Biomedical subjects

M Röllinghoff

Publications and source records attributed to M Röllinghoff.

At least 37 records · Page 2Linked to original sources

PubMed2000

Fibroblasts as host cells in latent leishmaniosis.

Intracellular parasites are known to persist lifelong in mammalian hosts after the clinical cure of the disease, but the mechanisms of persistence are poorly understood. Here, we show by confocal laser microscopy that in the draining lymph nodes of mice that had healed a cutaneous infection with Leishmania major, 40% of the persisting parasites were associated with fibroblasts forming the reticular meshwork of the lymph nodes. In vitro, both promastigotes and amastigotes of L. major infected primary skin or lymph node fibroblasts. Compared with macrophages, cytokine-activated fibroblasts had a reduced ability to express type 2 nitric oxide synthase and to kill intracellular L. major. These data identify fibroblasts as an important host cell for Leishmania during the chronic phase of infection and suggest that they might serve as safe targets for the parasites in clinically latent disease.

Animals↗

PubMed2000

Tumor necrosis factor sustains the generalized lymphoproliferative disorder (gld) phenotype.

Tumor necrosis factor (TNF) and Fas ligand (FasL) play major roles in the homeostasis of the peripheral immune system. This becomes dramatically obvious in the absence of a functional FasL. Mice with such a deficiency develop a profound lymphadenopathy, splenomegaly, hypergammaglobulinemia, and strain-dependent systemic autoimmune disease, and succumb to premature death. It is consequently termed generalized lymphoproliferative disorder (gld). By contrast, TNF deficiency alone does not result in a striking phenotype. Thus, we sought to determine what role TNF might play in contributing to the gld phenotype by creating C57BL/6.gld.TNF(-/-) mice. Contrary to the expected outcome, mice deficient for both FasL and TNF had a substantially milder gld phenotype with regard to mortality, lymphoaccumulation, germinal center formation, and hypergammaglobulinemia. To confirm these data in a strain highly permissive for the phenotype, C3H/HeJ.gld and C3H.HeJ.lpr mice were treated with a TNF-specific monoclonal antibody. This transient neutralization of TNF also resulted in a significantly attenuated lymphoproliferative phenotype. We conclude that TNF is necessary for the full manifestation of the lymphoproliferative disorder, in particular playing a critical role in lymphoaccumulation. Most importantly, absence of TNF protects gld mice against premature death.

Animals↗

PubMed2000

Regulation of type 2 nitric oxide synthase by type 1 interferons in macrophages infected with Leishmania major.

We recently reported that the infection of macrophages with Leishmania major led to the release of type 1 interferons (IFN-alpha /beta ). Moreover, at day 1 of infection of mice with L. major, IFN-alpha /beta was required for the expression of type 2 (inducible) NO synthase (NOS2 or iNOS) which, however, was restricted to a few macrophages in the dermis. Here, we further characterized the regulation of NOS2 by IFN-alpha /beta. Macrophages that were either simultaneously or sequentially exposed to L. major promastigotes and IFN-alpha /beta expressed NOS2 and anti-leishmanial activity. In contrast, when high amounts of IFN-alpha /beta were used or when IFN-alpha /beta was added to the macrophages 2 h prior to the parasites, almost no induction of NOS2 was observed. After pretreatment with IFN-alpha /beta, tyrosine phosphorylation and nuclear DNA binding of Stat1alpha, the degradation of the NF-kappaB inhibitor (IkappaBalpha and beta), and the nuclear translocation of NF-kappaB were strongly impaired compared with macrophages exposed to IFN-alpha /beta and L. major simultaneously. Thus, IFN-alpha /beta exerts agonistic or antagonistic effects on the expression of NOS2 in macrophages infected with a microbial pathogen, depending on the sequence of the stimuli and the amount of IFN-alpha /beta added. The limited number of NOS2-positive macrophages at day 1 of infection in vivo might result from a blockage of non-infected macrophages by IFN-alpha /beta that is released by neighboring infected cells.

Animals↗

PubMed2000

Biologic functions and signaling of the interleukin-4 receptor complexes.

IL-4 is a pleiotropic cytokine which plays a pivotal role in shaping immune responses. The effects of IL-4 are mediated after binding to high affinity receptor complexes present on hematopoietic as well as non-hematopoietic cells. This review will summarize the current knowledge on the molecular structure of the different types of IL-4 receptor (IL-4R) complexes as well as the signal transduction mechanisms induced by IL-4 leading to cellular proliferation and / or gene activation. IL-4 effects are modulated by soluble forms of the respective receptor molecules which are produced by several immune cells in a regulated manner. The biological impact of recently described IL-4R allotypes of mice and humans as well as the results of studies with IL-4R knockout mice will be particularly emphasized.

