Biomedical subjects

Stefan Ehlers

Publications and source records attributed to Stefan Ehlers.

At least 19 recordsLinked to original sources

PubMed2006

Helminth-derived products inhibit the development of allergic responses in mice.

RATIONALE: Epidemiological studies suggest that infections with helminths protect from the development of asthma. Supporting this view is our published finding that infection with Nippostrongylus brasiliensis decreased ovalbumin-induced Th2 responses in the lung of mice. OBJECTIVES: To evaluate if N. brasiliensis excretory-secretory products also prevent the development of asthma. METHODS: Mice were immunized with ovalbumin/alum intraperitoneally in the absence or presence of helminthic products and then challenged intranasally with ovalbumin. Six days later, we analyzed if the mice developed Th2 responses in the lung. MAIN RESULTS: The application of the helminthic products together with ovalbumin/alum during the sensitization period totally inhibited the development of eosinophilia and goblet cell metaplasia in the airways and also strongly reduced the development of airway hyperreactivity. Allergen-specific IgG1 and IgE serum levels were also strongly reduced. These findings correlated with decreased levels of IL-4 and IL-5 in the airways in product-treated animals. The suppressive effects on the development of allergic responses were independent of the presence of Toll-like receptors 2 and 4, IFN-gamma, and most important, IL-10. Interestingly, suppression was still observed when the helminthic products were heated or treated with proteinase K. Paradoxically, we found that strong helminth product-specific Th2 responses were induced in parallel with the inhibition of ovalbumin-specific responses. CONCLUSION: Our results suggest that helminths suppress the development of asthma by secreting substances that modulate allergic responses without affecting the generation of helminth-specific Th2 immunity. The identification of these products may lead to the design of novel therapeutic intervention strategies for the treatment of asthma.

Animals↗

PubMed2006

Mediator responses of alveolar macrophages and kinetics of mononuclear phagocyte subset recruitment during acute primary and secondary mycobacterial infections in the lungs of mice.

Little is known about the activation programme induced in alveolar macrophages upon phagocytosis of mycobacteria and the concomitant mononuclear phagocyte migratory responses that shape the acute phase of mycobacterial infection. Using high-speed cell sorting in conjunction with real-time RT-PCR analysis, we show that sorted alveolar macrophages of transgenic CX3CR1+/GFP mice infected with red fluorescent-labelled Mycobacterium bovis BCG exhibited weak transcriptional changes of lysosomal cathepsins B, L, D, K and S, whereas antimicrobial cathepsin G was strongly induced upon infection. Moreover, mRNA levels of pattern recognition receptors TLR2, TLR4 and NOD2 were downregulated as were neutrophil chemoattractants KC, MIP-2 and IP-10, whereas highly upregulated mRNA levels of the monocyte chemoattractant CCL2 were observed. M. bovis BCG infection of the mice elicited the alveolar accumulation of highly CX3CR1-positive alveolar dendritic cells but not neutrophils within the alveolar compartment, whereas no increased accumulation of CX3CR1high lung parenchymal dendritic cells (lung DC) or CX3CR1neg lung macrophages compared with controls was noted. In contrast, CX3CR1+/GFP mice previously immunized with M. bovis BCG rapidly responded with increased accumulations of both CX3CR1high alveolar and lung parenchymal DC and CX3CR1neg lung macrophages upon intratracheal M. bovis BCG challenge. Moreover, alveolar and lung macrophages and lung DC were found to contribute to early uptake of M. bovis BCG. Together, acute mycobacterial infection triggers both rapid changes in gene expression profiles in infected alveolar macrophages and a compartment-specific accumulation of mononuclear phagocyte subsets contributing to M. bovis BCG uptake in vivo.

Animals↗

PubMed2006

Characterization of a Mycobacterium tuberculosis mutant deficient in pH-sensing adenylate cyclase Rv1264.

Mycobacterium tuberculosis open reading frame Rv1264 encodes an adenylate cyclase that exhibits its highest enzymatic activity at an acidic pH of 6.0. This is the pH M. tuberculosis encounters in the phagosome. Consequently Rv1264 has been suggested to sense the phagosomal milieu resulting in adaption of M. tuberculosis to its intracellular niche. A targeted knock-out mutant deficient in Rv1264, however, exhibits wild-type virulence.

Adenylyl Cyclases↗

PubMed2006

Alternatively activated macrophages express the IL-27 receptor alpha chain WSX-1.

