
Authors: Arora Shiwani K. , Dasgupta Nandini , Lory Stephen , Ramphal Reuben
Published: July 2002

Binding of
Pseudomonas aeruginosa
strain PAK to mucin has been shown to be mediated by the flagellar cap protein, product of the
fliD
gene. Since the flagellar cap is very likely an exposed structure, the FliD polypeptide should be recognized by the host immune system, analogous to the recognition of dominant epitopes located in the exposed parts of the flagellin polypeptide within the assembled flagellum. In
P. aeruginosa
, a number of distinct flagellin variants are made, and these variable sequences presumably allow the newly infected
P. aeruginosa
to escape recognition by the antibody induced during a previous infection. Since similar mechanisms may direct the selection of FliD variants, we examined the extent of sequence heterogeneity among various FliD sequences among a selected group of
P. aeruginosa
. The results of PCR and nucleotide sequencing of the
fliD
region of eight different
P. aeruginosa
strains (laboratory strains PAK, PAO1, and PA103; clinical strains 1244, CS2, and CS32; cystic fibrosis strains CS29 and MDR) suggested that there were two distinct types of FliD in
P. aeruginosa
, which we named A type and B type. The results of Western blotting using the polyclonal antibodies raised against the purified FliD of A type (PAK) or B type (PAO1) further confirmed the existence of two distinct antigenic types of FliD proteins, with no cross-reactivity between the two serotypes. Further Western immunoblot analysis of the same strains using polyclonal FliC antibody showed that the strains with A-type FliD possessed a-type FliC and those with B-type FliD had b-type FliC. Similar Western blot analyses of 50 more
P. aeruginosa
strains obtained from varied sources revealed that all strains contained either A-type or B-type FliD, suggesting the existence of only two types of FliD in
P. aeruginosa
and indicating that
fliC
and
fliD
were coinherited. This limited diversity of FliC and FliD serotypes seems to be a unique feature of flagellar proteins. A chromosomal mutant having an insertion in the
fliD
gene of
P. aeruginosa
PAO1 was constructed. The motility defect of this mutant and a previously constructed PAK
fliD
mutant was better complemented with the
fliD
gene of the homologous types.
Staphylococcal Enterotoxin A Acts through Nitric Oxide Synthase Mechanisms in Human Peripheral Blood Mononuclear Cells To Stimulate Synthesis of Pyrogenic Cytokines
Authors: Won Shen-Jeu , Huang Wu-Tein , Lai Yih-Shyong , Lin Mao-Tsun
Published: July 2002

The pyrogenic response to supernatant fluids obtained from human peripheral blood mononuclear cells (PBMC) stimulated with staphylococcal enterotoxin A (SEA) was characteristic of a response to an endogenous pyrogen in that it was brief and monophasic and was destroyed by heating supernatant fluids at 70°C for 30 min. The febrile responses were in parallel with the levels of interleukin-1 (IL-1), tumor necrosis factor (TNF), interferon-γ (IFN-γ), IL-2, and IL-6 in supernatant fluids obtained from PBMC treated with SEA. Both the pyrogenicity and the levels of IL-1, TNF, IFN-γ, IL-2, and IL-6 in supernatant fluids started to rise at 6 to 18 h and reached their peak levels at 24 to 96 h after SEA incubation. Both the fever and the increased levels of IL-1, TNF, IFN-γ, IL-2, and IL-6 in supernatant fluids obtained from the SEA-stimulated PBMC were decreased by incubating SEA-PBMC with anisomycin (a protein synthesis inhibitor), aminoguanidine (an inhibitor of inducible nitric oxide synthase [NOS]), or dexamethasone (an inhibitor of NOS). The febrile response to supernatant fluids obtained from the SEA-stimulated PBMC was attenuated by adding either anti-IL-1β, anti-TNF-α, or anti-IFN-γ monoclonal antibody (MAb) to supernatant fluids. The antipyretic effects exerted by anti-IL-1β MAb were greater than those exerted by anti-TNF-α or anti-IFN-γ MAb. The data suggest that SEA acts through the NOS mechanisms in PBMC to stimulate synthesis of pyrogenic cytokines (in particular, the IL-1β).
Interleukin-12 Neutralization Alters Lung Inflammation and Leukocyte Expression of CD80, CD86, and Major Histocompatibility Complex Class II in Mice Infected withHistoplasma capsulatum
Authors: Cain Judith A. , Deepe George S.
Published: July 2002