Animals↗

PubMed2000

Reactive oxygen and reactive nitrogen intermediates in innate and specific immunity.

Nitric oxide, nitric oxide derivatives and reactive oxygen intermediates are toxic molecules of the immune system which contribute to the control of microbial pathogens and tumors. There is recent evidence for additional functions of these oxygen metabolites in innate and adaptive immunity; these functions include the modulation of the cytokine response of lymphocytes and the regulation of immune cell apoptosis, as well as immunodeviating effects. Components of several signal transduction pathways have been identified as intracellular targets for reactive nitrogen and oxygen intermediates.

Anti-Infective Agents↗

PubMed2000

The role of nitric oxide in innate immunity.

Type 2 nitric oxide synthase (iNOS or NOS2) was originally described as an enzyme that is expressed in activated macrophages, generates nitric oxide (NO) from the amino acid L-arginine, and thereby contributes to the control of replication or killing of intracellular microbial pathogens. Since interferon (IFN)-gamma is the key cytokine for the induction of NOS2 in macrophages and the prototypic product of type 1 T-helper cells, high-level expression of NOS2 has been regarded to be mostly restricted to the adaptive phase of the immune response. In this review, we summarize data that demonstrate a prominent role of NOS2/NO also during innate immunity. During the early phase of infection with the intracellular pathogen Leishmania major, focally expressed NOS2/NO not only exerts antimicrobial activities but also controls the function of natural killer cells and the expression of cytokines such as IFN-gamma or transforming growth factor-beta. Some of these effects result from the function of NOS2/NO as an indispensable co-factor for the activation of Tyk2 kinase and, thus, for interleukin-12 and IFN-alpha/beta signaling in natural killer cells.

Animals↗

PubMed2000

The TEA/ATTS transcription factor CaTec1p regulates hyphal development and virulence in Candida albicans.

The temporal and spatial expression of stage-specific genes during morphological development of fungi and higher eukaryotes is controlled by transcription factors. In this study, we report the cloning and functional analysis of the Candida albicans TEC1 (CaTEC1) gene, a new member of the TEA/ATTS family of transcription factors that regulates C. albicans virulence. The promoters of the type 4, 5 and 6 proteinase isogenes (SAP4-6) contain repetitive TEA/ATTS consensus sequence motifs. This finding suggests a possible role for a homologue of Saccharomyces cerevisiae TEC1 during the activation of proteinase gene expression in C. albicans. CaTEC1 is predominantly expressed in the hyphal form of C. albicans. In vitro, serum-induced hyphal formation as well as evasion from MPhi after phagocytosis is suppressed in catec1/catec1 mutant cells. Furthermore, expression of the proteinase isogenes SAP4-6 is no longer inducible in these mutant cells. The deletion of the CaTEC1 gene attenuates virulence of C. albicans in a systemic model of murine candidiasis, although both mutant and revertant cells that were prepared from infected tissues or the vaginal mucosa grew in a hyphal morphology in vivo. CaTEC1 complements the pseudohyphal and invasive growth defect of haploid and diploid S. cerevisiae tec1/tec1 mutant cells and strongly activates the promoter of FLO11, a gene required for pseudohyphal growth. This study provides the first evidence pointing to an essential role for a member of the TEA/ATTS transcription factor family that had so far only been ascribed to function during development as a virulence regulator in microbial pathogenesis.

Animals↗

PubMed2000

Repression of hyphal proteinase expression by the mitogen-activated protein (MAP) kinase phosphatase Cpp1p of Candida albicans is independent of the MAP kinase Cek1p.

Cpp1p is a putative mitogen-activated protein (MAP) kinase phosphatase that suppresses Candida albicans hyphal formation at 25 degrees C through its probable substrate, the Cek1p filamentation MAP kinase. Here we report that expression of the serum-induced genes SAP4-6 and HYR1 increased several fold in hyphal forms of a cpp1/cpp1 null mutant, while the rate and extent of hyphal development up to 5 h were normal. Therefore, we provide evidence that Cpp1p represses hyphal gene expression by acting through a Cek1p-independent mechanism. SAP4-6 and HYR1 transcripts were undetectable in a null mutant of another key regulator of filamentation, Efg1p; thus, Efg1p and Cpp1p oppose each other during the expression of these genes in hyphal forms.