The interleukin (IL)-27 receptor-alpha WSX-1 is one component of the heterodimeric IL-27 receptor that is expressed on various cell types including macrophages. We previously demonstrated that IL-27 induces STAT-3 and is able to inhibit the production of pro-inflammatory cytokines in activated macrophages suggesting a novel feed-back mechanism by which IL-27 can modulate excessive inflammation. Because IL-4 receptor-alpha (IL-4Ralpha)-induced alternatively activated macrophages have also been described to attenuate pathological inflammatory immune responses, we analyzed the contribution of IL-27 in alternative macrophage activation. In the present study, like IL-10 and IL-4, IL-27 was found to suppress IL-12/23p40 production in activated bone marrow-derived macrophages. Whereas IL-10 induced the upregulation of the IL-4Ralpha on macrophages, receptor expression was not triggered by IL-27. In contrast to IL-4, IL-27 did not induce alternative macrophage activation but IL-4 strongly upregulated the expression of WSX-1 on macrophages and alternative macrophage activation enhanced IL-27-mediated signalling. We therefore conclude from our study that IL-10, IL-4 and IL-27 collaborate in modulating macrophage activation by successive upregulation of the IL-4Ralpha and WSX-1 on alternatively activated macrophages.

Animals↗

PubMed2006

The Wingless homolog WNT5A and its receptor Frizzled-5 regulate inflammatory responses of human mononuclear cells induced by microbial stimulation.

Microarray--assisted gene--expression screens of human macrophages revealed WNT5A, a homolog of Wingless, a key regulator of Drosophila melanogaster embryonic segmentation and patterning, to be consistently up-regulated following stimulation with different mycobacterial species and conserved bacterial structures. The expression of WNT5A required Toll-like receptor signaling and NF-kappaB activation, which identifies a novel induction pathway for a Wingless homolog. We show that human peripheral-blood mononuclear cells express the WNT5A receptor Frizzled-5 (FZD5). Both WNT5A and FZD5 also were detected in granulomatous lesions in the lungs of Mycobacterium tuberculosis-infected patients. Functional studies showed that WNT5A and FZD5 regulate the microbially induced interleukin-12 response of antigen-presenting cells and interferon-gamma production by mycobacterial antigen-stimulated T cells. Our findings implicate the evolutionarily conserved WNT/Frizzled signaling system in bridging innate and adaptive immunity to infections.

Antigen-Presenting Cells↗

PubMed2006

Selectin ligand-independent priming and maintenance of T cell immunity during airborne tuberculosis.

Immunity to Mycobacterium tuberculosis infection is critically dependent on the timely priming of T effector lymphocytes and their efficient recruitment to the site of mycobacterial implantation in the lung. E-, P-, and L-selectin counterreceptors control lymphocyte homing to lymph nodes and leukocyte trafficking to peripheral sites of acute inflammation, their adhesive function depending on fucosylation by fucosyltransferases (FucT) IV and VII. To address the relative importance of differentially glycosylated selectin counterreceptors for priming of T cell effector functions in a model of mycobacteria-induced granulomatous pulmonary inflammation, we used aerosol-borne M. tuberculosis to infect FucT-IV-/-, FucT-VII-/-, FucT-IV-/-/FucT-VII-/-, or wild-type control mice. In lymph nodes, infected FucT-IV-/-/FucT-VII-/- and, to a lesser extent, FucT-VII-/- mice had severely reduced numbers of T cells and reduced Ag-specific effector responses. By contrast, recruitment of activated T cells into the lungs was similar in all four groups of mice during infection and expression of T cell, and macrophage effector functions were only delayed in lungs of FucT-IV-/-/FucT-VII-/- mice. Importantly, lungs from all groups expressed CXCL13, CCL21, and CCL19 and displayed organized follicular neolymphoid structures after infection with M. tuberculosis, which suggests that the lung served as a selectin ligand-independent priming site for immune responses to mycobacterial infection. All FucT-deficient strains were fully capable of restricting M. tuberculosis growth in infected organs until at least 150 days postinfection. Our observations indicate that leukocyte recruitment functions dictated by FucT-IV and FucT-VII-dependent selectin ligand activities are not critical for inducing or maintaining T cell effector responses at levels necessary to control pulmonary tuberculosis.

Animals↗

PubMed2006

Interleukin-15 mediates protection against experimental tuberculosis: a role for NKG2D-dependent effector mechanisms of CD8+ T cells.

