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Biomedical subjects

Hermann Wagner

Publications and source records attributed to Hermann Wagner.

At least 91 records · Page 5Linked to original sources
PubMed2002

Cutting edge: myeloid differentiation factor 88 deficiency improves resistance against sepsis caused by polymicrobial infection.

Toll-like receptors (TLRs) are important for the activation of innate immune cells upon encounter of microbial pathogens. The present study investigated the potential roles of TLR2, TLR4, and the signaling protein myeloid differentiation factor 88 (MyD88) in polymicrobial septic peritonitis. Whereas both TLR2 and TLR4 were dispensable for host defense against septic peritonitis, MyD88-deficient mice were protected in this infection model. Recruitment of neutrophils to the septic focus and bacterial clearance were normal in MyD88-deficient mice. In contrast, the systemic inflammatory response was strongly attenuated in the absence of MyD88. Surprisingly, MyD88 deficiency did not alter cytokine and chemokine production in spleen, but markedly reduced the inflammatory response in liver and lung. Production of monocyte chemoattractant protein-1 and macrophage-inflammatory protein-1alpha was entirely independent of MyD88. These results imply a central role of MyD88 for the systemic immune pathology of polymicrobial sepsis and show that cytokine production in spleen and induction of certain chemokines are MyD88 independent.

Adaptor Proteins, Signal Transducing↗
PubMed2002

Caspase-9/-3 activation and apoptosis are induced in mouse macrophages upon ingestion and digestion of Escherichia coli bacteria.

A number of highly virulent, intracellular bacteria are known to induce cell death by apoptosis in infected host cells. In this work we demonstrate that phagocytosis of bacteria from the Escherichia coli laboratory strain K12 DH5alpha is a potent cell death stimulus for mouse macrophages. RAW264.7 mouse macrophages took up bacteria and digested them within 2-4 h as investigated with green fluorescent protein-expressing bacteria. No evidence of apoptosis was seen at 8 h postexposure, but at 24 h approximately 70% of macrophages displayed an apoptotic phenotype by a series of parameters. Apoptosis was blocked by inhibition of caspases or by forced expression of the apoptosis-inhibiting protein Bcl-2. Processing of caspase-3 and caspase-9 but not caspase-8 was seen suggesting that the mitochondrial branch of the apoptotic pathway was activated. Active effector caspases could be detected in two different assays. Because the adapter molecule myeloid differentiation factor 88 (MyD88) has been implicated in apoptosis, involvement of the Toll-like receptor pathway was investigated. In RAW264.7 cells, heat-treated bacteria were taken up poorly and failed to induce significant apoptosis. However, cell activation was almost identical between live and heat-inactivated bacteria as measured by extracellular signal-regulated kinase activation, generation of free radicals, and TNF secretion. Furthermore, primary bone marrow-derived macrophages from wild-type as well as from MyD88-deficient mice underwent apoptosis upon phagocytosis of bacteria. These results show that uptake and digestion of bacteria leads to MyD88-independent apoptosis in mouse macrophages. This form of cell death might have implications for the generation of the immune response.

Animals↗
PubMed2002

Postexposure prophylaxis against prion disease with a stimulator of innate immunity.

The absence of an immune response to prions--the infectious agents of scrapie, bovine spongiform encephalopathy (BSE), and Creutzfeldt-Jakob disease--might be related to the fact that these agents do not contain nucleic acids. We aimed to use CpG oligodeoxynucleotides, which have been shown to stimulate innate immunity, as a form of postexposure prophylaxis in mice. We inoculated healthy mice with brain homogenates from mice infected with the RML scrapie prion, and then injected CpG oligodeoxynucleotides. This postexposure prophylaxis with CpG oligodeoxynucleotides resulted in 38% longer survival times than treatment with saline (p<0.0001), or even longer after repeated application. The explanation for this finding remains to be elucidated, but the most likely is stimulation of TLR9-expressing cells of the innate immune system such as macrophages, monocytes, and dendritic cells. CpG oligodeoxynucleotides have not been shown to have adverse effects to human health and could therefore be considered as a therapeutic choice in postexposure prophylaxis.

Adjuvants, Immunologic↗
PubMed2002

The endoplasmic reticulum-resident heat shock protein Gp96 activates dendritic cells via the Toll-like receptor 2/4 pathway.