Histoplasma capsulatuminduces a cell-mediated immune response in lungs and lymphoid organs of mammals. Resolution of primary infection in mice depends on interleukin-12 (IL-12), since neutralization of this monokine increases susceptibility to infection. The present study was designed to determine if blockade of IL-12 disrupts the protective immune response by altering the influx of lineage-specific cells into infected lungs and the numbers of cells expressing CD80, CD86, CD119, and major histocompatibility complex class II (MHC II) molecules. In mice given anti-IL-12, there was a 2.5-fold decrease in total numbers of T cells on days 3 to 10 of infection and a 4-fold increase in Mac-1/Gr-1+cells on days 7 and 10 compared to infected controls. CD80+lung cells from anti-IL-12-treated mice were 2- to 3-fold greater than those from controls on days 7 and 10, whereas the total numbers of CD86+cells were 2- to 3-fold less and MHC II+cells were 1.5- to 2-fold less on days 3 and 5. Cells expressing CD119 were reduced 1.5-fold on day 5. Treatment with monoclonal antibodies (MAb) to CD80, CD86, or both reduced the fungal burden slightly compared to that in rat immunoglobulin G-treated controls, whereas after IL-12 neutralization, blocking of CD80 reduced the tissue burden by 2.5-fold and this correlated with a decrease in IL-4. Regardless, mortality was not altered by treatment with MAb to CD80 or CD86. We conclude that (i) IL-12 neutralization alters the nature of the inflammatory response in lungs and the expression of CD80 and CD86 on lineage-specific cells, (ii) the immune response during infection withH. capsulatumis controlled via mechanisms independent of the CD80 and CD86 costimulatory pathways, and (iii) decreased expression of CD86 and MHC II may modulate generation of optimal protective immunity.
Superantigen YPMa Exacerbates the Virulence ofYersinia pseudotuberculosisin Mice
Authors: Carnoy Christophe , Mullet Chantal , Müller-Alouf Heide , Leteurtre Emmanuelle , Simonet Michel
Published: July 2002

Yersinia pseudotuberculosis, a gram-negative bacterium responsible for enteric and systemic infection in humans, produces a superantigenic toxin designated YPMa (Y. pseudotuberculosis-derived mitogen). To assess the role of YPMa in the pathogenesis ofY. pseudotuberculosis, we constructed a superantigen-deficient mutant and compared its virulence in a mouse model of infection to the virulence of the wild-type strain. Determination of the survival rate after intravenous (i.v.) bacterial inoculation of OF1 mice clearly showed that inactivation ofypmA, encoding YPMa, reduced the virulence ofY. pseudotuberculosis. Mice infected i.v. with 104and 105wild-type bacteria died within 9 days, whereas mice infected with theypmAmutant survived 12 and 3 days longer, respectively. This decreased virulence of theypmAmutant strain was not due to an impaired colonization of the spleen, liver, or lungs. In contrast to i.v. challenge, bacterial inoculation by the intragastric (i.g.) route did not reveal any difference in virulence between wild-typeY. pseudotuberculosisand theypmAmutant since the 50% lethal doses were identical for both strains. Moreover, inactivation ofypmAgene did not affect the bacterial growth ofY. pseudotuberculosisin Peyer's patches, mesenteric lymph nodes (MLNs), and spleen after oral infection. Histological studies of spleen, liver, lungs, heart, Peyer's patches, and MLNs after i.v. or i.g. challenge with the wild type or theypmAmutant did not reveal any feature that can be specifically related to YPMa. Our data show that the superantigenic toxin YPMa contributes to the virulence ofY. pseudotuberculosisin systemic infection in mice.
Mutation ofwaaNReducesSalmonella entericaSerovar Typhimurium-Induced Enteritis and Net Secretion of Type III Secretion System 1-Dependent Proteins
Authors: Watson Patricia R. , Benmore Annette , Khan Shahid A. , Jones Philip W. , Maskell Duncan J. , Wallis Timothy S.
Published: July 2002

Mutation ofwaaN, a gene involved in lipid A biosynthesis, reduced enteropathogenic responses induced bySalmonella entericaserovar Typhimurium in bovine ligated ileal loops. However, the secretion of key virulence determinants was also reduced, and therefore the reduction in enteropathogenicity cannot be solely attributed to a reduction in biological activity of lipid A.
Interleukin-10 Has Different Effects on Proliferation ofListeria monocytogenesin Livers and Spleens of Mice
Authors: Samsom Janneke N. , Annema Akke , Geertsma Minke F. , Langermans Jan A. M. , Groeneveld Paul H. P. , de Heer Emile , van Furth Ralph
Published: July 2002