CD8+ T cells are involved in protection against Mycobacterium tuberculosis infection and represent a promising target for new vaccine strategies. Because IL-15 is important for the homeostasis of CD8+ T cells, we studied the immune response in IL-15-deficient mice during tuberculosis. In the absence of IL-15, CD8+ T cells failed to efficiently accumulate in draining lymph nodes and at the site of infection. The expression of antigen-specific effector functions, such as the production of interferon-gamma and cytotoxicity, were impaired in CD8+ T cells, but not CD4+ T cells, from IL-15-deficient mice. This defect was associated with an increased mortality of IL-15-deficient mice during the chronic phase of infection. The lectin-like stimulatory receptor natural killer group 2D (NKG2D) was up-regulated on CD8+ T cells only from wild-type mice, but not from IL-15-deficient mice. Mechanistically, blocking NKG2D function with an mAb inhibited M. tuberculosis-directed CD8+ T cell responses in vitro. We conclude that in addition to regulating the expansion of CD8+ T cells, IL-15 is also necessary for inducing effector mechanisms in CD8+ T cells that depend on NKG2D expression. Hence, our results implicate IL-15 and NKG2D as promising targets for modulating CD8+ T cell-mediated protection against tuberculosis.

Animals↗

PubMed2006

Genetically determined susceptibility to tuberculosis in mice causally involves accelerated and enhanced recruitment of granulocytes.

Classical twin studies and recent linkage analyses of African populations have revealed a potential involvement of host genetic factors in susceptibility or resistance to Mycobacterium tuberculosis infection. In order to identify the candidate genes involved and test their causal implication, we capitalized on the mouse model of tuberculosis, since inbred mouse strains also differ substantially in their susceptibility to infection. Two susceptible and two resistant mouse strains were aerogenically infected with 1,000 CFU of M. tuberculosis, and the regulation of gene expression was examined by Affymetrix GeneChip U74A array with total lung RNA 2 and 4 weeks postinfection. Four weeks after infection, 96 genes, many of which are involved in inflammatory cell recruitment and activation, were regulated in common. One hundred seven genes were differentially regulated in susceptible mouse strains, whereas 43 genes were differentially expressed only in resistant mice. Data mining revealed a bias towards the expression of genes involved in granulocyte pathophysiology in susceptible mice, such as an upregulation of those for the neutrophil chemoattractant LIX (CXCL5), interleukin 17 receptor, phosphoinositide kinase 3 delta, or gamma interferon-inducible protein 10. Following M. tuberculosis challenge in both airways or peritoneum, granulocytes were recruited significantly faster and at higher numbers in susceptible than in resistant mice. When granulocytes were efficiently depleted by either of two regimens at the onset of infection, only susceptible mice survived aerosol challenge with M. tuberculosis significantly longer than control mice. We conclude that initially enhanced recruitment of granulocytes contributes to susceptibility to tuberculosis.

Animals↗

PubMed2005

Towards a comprehensive view of the bacterial cell wall.

Direct in vivo visualization, in full atomic detail, of the microbial cell wall and its stress-bearing structural architecture remains one of the prime challenges in microbiology. In the meantime, molecular modeling can provide a framework for explaining and predicting mechanisms involved in morphogenesis, bacterial cell growth and cell division, during which the wall and its major structural component--murein--have to protect the cell from osmotic pressure and multiple tensile forces. Here, we illustrate why the scaffold concept of murein architecture provides a more comprehensive representation of bacterial cell wall physiology than previous models.

Bacteria↗

PubMed2005

Tumor necrosis factor and its blockade in granulomatous infections: differential modes of action of infliximab and etanercept?

Tumor necrosis factor (TNF) is a critical component of both the antibacterially protective and the inflammatory responses against infections, particularly infections with intracellularly viable microorganisms. It is, therefore, not surprising that some treatment regimens that target TNF function have resulted in an increase in complications associated with infections due to such pathogens as Mycobacterium tuberculosis, Listeria monocytogenes, and Histoplasma capsulatum; organized granuloma formation is required to keep such infections under control. However, treatment with anti-TNF monoclonal antibodies (i.e., infliximab) has been associated with a higher incidence of granulomatous infections than has treatment with a TNF receptor (TNFR) p75 immunoglobulin G-fusion construct (i.e., etanercept). Three hypotheses concerning the mode of action of these 2 agents that might explain this difference are discussed here: differential induction of apoptosis or lysis in membrane TNF-expressing macrophages and T cells, differential inhibition of signaling via TNFRp55 and TNFRp75, and different net neutralizing capacities resulting from different pharmacologic properties.

Antibodies, Monoclonal↗

PubMed2005

The IL-27 receptor chain WSX-1 differentially regulates antibacterial immunity and survival during experimental tuberculosis.