The heat shock protein Gp96 has been shown to induce specific immune responses. On one hand, this phenomenon is based on the specific interaction with CD91 that mediates endocytosis and results in major histocompatibility complex class I-restricted representation of the Gp96-associated peptides. On the other hand, Gp96 induces activation of professional antigen-presenting cells, resulting in the production of pro-inflammatory cytokines and up-regulation of costimulatory molecules by unknown mechanisms. In this study, we have analyzed the consequences of Gp96 interaction with cells expressing different Toll-like receptors (TLRs) and with bone marrow-derived dendritic cells from mice lacking functional TLR2 and/or TLR4 molecules. We find that the Gp96-TLR2/4 interaction results in activation of nuclear factor kappaB-driven reporter genes and mitogen- and stress-activated protein kinases and induces IkappaBalpha degradation. Bone marrow-derived dendritic cells of C3H/HeJ and more pronounced C3H/HeJ/TLR2(-/-) mice fail to respond to Gp96. Interestingly, activation of bone marrow-derived dendritic cells depends on endocytosis of Gp96 molecules. Our results provide, for the first time, the molecular basis for understanding the Gp96-mediated activation of antigen-presenting cells by describing the simultaneous stimulation of the innate and adaptive immune system. This feature explains the remarkable ability of Gp96 to induce specific immune responses against tumors and pathogens.

Animals↗
PubMed2002

The Helicobacter pylori blood group antigen-binding adhesin facilitates bacterial colonization and augments a nonspecific immune response.

Presence of the Helicobacter pylori adherence factor blood group Ag-binding adhesin (BabA; binding to Lewis(b) (Le(b))) is associated with ulcer disease, adenocarcinoma, and precancerous lesions. The importance of BabA for bacterial colonization and the inflammatory response is unknown. A total of 141 antral biopsies from H. pylori-infected patients were assessed in regard to the degree of granulocytic (G0 degrees--G3 degrees) and lymphocytic (L1 degrees--L3 degrees) infiltration. DNA genotypes of babA2 (the transcriptionally active gene of BabA), cagA, and vacAs1/2 were determined by PCR. Colonization density and Le(b) status on gastric epithelial cells were determined by immunohistochemistry. Real-time quantitative (TaqMan) RT-PCR determined mRNA expression of IL-8, TNF -alpha, and the Th1 markers IFN-gamma and the IL-12R beta2 chain. A total of 91% of infected patients were Le(b) positive. The vacAs1(+)/cagA(+) strains harboring babA2 showed significantly higher levels of granulocytic infiltration, bacterial colonization, and IL-8 mRNA than vacAs1(+)/cagA(+) strains lacking babA2. IL-8 mRNA and protein production by KATO III cells in vitro increased dose dependently with addition of different numbers of type 1 strains (G27 and 2808 strains, 0.1--20 bacteria/cell). The mRNA expression of TNF-alpha, IFN-gamma, and IL-12R beta2 was higher in H. pylori-positive patients than in controls, but it did not differ significantly between patients infected with different strain types. These data suggest that BabA facilitates colonization of H. pylori and thereby increases IL-8 response, resulting in enhanced mucosal inflammation. Infection with strains harboring BabA thereby augment a nonspecific immune response, whereas the Th1 response toward H. pylori appears to be independent of BabA, cytotoxin-associated gene A, or vacuolating cytotoxin.

Adhesins, Bacterial↗
PubMed2002

HSP70 as endogenous stimulus of the Toll/interleukin-1 receptor signal pathway.

Human heat-shock protein (HSP)70 activates innate immune cells and hence requires no additional adjuvants to render bound peptides immunogenic. Here we tested the assumption that endogenous HSP70 activates the Toll/IL-1 receptor signal pathway similar to HSP60 and pathogen-derived molecular patterns. We show that HSP70 induces interleukin-12 (IL-12) and endothelial cell-leukocyte adhesion molecule-1 (ELAM-1) promoters in macrophages and that this is controlled by MyD88 and TRAF6. Furthermore, HSP70 causes MyD88 relocalization and MyD88-deficient dendritic cells do not respond to HSP70 with proinflammatory cytokine production. Using the system of genetic complementation with Toll-like receptors (TLR) we found that TLR2 and TLR4 confer responsiveness to HSP70 in 293T fibroblasts. The expanding list of endogenous ligands able to activate the ancient Toll/IL-1 receptor signal pathway is in line with the "danger hypothesis" proposing that the innate immune system senses danger signals even if they originate from self.