The aim of this study was to investigate the effect of interleukin-10 (IL-10) on the course ofListeria monocytogenesinfection in naive and immune mice. Treatment with IL-10 during the course of a primary infection significantly decreased the number of bacteria in the spleen and did not affect the number in the liver. During a secondary infection in immune mice treated with IL-10, the number of bacteria was significantly lower in the spleen but significantly higher in the liver in comparison to mock-treated immune mice. IL-10 treatment during a primaryListeriainfection decreased the concentration of gamma interferon (IFN-γ) in plasma and the toxoplasmastatic activity of macrophages, whereas it increased the percentage of mildly CD3-positive T cells in the spleen. During a secondary infection, the concentration of IFN-γ in plasma was decreased on day 1 but remained unaffected during later days of infection. From these results, we conclude that IL-10 has different effects on the proliferation ofL. monocytogenesin the spleen and liver during primary and secondaryListeriainfections.
Immunohistochemical Study of Lymphocyte Populations Infiltrating the Gastric Mucosa of Beagle Dogs Experimentally Infected withHelicobacter pylori
Authors: Rossi Giacomo , Fortuna Damiano , Pancotto Laura , Renzoni Giacomo , Taccini Ennio , Ghiara Paolo , Rappuoli Rino , Del Giudice Giuseppe
Published: July 2002

Experimental infection of beagle dogs withHelicobacter pyloriinduces recruitment to the gastric mucosae of neutrophils at early stages and later of mononuclear cells that organize into lymphoid follicles. These structures become macroscopically evident and consist of peripheral CD4+T lymphocytes and central CD21+B lymphocytes. Furthermore, transient expression of interleukin-8 (IL-8) parallels the presence of neutrophils in the gastric mucosae, whereas expression of IL-6 tends to persist chronically.
Nonimmune Interaction of the SfbI Protein ofStreptococcus pyogeneswith the Immunoglobulin G F(ab′)2Fragment
Authors: Medina Eva , Schulze Kai , Chhatwal Gursharan Singh , Guzmán Carlos Alberto
Published: July 2002

Fibronectin-binding protein I (SfbI) ofStreptococcus pyogenesbinds to mouse immunoglobulin G (IgG) but not to IgA or IgM in a nonimmune fashion. The fibronectin-binding domains of SfbI were responsible for this activity, which was targeted to the IgG F(ab′)2fragment. SfbI also binds to B cells but not to CD4+or CD8+lymphocytes.
Burkholderia pseudomallei
Induces Cell Fusion and Actin-Associated Membrane Protrusion: a Possible Mechanism for Cell-to-Cell Spreading
Authors: Kespichayawattana W. , Rattanachetkul S. , Wanun T. , Utaisincharoen P. , Sirisinha S.
Published: July 2002

Burkholderia pseudomallei
, a facultative intracellular bacterium, is the causative agent of a broad spectrum of diseases collectively known as melioidosis. Its ability to survive inside phagocytic and nonphagocytic cells and to induce multinucleated giant cell (MNGC) formation has been demonstrated. This study was designed to assess a possible mechanism(s) leading to this cellular change, using virulent and nonvirulent strains of
B. pseudomallei
to infect both phagocytic and nonphagocytic cell lines. We demonstrated that when the cells were labeled with two different cell markers (CMFDA or CMTMR), mixed, and then infected with
B. pseudomallei
, direct cell-to-cell fusion could be observed, leading to MNGC formation. Staining of the infected cells with rhodamine-conjugated phalloidin indicated that immediately after the infection, actin rearrangement into a comet tail appearance occurred, similar to that described earlier for other bacteria. The latter rearrangement led to the formation of bacterium-containing, actin-associated membrane protrusions which could lead to a direct cell-to-cell spreading of
B. pseudomallei
in the infected hosts. Results from 4′,6′-diamidine-2-phenylindole dihydrochloride (DAPI) nuclear staining, poly-ADP ribose polymerase cleavage, staining of infected cells for phosphatidylserine exposure with annexin V, and electrophoresis of the DNA extracted from these infected cells showed that
B. pseudomallei
could kill the host cells by inducing apoptosis in both phagocytic and nonphagocytic cells.
Prevotella intermedia
Stimulates Expansion of Vβ-Specific CD4
+
T Cells
Authors: Leung K.-P. , Torres Barbara A.
Published: July 2002