Aspartic Acid Endopeptidases↗

PubMed1999

Requirement for type 2 NO synthase for IL-12 signaling in innate immunity.

Interleukin-12 (IL-12) and type 2 NO synthase (NOS2) are crucial for defense against bacterial and parasitic pathogens, but their relationship in innate immunity is unknown. In the absence of NOS2 activity, IL-12 was unable to prevent spreading of Leishmania parasites, did not stimulate natural killer (NK) cells for cytotoxicity or interferon-gamma (IFN-gamma) release, and failed to activate Tyk2 kinase and to tyrosine phosphorylate Stat4 (the central signal transducer of IL-12) in NK cells. Activation of Tyk2 in NK cells by IFN-alpha/beta also required NOS2. Thus, NOS2-derived NO is a prerequisite for cytokine signaling and function in innate immunity.

Animals↗

PubMed1999

Inhibition of inducible nitric oxide synthase exacerbates chronic cerebral toxoplasmosis in Toxoplasma gondii-susceptible C57BL/6 mice but does not reactivate the latent disease in T. gondii-resistant BALB/c mice.

Infection of C57BL/6 mice with Toxoplasma gondii leads to progressive and ultimately fatal chronic Toxoplasma encephalitis (TE). Genetic deletion or inhibition of inducible nitric oxide synthase (iNOS) from the beginning of infection increased the number of T. gondii cysts in the brain and markedly reduced the time-to-death in this mouse strain. In the present study, we addressed whether iNOS also contributes to the control of intracerebral parasites in a clinically stable latent infection that develops in T. gondii-resistant BALB/c mice after resolution of the acute phase of TE. iNOS was expressed in the inflammatory cerebral infiltrates of latently infected BALB/c mice, but the number of iNOS+ cells was significantly lower than in the brains of chronically infected T. gondii-susceptible C57BL/6 mice. In BALB/c mice with latent TE (> 30 days of infection), treatment with the iNOS inhibitors L-N6-iminoethyl-lysine or L-nitroarginine-methylester for < or = 40 days did not result in an increase of the intracerebral parasitic load and a reactivation of the disease, despite the presence of iNOS-suppressive inhibitor levels in the brain. However, L-nitroarginine-methylester treatment had remarkably toxic effects and induced a severe wasting syndrome with high mortality. In contrast to BALB/c mice, L-N6-iminoethyl-lysine treatment rapidly exacerbated the already established chronic TE of C57BL/6 mice. Thus, the containment of latent toxoplasms in T. gondii-resistant BALB/c mice is independent of iNOS, whereas the temporary control of intracerebral parasites in T. gondii-susceptible C57BL/6 mice with chronic TE requires iNOS activity.

Administration, Oral↗

PubMed1999

Human anti-FcepsilonRIalpha autoantibodies isolated from healthy donors cross-react with tetanus toxoid.

Natural antibodies (Ab) reacting with self antigens have been shown to be present in all individuals. These autoantibodies (auto-Ab) can be either pathogenic or non-pathogenic. Auto-Ab reacting with the alpha-subunit of the high-affinity receptor for IgE (FcepsilonRIalpha) have been implicated in the pathogenesis of a subset of patients with chronic idiopathic urticaria (CIU). Intravenous immunoglobulin (IVIg) preparations have been used with variable clinical benefit in the treatment of these patients. Here we show that anti-FcepsilonRIalpha auto-Ab are present in a therapeutic IVIg preparation as well as in atopic and chronic urticaria patients and healthy individuals. We affinity-purified the anti-FcepsilonRIalpha Ab from an IVIg preparation using recombinant FcepsilonRIalpha. Interestingly, these anti-FcepsilonRIalpha auto-Ab showed no evidence of histamine release but strongly cross-reacted with an external antigen, tetanus toxoid (TTd) with a higher affinity for TTd than for the FcepsilonRIalpha. Since the cross-reacting Ab are non-anaphylactogenic, there is no evidence that TTd immunization may contribute to the pathogenesis of CIU. However, our results may indicate that the anti-FcepsilonRIalpha auto-Ab belong to the natural Ab and serve as the parental Ab for some anti-TTd Ab.

Animals↗

PubMed1999

Analysis of the Yersinia enterocolitica 0:8 V antigen for cross protectivity.