Animals↗
PubMed2002

Generation of neutralizing mouse anti-mouse IL-18 antibodies for inhibition of inflammatory responses in vivo.

The proinflammatory cytokine IL-18 mediates IFN-gamma production as well as the induction of Th1 polarized immune responses in synergy with IL-12. In this study, we describe the production of isogeneic monoclonal antibodies (Mabs) directed against murine IL-18 (mIL-18). Immunization of IL-18-deficient mice with recombinant mIL-18 in the presence of CpG-oligodeoxynucleotides (CpG-ODN) and alum as adjuvant resulted in high anti-IL-18 serum titers. We could identify two Mabs, SK721-2 and SK113AE-4, which were able to bind to IL-18 and neutralize its IFN-gamma inducing effect in vitro with an IC(50) of 40-100 ng/ml. In vivo, LPS-induced IFN-gamma production was reduced by 60-85% following a single administration of Mabs SK113AE-4 or SK721-2. Since IL-18 is likely to be involved in the pathogenesis of inflammatory diseases such as rheumatoid arthritis or Crohn's disease, neutralizing mouse anti-mouse IL-18 Mabs have the potential to become valuable tools for the therapeutic exploration of long-term IL-18 blockade in vivo.

Adjuvants, Immunologic↗
PubMed2002

Bacterial CpG-DNA and lipopolysaccharides activate Toll-like receptors at distinct cellular compartments.

Recognition by innate immune cells of the pathogen associated molecular patterns (PAMP) lipopolysaccharide (LPS) from Gram-negative bacteria and bacterial CpG-DNA depends on Toll-like receptor4 (TLR4) and TLR9, respectively. To define differences in the response to these distinct PAMP we compared a key intracellular event, namely recruitment of myeloid differentiation marker 88 (MyD88) to the respective PAMP-initiated TLR signaling. Using MyD88-GFP fusion protein expressing macrophages we demonstrate that LPS and CpG-DNA trigger signaling from two different cellular locations: theformer at the cell membrane and the latter at the lysosomal compartment. While LPS does not require endocytosis to functionally associate with the membrane expressed TLR4/MD2 complex, internalization and endosomal maturation is conditional for CpG-DNA to activate TLR9. In support of these data TLR9 is not localized at the cell surface, but intracellularily. These data stress the need to characterize individual TLR at the very beginning of signal initiation in order to understand their diverse biological functions.

Adaptor Proteins, Signal Transducing↗
PubMed2002

CpG-DNA aided cross-presentation of soluble antigens by dendritic cells.

For cross-presentation immature dendritic cells (DC) require enhanced antigen (Ag) uptake and a maturation signal to prime for MHC class I-restricted CTL responses in vivo. While immunostimulatory CpG-DNA provides, via TLR9, the maturation signal, CpG-DNA linked to Ag augments cellular Ag uptake. In this study we show that CpG-DNA ovalbumin (OVA) conjugates trigger in vivo peptide-specific CTL responses at tenfold lower Ag doses compared to a mixture of CpG-DNA plus OVA. We provide evidence that CpG-DNA-OVA conjugates shift OVA uptake by immature DC from the presumably inefficient fluid phase pinocytosis to efficient DNA receptor-mediated endocytosis. Since the DNA-binding receptor mediating endocytosis lacks any sequence specificity, cellular uptake of OVA conjugated with either stimulatory or non-stimulatory oligonucleotides (ODN) is equally enhanced. As a consequence cross-linking of OVA with either stimulatory or non-stimulatory DNA yields, via enhanced OVA uptake, efficient generation and presentation of the dominant OVA-CTL epitope SIINFEKL. However, only stimulatory CpG-ODN cross-linked to OVA provide the DC maturation signal required to trigger robust primary CTL responses towards the cross-presented MHC class I complexed T cell epitope SIINFEKL. Our studies show that stimulatory CpG-ODN linked to Ag fulfill a dual role: enhancement of Ag uptake yielding efficient Ag cross-presentation by DC and in addition, their activation into professional DC.

Adjuvants, Immunologic↗
PubMed2002

Role of chlamydial heat shock protein 60 in the stimulation of innate immune cells by Chlamydia pneumoniae.