Recent evidence suggests that certain periodontal pathogens preferentially stimulate T cells expressing specific variable regions on the β chain (Vβ) of the T-cell receptor, which may indicate the presence of a superantigen. Superantigens are microbial proteins that activate large numbers of CD4
+
T cells in a Vβ-specific manner. The purpose of this study was to determine whether
Prevotella intermedia
, a putative periodontal pathogen, activates populations of specific Vβ on CD4
+
T cells. Among the bacterial strains tested,
P. intermedia
strain 17, a clinical isolate, induced the strongest proliferative response in peripheral blood mononuclear cells. Antibodies raised against whole cells of this organism blocked the proliferative activity.
P. intermedia
-induced proliferation was T-cell specific and required the presence of antigen-presenting cells. Flow cytometric analysis showed that CD4
+
T-cell subsets expressing Vβ8, Vβ12, and Vβ17 expanded in response to
P. intermedia
strain 17. The ability of
P. intermedia
to stimulate CD4
+
-T-cell proliferation was further supported by the production profiles of key T-cell cytokines, gamma interferon and interleukin-2. The data collectively suggest that certain strains of
P. intermedia
can activate Vβ-specific T cells in a manner similar to that of other known microbial superantigens.
Signaling via Interleukin-4 Receptor α Chain Is Required for Successful Vaccination against Schistosomiasis in BALB/c Mice
Authors: Mountford Adrian P. , Hogg Karen G. , Coulson Patricia S. , Brombacher Frank
Published: July 2002

Although protective immunity in C57BL/6 mice induced by a single dose of the radiation-attenuated schistosome vaccine is believed to be mediated by Th1-type immune responses, we here report that in BALB/c mice protection can also depend upon signaling via the interleukin-4 (IL-4) receptor which conventionally governs the development of Th2-type immune responses. We show that in BALB/c mice deficient for the IL-4 receptor α chain (IL-4Rα−/−), which are unresponsive to IL-4 and IL-13, vaccine-induced protection is abrogated compared with that in wild-type (WT) mice. In vaccinated IL-4Rα−/−mice, IL-12p40 production by cells from the skin exposure site was elevated, although gamma interferon (IFN-γ) production in draining lymphoid tissues was similar in WT and IL-4Rα−/−mice. Nevertheless, the effector response in IL-4Rα−/−mice was Th1 biased with elevated IFN-γ in the lungs and higher immunoglobulin G2a (IgG2a) and IgG2b titers but negligible quantities of Th2-associated IgG1 and IgE. Interestingly, levels of IL-4 were equivalent in WT and IL-4Rα−/−mice, indicating that Th2 responses were not dependent upon signaling by IL-4 or IL-13. No differences in the phenotype and composition of the pulmonary effector mechanism that might explain the failure to induce protection in IL-4Rα−/−mice were detected. However, passive transfer of partial protection to naive IL-4Rα−/−mice, using serum from vaccinated WT mice, indicates that Th2-associated antibodies such as IgG1 have a role in parasite elimination in BALB/c strain mice and that signaling via IL-4R can be an important factor in the generation of protection.
Synthesis and Surface Expression of CD14 by Human Endothelial Cells
Authors: Jersmann Hubertus P. A. , Hii Charles S. T. , Hodge Greg L. , Ferrante Antonio
Published: July 2002

Previous studies have reported that human vascular endothelial cells lack the membrane-bound lipopolysaccharide (LPS) receptor, CD14 (mCD14). By optimizing assay conditions, including the selection of anti-CD14 monoclonal antibody, we now demonstrate that human umbilical vein endothelial cells (HUVEC) express CD14 on the cell surface. Single-passage HUVEC showed approximately 20 times less expression of CD14 than monocytes. Interestingly, there was significant loss of surface CD14 expression with increasing numbers of culture passages. Evidence for synthesis of CD14 by HUVEC was provided by the finding thatl-[35S]methionine was incorporated into CD14. In addition, the expression of CD14 on HUVEC was upregulated by LPS, lysophosphatidic acid, and tissue culture supplements, and this upregulation was dependent on protein synthesis. Furthermore, the results imply that mCD14 is required for LPS-induced activation of endothelial cells in the absence of serum and that it acts in concert with serum factors (soluble CD14). Our results provide evidence that CD14 is expressed by endothelial cells and suggest that the previous inability to observe expression of this molecule has been due to culture and staining conditions. This finding has important implications for the understanding of the mechanisms by which LPS stimulates endothelial cells and the management of sepsis caused by gram-negative bacteria.
Molecular Cloning of aBacteroides caccaeTonB-Linked Outer Membrane Protein Identified by an Inflammatory Bowel Disease Marker Antibody
Authors: Wei Bo , Dalwadi Harnisha , Gordon Lynn K. , Landers Carol , Bruckner David , Targan Stephan R. , Braun Jonathan
Published: July 2002