IL-12 is a potent inducer of IFN-gamma production and promotes a protective cell-mediated immune response after Mycobacterium tuberculosis infection. Recently, the IL-12-related cytokine IL-27 was discovered, and WSX-1 was identified as one component of the IL-27R complex. To determine the functional significance of IL-27/WSX-1 during tuberculosis, we analyzed the course of infection and the immune response in WSX-1-KO mice after aerosol infection with M. tuberculosis. In the absence of WSX-1, an increased production of the proinflammatory cytokines TNF and IL-12p40 resulted in elevated CD4+ T cell activation and IFN-gamma production, which enhanced macrophage effector functions and reduced bacterial loads. This is the first occasion of a selectively gene-deficient mouse strain showing higher levels of protective immunity against M. tuberculosis infection than wild-type mice. However, a concomitantly increased chronic inflammatory response also accelerated death of infected WSX-1-KO mice. In vitro, IL-27 induced STAT3 phosphorylation and inhibited TNF and IL-12 production in activated peritoneal macrophages, indicating a novel feedback mechanism by which IL-27 can modulate excessive inflammation. In conclusion, IL-27 both prevents optimal antimycobacterial protection and limits the pathological sequelae of chronic inflammation.

Animals↗

PubMed2005

Sarcoidosis is associated with a truncating splice site mutation in BTNL2.

Sarcoidosis is a polygenic immune disorder with predominant manifestation in the lung. Genome-wide linkage analysis previously indicated that the extended major histocompatibility locus on chromosome 6p was linked to susceptibility to sarcoidosis. Here, we carried out a systematic three-stage SNP scan of 16.4 Mb on chromosome 6p21 in as many as 947 independent cases of familial and sporadic sarcoidosis and found that a 15-kb segment of the gene butyrophilin-like 2 (BTNL2) was associated with the disease. The primary disease-associated variant (rs2076530; P(TDT) = 3 x 10(-6), P(case-control) = 1.1 x 10(-8); replication P(TDT) = 0.0018, P(case-control) = 1.8 x 10(-6)) represents a risk factor that is independent of variation in HLA-DRB1. BTNL2 is a member of the immunoglobulin superfamily and has been implicated as a costimulatory molecule involved in T-cell activation on the basis of its homology to B7-1. The G --> A transition constituting rs2076530 leads to the use of a cryptic splice site located 4 bp upstream of the affected wild-type donor site. Transcripts of the risk-associated allele have a premature stop in the spliced mRNA. The resulting protein lacks the C-terminal IgC domain and transmembrane helix, thereby disrupting the membrane localization of the protein, as shown in experiments using green fluorescent protein and V5 fusion proteins.

Bronchoalveolar Lavage↗

PubMed2005

Interleukin-12p40 mediates transient protection against Mycobacterium avium infection in the absence of interleukin-12.

Produced by macrophages and dendritic cells, interleukin (IL)-12 is composed of a p35 and a p40 subunit and promotes protection against intracellular pathogens through the development of interferon-gamma (IFNgamma) -producing T cells. The p40 subunit is also shared by the dimeric cytokines IL-12p40 homodimer and IL-23. In man, genetic defects in IL-12p40-mediated mechanisms are responsible for the familial occurrence of nontuberculous mycobacterial infections, the most common of which is infection with Mycobacterium avium. To experimentally differentiate the contribution of IL-12p40-containing cytokines in the outcome of M. avium infection, we studied wild-type, p35- and p35/p40 doubly deficient mice in an intravenous infection model which reflects many parameters of the disseminated infection in humans. Our study shows that in contrast to p35/p40 doubly deficient mice, p35-deficient mice mount a transient antibacterially protective response against M. avium although such animals were unable to produce detectable levels of IFNgamma or generate efficient granulomas. In conclusion, our results identify an antibacterial effector mechanism preserved in p35-deficient mice that is absent in mice devoid of p35 and p40. This phenotype probably reflects an IL-12p40-dependent effect on macrophage activation at the level of innate immunity.

Animals↗

PubMed2005

Tumor necrosis factor and granuloma biology: explaining the differential infection risk of etanercept and infliximab.

Several studies show that the risk of granulomatous infections following therapy with the anti-tumor necrosis factor (TNF) antibody infliximab is higher than after treatment with the soluble TNFRp75 immunoglobulin fusion construct etanercept. Therefore, despite sharing a common target, it is possible that the actual mode of action of the 2 biologicals differs in vivo. TNF is known to participate in the induction and maintenance of protective granulomas at multiple steps, and evidence supporting a differential inhibition of TNF bioactivity and signaling by the 2 drugs is discussed.