Chlamydia pneumoniae stimulates potently maturation of and cytokine secretion by bone marrow-derived dendritic cells (BMDDC). BMDDC responses depend mainly on Toll-like receptor (TLR)2 and to a minor extent on TLR4. We demonstrate here using C. pneumoniae in an infectious model with the replication-permissive epithelial cell line HEp2 that HSP60 is produced in substantial amounts in chlamydial inclusions during infection. Electron microscopy of chlamydial inclusions revealed that HSP60 was mainly associated with reticulate bodies, but was also located in between the different chlamydial developmental forms. Supernatants of permissive HEp2 cells infected with C. pneumoniae contained soluble chlamydial HSP60 as demonstrated by Western blotting and were able to stimulate BMDDC of wild-type mice. The stimulatory capacity of culture supernatants correlated with the presence of chlamydial HSP60. In contrast, BMDDC from TLR4-mutant mice crossed to TLR2-deficient mice were not stimulated by the culture supernatant, indicating that chlamydial HSP60 but not cytokines, possibly secreted by infected HEp2 cells, are responsible for the observed stimulation of BMDDC. Purified recombinant HSP60 from C. pneumoniae stimulated BMDDC in a TLR2- and TLR4-dependent fashion similar to the whole microorganism. In summary, these data suggest chlamydial HSP60 as an important mediator of inflammatory responses during infection with C. pneumoniae.

Animals↗
PubMed2002

Depth generalization from stereo to motion parallax in the owl.

Although many sources of three-dimensional information have been isolated and demonstrated to contribute independently, to depth vision in animal studies, it is not clear whether these distinct cues are perceived to be perceptually equivalent. Such ability is observed in humans and would seem to be advantageous for animals as well in coping with the often co-varying (or ambiguous) information about the layout of physical space. We introduce the expression primary-depth-cue equivalence to refer to the ability to perceive mutually consistent information about differences in depth from either stereopsis or motion-parallax. We found that owls trained to detect relative depth as a perceptual category (objects versus holes) when specified by binocular disparity alone (stereopsis), immediately transferred this discrimination to novel stimuli where the equivalent depth categories were available only through differences in motion information produced by head movements (observer-produced motion-parallax). Motion-parallax discrimination did occur under monocular viewing conditions and reliable performance depended heavily on the amplitude of side-to-side head movements. The presence of primary-depth-cue equivalence in the visual system of the owl provides further conformation of the hypothesis that neural systems evolved to detect differences in either disparity or motion information are likely to share similar processing mechanisms.

Animals↗
PubMed2002

Visual depth encoding in populations of neurons with localized receptive fields.

Stereopsis is the ability to perceive three-dimensional structure from disparities between the two-dimensional retinal images. Although disparity-sensitive neurons have been proposed as a neural representation of this ability many years ago, it is still difficult to link all qualities of stereopsis to properties of the neural correlate of binocular disparities. The present study wants to support efforts directed at closing the gap between electrophysiology and psychophysics. Populations of disparity-sensitive neurons in V1 were simulated using the energy-neuron model. Responses to different types of stimuli were evaluated with an efficient statistical estimator and related to psychophysical findings. The representation of disparity in simulated population responses appeared to be very robust. Small populations allowed good depth discrimination. Two types of energy neurons (phase- and position-type models) that are discussed as possible neural implementations of disparity-selectivity could be compared to each other. Phase-type coding was more robust and could explain a tendency towards zero disparity in degenerated stimuli and, for high-pass stimuli, exhibited the breakdown of disparity discrimination at a maximum disparity value. Contrast-inverted stereograms led to high variances in disparity representation, which is a possible explanation of the absence of depth percepts in large contrast-inverted stimuli. Our study suggests that nonlocal interactions destroy depth percepts in large contrast-inverted stereograms, although these percepts occur for smaller stimuli of the same class.

Action Potentials↗
PubMed2002

Human and mouse plasmacytoid dendritic cells.

By virtue of their enormous potential to produce type I interferons it is clear that plasmacytoid dendritic cells are major players in the host defense against viruses and various pathogens. Plasmacytoid dendritic cells were first identified in humans and very recently the mouse equivalents have been isolated. This review focuses, where possible, on a comparison between the cells in these two species. The mouse and human plasmacytoid DC are remarkably similar in surface phenotype and they display similar responses to pathogenic stimuli. The discovery of the mouse plasmacytoid dendritic cells has allowed for further biological properties of these cells to be investigated and it is now clear that they represent a long-lived family of cells that differentiate into novel dendritic cells upon microbiologic stimulation.