Commensal enteric bacteria are a required pathogenic factor in inflammatory bowel disease (IBD), but the identity of the pertinent bacterial species is unresolved. Using an IBD-associated pANCA monoclonal antibody, a 100-kDa protein was recently characterized from an IBD clinical isolate ofBacteroides caccae(p2Lc3). In this study, consensus oligonucleotides were designed from 100-kDa peptides and used to identify a single-copy gene from the p2Lc3 genome. Sequence analysis of the genomic clone revealed a 2,844-bp (948 amino acid) open reading frame encoding features typical of the TonB-linked outer membrane protein family. This gene, termedompW,was detected by Southern analysis only inB. caccaeand was absent in other species ofBacteroidesand gram-negative coliforms. The closest homologues of OmpW included the outer membrane proteins SusC ofBacteroides thetaiotaomicronand RagA ofPorphyromonas gingivalis. Recombinant OmpW protein was immunoreactive with the monoclonal antibody, and serum anti-OmpW immunoglobulin A levels were elevated in a Crohn's disease patient subset. These findings suggest that OmpW may be a target of the IBD-associated immune response and reveal its structural relationship to a bacterial virulence factor ofP. gingivalisand periodontal disease.
Shiga Toxins Induce, Superinduce, and Stabilize a Variety of C-X-C Chemokine mRNAs in Intestinal Epithelial Cells, Resulting in Increased Chemokine Expression
Authors: Thorpe Cheleste M. , Smith Wendy E. , Hurley Bryan P. , Acheson David W. K.
Published: July 2002

Exposure of humans to Shiga toxins (Stxs) is a risk factor for hemolytic-uremic syndrome (HUS). Because Stx-producing
Escherichia coli
(STEC) is a noninvasive enteric pathogen, the extent to which Stxs can cross the host intestinal epithelium may affect the risk of developing HUS. We have previously shown that Stxs can induce and superinduce IL-8 mRNA and protein in intestinal epithelial cells (IECs) in vitro via a ribotoxic stress response. We used cytokine expression arrays to determine the effect of Stx1 on various C-X-C chemokine genes in IECs. We observed that Stx1 induces multiple C-X-C chemokines at the mRNA level, including interleukin-8 (IL-8), GRO-α, GRO-β, GRO-γ, and ENA-78. Like that of IL-8, GRO-α and ENA-78 mRNAs are both induced and superinduced by Stx1. Furthermore, Stx1 induces both IL-8 and GRO-α protein in a dose-response fashion, despite an overall inhibition in host cell protein synthesis. Stx1 treatment stabilizes both IL-8 and GRO-α mRNA. We conclude that Stxs are able to increase mRNA and protein levels of multiple C-X-C chemokines in IECs, with increased mRNA stability at least one mechanism involved. We hypothesize that ribotoxic stress is a pathway by which Stxs can alter host signal transduction in IECs, resulting in the production of multiple chemokine mRNAs, leading to increased expression of specific proteins. Taken together, these data suggest that exposing IECs to Stxs may stimulate a proinflammatory response, resulting in influx of acute inflammatory cells and thus contributing to the intestinal tissue damage seen in STEC infection.
HLA-B27 Expression Does Not Modulate IntracellularChlamydia trachomatisInfection of Cell Lines
Authors: Young J. L. , Smith L. , Matyszak M. K. , Gaston J. S. H.
Published: July 2002

Chlamydia trachomatisis an obligate intracellular pathogen. Infection of susceptible individuals with this bacterium can trigger the development of reactive arthritis, an acute inflammation that is associated with the expression of the class I major histocompatibility antigen, HLA-B27. Other facultative intracellular pathogens, such asYersiniaandSalmonellaspp., are also known triggers of reactive arthritis. Previous studies report conflicting results concerning whether the presence of HLA-B27 modulates the infection of cells with these enteric pathogens. In the present study, we have examined whether the expression of HLA-B27 can influence the infection of cell lines withC. trachomatisand also whether the replication of these bacteria is altered in HLA-B27-expressing cell lines. To do this, we have used a sensitive flow cytometric approach. We fixed and permeabilized cells and used fluorescein isothiocyanate-conjugated monoclonal antibody specific for chlamydia lipopolysaccharide to detect intracellular bacteria. The staining pattern obtained closely resembled the intracellular life cycle of chlamydia, with the appearance of brightly staining cells correlating to the microscopic detection of mature inclusion bodies. Moreover, since the percentage of cells that stained with the antibody was proportional to the infectious inoculum used, we were able to use the technique to quantitate the number of infectious organisms recoverable from infected cell lines. An important component of our study was the use of heparin to prevent reinfection of cells and thus enable the infection to be followed from a discrete time point. Our results suggest that HLA-B27 influences neither the infection nor replication ofC. trachomatisserovar L2 within cell lines. Consequently, the role of HLA-B27 in the pathogenesis of reactive arthritis may lie downstream of the invasion and replication stages of the triggering pathogenic infection.
Sequence Analysis of Four
Shigella boydii
O-Antigen Loci: Implication for
Escherichia coli
and
Shigella
Relationships
Authors: Wang Lei , Qu Wenjia , Reeves Peter R.
Published: July 2002