Animals↗

PubMed2005

Interaction of Rv1625c, a mycobacterial class IIIa adenylyl cyclase, with a mammalian congener.

The adenylyl cyclase Rv1625c from Mycobacterium tuberculosis codes for a protein with six transmembrane spans and a catalytic domain, i.e. it corresponds to one half of the pseudoheterodimeric mammalian adenylyl cyclases (ACs). Rv1625c is active as a homodimer. We investigated the role of the Rv1625c membrane domain and demonstrate that it efficiently dimerizes the protein resulting in a 7.5-fold drop in K(m) for ATP. Next, we generated a duplicated Rv1625c AC dimer by a head-to-tail concatenation. This produced an AC with a domain order exactly as the mammalian pseudoheterodimers. It displayed positive cooperativity and a 60% increase of v(max) compared with the Rv1625c monomer. Further, we probed the compatibility of mycobacterial and mammalian membrane domains. The second membrane anchor in the Rv1625c concatamer was replaced with membrane domain I or II of rabbit type V AC. The mycobacterial and either mammalian membrane domains are compatible with each other and both recombinant proteins are active. A M. tuberculosis Rv1625c knockout strain was assayed in a mouse infection model. In vitro growth characteristics and in vivo organ infection and mortality were unaltered in the knockout strain indicating that AC Rv1625c alone is not a virulence factor.

Adenylyl Cyclases↗

PubMed2005

Common and unique gene expression signatures of human macrophages in response to four strains of Mycobacterium avium that differ in their growth and persistence characteristics.

Classification of pathogenic species according to the distinct host transcriptional responses that they elicit may become a relevant tool for microarray-based diagnosis of infection. Individual strains of Mycobacterium avium, an opportunistic pathogen in humans, have previously been shown to differ in terms of growth and persistence. In order to cover a wide spectrum of virulence, we selected four M. avium isolates (2151SmO, 2151SmT, SE01, TMC724) that have distinct intramacrophage replication characteristics and cause differential activation in human macrophages. Following infection with each of these strains, the expression of 12,558 genes in human macrophages was systematically analyzed by microarray technology. Fifty genes (including genes encoding proinflammatory cytokines, chemokines, signaling, and adhesion molecules) were differentially expressed more than twofold in response to all of the M. avium isolates investigated and therefore constitute a common macrophage signature in response to M. avium. The magnitude of regulation of most of these genes was directly correlated with the host cell-activating capacity of the particular M. avium strain. The regulation of a number of genes not previously associated with mycobacterial infections was apparent; these genes included genes encoding lymphocyte antigen 64 and myosin X. In addition, individual response patterns typical for some M. avium isolates could be defined by the pronounced upregulation of interleukin-12p40 (IL-12p40) (in the case of 2151SmO) or the specific upregulation of SOCS-1 and IL-10 (in the case of SE01) in macrophages. TMC724, a strain of avian origin, could not be classified by any one of these schemes, possibly indicating the limits of pathogen categorization solely by immune response signatures.

Cells, Cultured↗

PubMed2005

Why does tumor necrosis factor targeted therapy reactivate tuberculosis?

Treatment of chronic inflammatory conditions, such as rheumatoid arthritis and Crohn's disease, with tumor necrosis factor (TNF) targeted biologics is associated with an increased risk of infectious complications, especially tuberculosis (TB). Clinical studies have revealed that monoclonal anti-TNF antibodies (e.g., infliximab) more frequently reactivate TB than a TNF receptor p75 immunoglobulin fusion construct (etanercept). Experimental studies in mice have shown TNF to be an essential component of protective granuloma formation. Based on these studies and the known pharmacological properties of the 2 prototype TNF targeted biologic agents, this review discusses 3 hypotheses that might explain the unpredicted differential risk of infectious complications: differential induction of target cell death, differential TNF receptor signaling, and differential net inhibition of TNF bioavailability.

Animals↗

PubMed2004

PAF-mediated pulmonary edema: a new role for acid sphingomyelinase and ceramide.

Platelet-activating factor (PAF) induces pulmonary edema and has a key role in acute lung injury (ALI). Here we show that PAF induces pulmonary edema through two mechanisms: acid sphingomyelinase (ASM)-dependent production of ceramide, and activation of the cyclooxygenase pathway. Agents that interfere with PAF-induced ceramide synthesis, such as steroids or the xanthogenate D609, attenuate pulmonary edema formation induced by PAF, endotoxin or acid instillation. Our results identify acid sphingomyelinase and ceramide as possible therapeutic targets in acute lung injury.

Animals↗