Animals↗
PubMed2002

Vaccination of mice against invasive aspergillosis with recombinant Aspergillus proteins and CpG oligodeoxynucleotides as adjuvants.

In a murine model of invasive pulmonary aspergillosis, dendritic cells (DCs) pulsed with Aspergillus antigens induced the activation of CD4(+) Th1 cells capable of conferring resistance to the infection. Here we show that the combined, local delivery of unmethylated CpG oligodeoxynucleotides (ODNs) and the Asp f 16 Aspergillus allergen resulted in the functional maturation and activation of airway DCs capable of inducing Th1 priming and resistance to the fungus. Therefore, ODNs act as a potent adjuvant for the vaccine-induced protection against the fungus by promoting dominant Th1 response to Aspergillus antigens and allergens.

Adjuvants, Immunologic↗
PubMed2002

Interactions between bacterial CpG-DNA and TLR9 bridge innate and adaptive immunity.

The astounding adjuvanticity and Th1-polarizing immunobiology of bacterial CpG-DNA and mimicking CpG-oligonucleotides continue to mirror promising therapeutic potential. The past year has witnessed some particularly impressive progress in knowledge of its molecular mode of action. Accordingly, CpG-DNA acts as a "pathogen-associated" molecular pattern that is recognized by TLR9 expressed, in particular, by dendritic cells. Interactions between CpG-DNA and TLR9 rapidly activate antigen-presenting dendritic cells through the ancient Toll/IL-1-receptor signaling pathway to upregulate co-stimulatory molecules and to produce Th1-polarizing cytokines, such as interleukin-12 and interleukin-18. Thus, interactions between CpG-DNA and TLR9 effectively bridge innate and acquired immunity.

Animals↗
PubMed2002

Reversible MHC multimer staining for functional isolation of T-cell populations and effective adoptive transfer.

Recently developed major histocompatibility complex (MHC) multimer technologies allow visualization and isolation of antigen-specific T cells. However, functional analysis and in vivo transfer of MHC multimer-stained cells is hampered by the persistence of T-cell receptor (TCR) MHC interactions and subsequently induced signaling events. As MHC monomers do not stably bind to TCRs, we postulated that targeted disassembly of multimers into MHC monomers would result in dissociation of surface-bound TCR ligands. We generated a new type of MHC multimers, which can be monomerized in the presence of a competitor, resulting in rapid loss of the staining reagent. Following dissociation, the T cells are phenotypically and functionally indistinguishable from untreated cells. This 'reversible' T-cell staining procedure, which maintains the specificity and sensitivity of MHC multimer staining while preserving the functional status of T lymphocytes, may be of broad benefit for ex vivo investigation of T-cell functions and clinical applications.

Adoptive Transfer↗
PubMed2002

Response properties of neurons in the core of the central nucleus of the inferior colliculus of the barn owl.

The central nucleus of the inferior colliculus (ICC) is particularly important for the processing of interaural time differences (ITDs). In the barn owl, neuronal best frequencies in a subnucleus of the ICC, the ICCcore, span the animal's entire hearing range (approximately equal to 200-10 000 Hz). This means that low-frequency ITD-sensitive ICCcore neurons in the owl can be directly compared to ITD-sensitive mammalian ICC neurons with similar best frequencies as well as to the high-frequency ITD-sensitive neurons usually studied in owls. This report represents a first attempt to systematically describe important physiological properties of ICCcore neurons in the barn owl, with particular attention to the low-frequency region (< 2 kHz). Responses were obtained from 133 neurons or small clusters of neurons; recording sites were confirmed by histological reconstruction of electrode tracks based on electrolytic lesions. Iso-intensity frequency response functions were typically approximately equal to 1 octave wide in the low-frequency range and approximately equal to 1/3 octave wide in the high-frequency range. Most neurons were ITD-tuned; both noise and pure tone stimuli yielded periodic ITD tuning curves with several equivalent response maxima. In most cases ITD tuning curves had a response peak within the barn owl's physiological ITD range. ITD tuning widths were inversely correlated with neuronal best frequency. None of the ICCcore neurons studied were sensitive to interaural level differences. Monaural inputs to ICCcore cells were typically binaurally balanced, i.e. they exhibited similar response thresholds, dynamic ranges, slopes and saturation levels, for both left and right ear monaural stimulation.

Acoustic Stimulation↗