Shigella
strains are in reality clones of
Escherichia coli
and are believed to have emerged relatively recently (G. M. Pupo, R. Lan, and P. R. Reeves, Proc. Natl. Acad. Sci. USA 97:10567–10572, 2000). There are 33 O-antigen forms in these
Shigella
clones, of which 12 are identical to O antigens of other
E. coli
strains. We sequenced O-antigen gene clusters from
Shigella boydii
serotypes 4, 5, 6, and 9 and also studied the O53- and O79-antigen gene clusters of
E. coli
, encoding O antigens identical to those of
S. boydii
serotype 4 and
S. boydii
serotype 5, respectively. In both cases the
S. boydii
and
E. coli
O-antigen gene clusters have the same genes and organization. The clusters of both
S. boydii
6 and
S. boydii
9 O antigens have atypical features, with a functional insertion sequence and a
wzx
gene located in the orientation opposite to that of all other genes in
S. boydii
serotype 9 and an
rmlC
gene located away from other
rml
genes in
S. boydii
serotype 6. Sequences of O-antigen gene clusters from another three
Shigella
clones have been published, and two of them also have abnormal structures, with either the entire cluster or one gene being located on a plasmid in
Shigella sonnei
or
Shigella dysenteriae
, respectively. It appears that a high proportion of clusters coding for O antigens specific to
Shigella
clones have atypical features, perhaps indicating recent formation of these gene clusters.
Role of Glucan and Surface Protein BAD1 in Complement Activation byBlastomyces dermatitidisYeast
Authors: Zhang Mason X. , Brandhorst Tristan T. , Kozel Thomas R. , Klein Bruce S.
Published: July 2002

Our previous studies showed thatBlastomyces dermatitidisyeast activates the human complement system, leading to deposition of opsonic complement fragments onto the yeast surface. This report examines the influence of altered surface expression of glucan or BAD1 protein (formerly WI-1) on the yeast's ability to activate and bind C3. Compared to the wild type, a glucan-deficient mutant yeast delayed initiation of C3 deposition and reduced C3-binding capacity by 50%. Linkage of baker's-yeast β-glucan to the glucan-deficient yeast restored initial C3 deposition kinetics to the wild-type level and partially restored C3-binding capacity, suggesting that β-glucan is an initiator of complement activation and a C3 acceptor. The role of BAD1 inB. dermatitidisyeast-complement interaction was also assessed.BAD1knockout yeast initiated faster C3 deposition and increased C3-binding capacity compared to the wild-type yeast or aBAD1-reconstituted yeast, suggesting either a lack of an intrinsic ability in BAD1 or an inhibitory role of BAD1 in complement activation and binding. However, both complement activation and the capacity for C3 binding by the wild-type yeast were enhanced in normal human serum supplemented with an anti-BAD1 monoclonal antibody (MAb) or in immune sera from blastomycosis patients. Microscopic analysis revealed that more initial C3-binding sites were formed on yeast in the presence of both naturally occurring complement initiators and exogenous anti-BAD1 MAb, suggesting that anti-BAD1 antibody enhanced the ability ofB. dermatitidisyeast to interact with the host complement system. Thus, glucan and BAD1 have distinctly different regulatory effects on complement activation byB. dermatitidis.
Characterization ofChlamydia pneumoniaePersistence in HEp-2 Cells Treated with Gamma Interferon
Authors: Pantoja Laura G. , Miller Richard D. , Ramirez Julio A. , Molestina Robert E. , Summersgill James T.
Published: July 2002

Infection withChlamydia pneumoniaehas been implicated as a potential risk factor for atherosclerosis. This study demonstrated the effects of gamma interferon (IFN-γ)-mediated indoleamine 2,3-dioxygenase activity onC. pneumoniaepersistence in HEp-2 cells, inclusion morphology, and ultrastructure.C. pneumoniaereplication showed a dose-dependent decrease when treated with increasing concentrations of IFN-γ and a phenotypic switch resulting in a decrease in typical inclusions with an increase in smaller, less-dense atypical inclusions. Ultrastructural analysis of IFN-γ-treatedC. pneumoniaerevealed atypical inclusions containing large reticulatate-like aberrant bodies with no evidence of redifferentiation into elementary bodies.
Migration-Inhibitory Factor Gene-Deficient Mice Are Susceptible to CutaneousLeishmania majorInfection
Authors: Satoskar Abhay R. , Bozza Marcelo , Rodriguez Sosa Miriam , Lin Guoshing , David John R.
Published: July 2002