The plasmid encoded V antigen (Vag) of pathogenic Yersinia spp. is a major virulence factor as well as a protective immunogen. Recently, two main types of Vag, represented by either Yersinia enterocolitica 0:8 or Yersinia pseudotuberculosis, have been identified and it has been suggested, that antibodies generated against one type are unable to protect against Yersinia spp. carrying the other type. By using a recombinant Vag (rVagHis) of the Y. enterocolitica 0:8 type we show here, that actively immunized mice were completely protected against challenge with both, Y. enterocolitica 0:8 and Y. pseudotuberculosis serotype III. In addition, passive protection was possible with polyclonal rabbit anti-rVagHisIgG. However, while a single antibody dose (200 microgramg) was sufficient to protect against challenge with Y. enterocolitica 0:8, repetitive injections at intervals of 2 to 3 days were needed to protect against challenge with Y. pseudotuberculosis III. The apparent difference in protection correlated with a rapid disappearance of anti-rVagHisIgG from the circulation by days 3 to 4. The data therefore indicate, that expression of distinct types of Vag by Yersinia spp. does not necessarily exclude immunoprotection in mice immunized with the other type of Vag. It rather appears, that differences in immunoprotection between Yersinia species relate to the amount of cross-protective antibody. Finally, as revealed by the lack of complement-mediated killing and the lack of immunostaining of Yersiniae with anti-rVagHisantibodies, evidence is provided to indicate that immunoprotection does not occur via opsonisation or complement lysis.

Animals↗

PubMed1999

How do protozoan parasites survive inside macrophages?

During infections with intracellular microbes, macrophages have two roles. On the one hand, they are important effector cells for the control and killing of intracellular bacteria and protozoan parasites by oxidative and non-oxidative mechanisms. On the other hand, macrophages may also serve as long-term host cells that facilitate the replication and survival of the pathogens, for example, by protecting them against toxic components of the extracellular milieu. In this review, Christian Bogdan and Martin Röllinghoff summarize some of the more recently discovered mechanisms by which intracellular protozoan parasites, such as Leishmania spp, Trypanosoma cruzi and Toxoplasma gondii, manage to exploit macrophages as safe target cells.

Animals↗

PubMed1999

Prevalence of Borrelia burgdorferi and granulocytic and monocytic ehrlichiae in Ixodes ricinus ticks from southern Germany.

A total of 287 adult Ixodes ricinus ticks, collected in two regions of southern Germany (Frankonia and Baden-Württemberg) where Borrelia burgdorferi infections are known to be endemic, were examined for the presence of 16S ribosomal DNA specific for the Ehrlichia phagocytophila genogroup, E. chaffeensis, E. canis, and B. burgdorferi by nested PCR. Totals of 2.2% (6 of 275) and 21.8% (65 of 275) of the ticks were positive for the E. phagocytophila genogroup and B. burgdorferi, respectively. Two ticks (0.7%) were coinfected with both bacteria. Of 12 engorged I. ricinus ticks collected from two deer, 8 (67%) were positive for the E. phagocytophila genogroup and one (8%) was positive for B. burgdorferi. There was no evidence of infection with E. canis or E. chaffeensis in the investigated tick population. The nucleotide sequences of the 546-bp Ehrlichia PCR products differed at one or two positions from the original sequence of the human granulocytic ehrlichiosis (HGE) agent (S.-M. Chen, J. S. Dumler, J. S. Bakken, and D. H. Walker, J. Clin. Microbiol. 32:589-595, 1994). Three groups of sequence variants were detected; two of these were known to occur in other areas in Europe or the United States, whereas one has not been reported before. Thus, in the German I. ricinus tick population closely related granulocytic ehrlichiae are prevalent, which might represent variants of E. phagocytophila or the HGE agent.

Animals↗

PubMed1999

Quantitative detection of Borrelia burgdorferi by real-time PCR.