To determine the role of endogenous migration-inhibitory factor (MIF) in the development of protective immunity against cutaneous leishmaniasis, we analyzed the course of cutaneousLeishmania majorinfection in MIF gene-deficient mice (MIF−/−) and wild-type (MIF+/+) mice. Following cutaneousL. majorinfection, MIF−/−mice were susceptible to disease and developed significantly larger lesions and greater parasite burdens than MIF+/+mice. Interestingly, antigen-stimulated lymph node cells from MIF−/−mice produced more interleukin-4 (IL-4) and gamma interferon (IFN-γ) than those from MIF+/+mice, although the differences were statistically not significant. IFN-γ-activated resting peritoneal macrophages from MIF−/−mice showed impaired macrophage leishmanicidal activity and produced significantly lower levels of nitric oxide and superoxide in vitro. The macrophages from MIF−/−mice, however, produced much more IL-6 than macrophages from wild-type mice. These findings demonstrate that endogenous MIF plays an important role in the development of protective immunity againstL. majorin vivo. Furthermore, they indicate that the susceptibility of MIF−/−mice toL. majorinfection is due to impaired macrophage leishmanicidal activity rather than dysregulation of Th1 and Th2 responses.
Protective Cytotoxic T Lymphocyte Responses Induced by DNA Immunization against Immunodominant and Subdominant Epitopes ofListeria monocytogenesAre Noncompetitive
Authors: Yamada Takashi , Uchiyama Hiroshi , Nagata Toshi , Uchijima Masato , Suda Takafumi , Chida Kingo , Nakamura Hirotoshi , Koide Yukio
Published: July 2002

Taking advantage of the fact that plasmid DNA encoding a single cytotoxic T lymphocyte (CTL) epitope can induce CTLs, we examined the influence of T-cell responses to dominant epitopes on those to a subdominant epitope derived fromListeria monocytogenes. Our data suggest that interaction between T cells against dominant and subdominant epitopes does not operate in the generation of the hierarchy. Furthermore, we found that a single dominant epitope is sufficient for the induction of protective immunity.
Safety and Immunogenicity of Oral Inactivated Whole-Cell
Helicobacter pylori
Vaccine with Adjuvant among Volunteers with or without Subclinical Infection
Authors: Kotloff Karen L. , Sztein Marcelo B. , Wasserman Steven S. , Losonsky Genevieve A. , DiLorenzo Susan C. , Walker Richard I.
Published: July 2002

Helicobacter pylori
infection of the gastric mucosa can be found in approximately 50% of the world's population and is associated with a range of pathology, including peptic ulcer, atrophic gastritis, and gastric cancer. To explore immunization as a strategy for preventing and treating
H. pylori
-associated disease, we assessed the safety and immunogenicity in healthy adults of a formalin-inactivated, oral
H. pylori
whole-cell (HWC) vaccine, administered with or without mutant
Escherichia coli
heat-labile toxin (LT
R192G
) as a mucosal adjuvant. In a dose-response study, 23 subjects with or without
H. pylori
infection were vaccinated with either 2.5 × 10
6
HWC, 2.5 × 10
8
HWC, or 2.5 × 10
10
HWC, plus 25 μg of LT
R192G
. Thereafter, a randomized study was conducted in which 18
H. pylori
-infected subjects were assigned, in a double-blind fashion, to receive either 2.5 × 10
10
HWC plus placebo-adjuvant, placebo-vaccine plus 25 μg of LT
R192G
, placebo-vaccine plus placebo-adjuvant, or 2.5 × 10
10
HWC plus 25 μg of LT
R192G
. Diarrhea (six subjects), low-grade fever (five subjects), and vomiting (two subjects) were observed, usually after the first dose. Significant rises in geometric mean mucosal (fecal and salivary) anti-HWC immunoglobulin A antibodies occurred among
H. pylori
-infected and uninfected subjects following inoculation with 2.5 × 10
10
HWC plus 25 μg of LT
R192G
. Moreover, among
H. pylori
-negative volunteers, this regimen induced significant lymphoproliferative responses in 5 of 10 subjects and gamma interferon production responses to
H. pylori
sonicate in 7 of 10 subjects. There was no evidence that vaccination eradicated
H. pylori
in infected volunteers. These results suggest that it is possible to stimulate mucosal and systemic immune responses in humans to
H. pylori
antigens by using an HWC vaccine.
Calprotectin Expression In Vitro by Oral Epithelial Cells Confers Resistance to Infection byPorphyromonas gingivalis
Authors: Nisapakultorn K. , Ross K. F. , Herzberg M. C.
Published: July 2002

Calprotectin, an S100 calcium-binding protein with broad-spectrum antimicrobial activity in vitro, is expressed in neutrophils, monocytes, and gingival keratinocytes. In periodontitis, calprotectin appears upregulated and is detected at higher levels in gingival crevicular fluid and tissue specimens. How calprotectin contributes to the pathogenesis of periodontal diseases is unknown. To isolate the effects of calprotectin, a calprotectin-negative oral epithelial cell line was transfected with calprotectin genes to enable expression.Porphyromonas gingivaliswas permitted to bind and invade transfected cells expressing calprotectin and sham transfectants. Rates of invasion into both cell lines were compared using the antibiotic protection assay. Transfected cells expressing calprotectin showed 40 to 50% fewer internalizedP. gingivalisthan sham transfectants. Similarly, binding to calprotectin expressing cells was reduced approximately twofold at all time points (15, 30, 45, and 60 min) as estimated by immunofluorescence analysis. Independent of invasion, however, prolonged exposure toP. gingivalisinduced epithelial cell rounding and detachment from the substratum. These morphological changes were delayed, however, in cells expressing calprotectin. UsingP. gingivalisprotease-deficient mutants, we found that Arg-gingipain and Lys-gingipain contributed to epithelial cell rounding and detachment. In conclusion, expression of calprotectin appears to protect epithelial cells in culture against binding and invasion byP. gingivalis. In addition, cells expressing calprotectin are more resistant to detachment mediated by Arg-gingipain and Lys-gingipain. In periodontal disease, calprotectin may augment both the barrier protection and innate immune functions of the gingival epithelium to promote resistance toP. gingivalisinfection.
Role of
Enterococcus faecalis
Surface Protein Esp in the Pathogenesis of Ascending Urinary Tract Infection
Authors: Shankar Nathan , Lockatell C. Virginia , Baghdayan Arto S. , Drachenberg C. , Gilmore Michael S. , Johnson David E.
Published: July 2002

Enterococcus faecalis
bacteria isolated from patients with bacteremia, endocarditis, and urinary tract infections more frequently express the surface protein Esp than do fecal isolates. To assess the role of Esp in colonization and persistence of
E. faecalis
in an animal model of ascending urinary tract infection, we compared an Esp
+
strain of
E. faecalis
to its isogenic Esp-deficient mutant. Groups of CBA/J mice were challenged transurethrally with 10
8
CFU of either the parent or mutant strain, and bacteria in the urine, bladder, and kidneys were enumerated 5 days postinfection. Significantly higher numbers of bacteria were recovered from the bladder and urine of mice challenged with the parent strain than from the bladder and urine of mice challenged with the mutant. Colonization of the kidney, however, was not significantly different between the parent and mutant strains. Histopathological evaluations of kidney and bladder tissue done at 5 days postinfection did not show marked histopathological changes consistent with inflammation, mucosal hyperplasia, or apoptosis, and there was no observable difference between the mice challenged with the parent and those challenged with the mutant. We conclude that, while Esp does not influence histopathological changes associated with acute urinary tract infections, it contributes to colonization and persistence of
E. faecalis
at this site.
Plasmidic versus Insertional Cloning of Heterologous Genes inMycobacterium bovisBCG: Impact on In Vivo Antigen Persistence and Immune Responses
Authors: Méderlé I. , Bourguin I. , Ensergueix D. , Badell E. , Moniz-Peireira J. , Gicquel B. , Winter N.
Published: July 2002

Bivalent recombinant strains ofMycobacterium bovisBCG (rBCG) expressing the early regulatorynefand the structuralgag(p26) genes from the simian immunodeficiency virus (SIV) SIVmac251 were engineered so that both genes were cotranscribed from a synthetic operon. The expression cassette was cloned into a multicopy-replicating vector, and the expression levels of bothnefandgagin the bivalent rBCG(nef-gag) strain were found to be comparable to those of monovalent rBCG(nef) or rBCG(gag) strains. However, extrachromosomal cloning of thenef-gagoperon into a replicative plasmid resulted in strains of low genetic stability that rapidly lost the plasmid in vivo. Thus, thenef-gagoperon was inserted site specifically into the BCG chromosome by means of mycobacteriophage Ms6-derived vectors. The resulting integrative rBCG(nef-gag) strains showed very high genetic stability both in vitro and in vivo. The in vivo expression of the heterologous genes was much longer lived when the expression cassette was inserted into the BCG chromosome. In one of the strains obtained, integrative cloning did not reduce the expression levels of the genes even though a single copy was present. Accordingly, this strain induced cellular immune responses of the same magnitude as that of the replicative rBCG strain containing several copies of the genes.