Currently, no easy and reliable methods allowing for the quantification of Borrelia burgdorferi in tissues of infected humans or animals are available. Due to the lack of suitable assays to detect B. burgdorferi CFU and the qualitative nature of the currently performed PCR assays, we decided to exploit the recently developed real-time PCR. This technology measures the release of fluorescent oligonucleotides during the PCR. Flagellin of B. burgdorferi was chosen as the target sequence. A linear quantitative detection range of 5 logs with a calculated detection limit of one to three spirochetes per assay reaction mixture was observed. The fact that no signals were obtained with closely related organisms such as Borrelia hermsii argues for a high specificity of this newly developed method. A similar method was developed to quantify mouse actin genomic sequences to allow for the standardization of spirochete load. The specificity and sensitivity of the B. burgdorferi and the actin real-time PCR were not altered when samples were spiked with mouse cells or spirochetes, respectively. To evaluate the applicability of the real-time PCR, we used the mouse model of Lyme disease. The fate of B. burgdorferi was monitored in different tissues from inbred mice and from mice treated with antibiotics. Susceptible C3H/HeJ mice had markedly higher burdens of bacterial DNA than resistant BALB/c mice, and penicillin G treatment significantly reduced the numbers of spirochetes. Since these results show a close correlation between clinical symptoms and bacterial burden of tissues, we are currently analyzing human biopsy specimens to evaluate the real-time PCR in a diagnostic setting.

Animals↗

PubMed1998

Specific antagonism of type I IL-4 receptor with a mutated form of murine IL-4.

IL-4 is a pleiotropic cytokine that is essential for the differentiation of Th2 cells and is critically involved in the pathogenesis of certain infectious and allergic diseases. We have produced and functionally characterized a mutant of murine IL-4 (IL-4.Y119D) as a potential antagonist of IL-4. The analysis of IL-4R binding revealed no differences between wild-type and mutated IL-4. Despite this finding, IL-4.Y119D was unable to induce proliferation of several IL-4-responsive T cell lines mediated via the type I IL-4R (IL-4Ralpha/common gamma chain (gamma c chain)) and specifically inhibited the proliferative effect of wild-type IL-4. In contrast, with IL-4.Y119D we found induction of MHC class II and CD23 molecules on resting splenic B cells as well as proliferation of B9 plasmocytoma cells. In addition, IL-4.Y119D induced mRNA for soluble IL-4R, leading to the release of soluble IL-4R protein by spleen cells. In macrophages, mutated IL-4 in combination with IFN-gamma induced TNF-alpha-dependent killing of Leishmania major parasites such as wild-type IL-4. The agonistic effects of IL-4.Y119D were observed on cells expressing the IL-13R alpha-chain, including an IL-13R alpha-chain transfected T cell line, but were absent in T cells that lack this molecule, indicating that IL-4.Y119D conveys its activity via the type II IL-4R (IL-4Ralpha/IL-13Ralpha). The described IL-4 mutant, therefore, represents a new tool to use in dissecting different IL-4 functions that are mediated by either type I or type II IL-4R complexes.

Amino Acid Substitution↗

PubMed1998

Migration inhibitory factor induces killing of Leishmania major by macrophages: dependence on reactive nitrogen intermediates and endogenous TNF-alpha.

Macrophage migration inhibitory factor (MIF) is a product of activated T cells, anterior pituitary cells, and macrophages. MIF plays an important role in LPS-induced shock and delayed-type hypersensitivity. Furthermore, MIF exhibits a proinflammatory spectrum of action, promoting TNF-alpha production by macrophages, and counter-regulates glucocorticoid suppression of cytokine production. Here, we report that purified recombinant MIF activates murine macrophages to kill Leishmania major, with maximal effects at concentrations above 1 microg/ml. This MIF-mediated activation is specific, since it can be blocked completely by anti-MIF mAb. The MIF-mediated activation is dependent on TNF-alpha produced endogenously by macrophages, because the administration of anti-TNF-alpha antiserum markedly reduced the MIF effect. No MIF-mediated activation was observed in macrophages derived from TNF receptor p55 knockout mice, thus demonstrating the requirement of the smaller TNF receptor molecule for autocrine TNF-alpha signaling. A highly specific inhibitor of the inducible nitric oxide synthase (iNOS), L-N6-(1-iminoethyl)lysine, dihydrochloride, also inhibited the action of MIF, suggesting an important role for iNOS in the antiparasitic properties of MIF. In line with this, no MIF-mediated activation was detected analyzing macrophages derived from iNOS-deficient mice. The effect of MIF was blocked completely by the macrophage-deactivating cytokines IL-10, IL-13, and TGF-beta. Finally, the expression of MIF mRNA and protein was up-regulated in lymph nodes of mice during the first week after infection with L. major. MIF therefore represents a cytokine involved not only in the recruitment of proinflammatory cells during infection but also in the complex regulation of the antimicrobial activity of these cells.

Animals↗

PubMed1998

Costimulation via TCR and IL-1 receptor reveals a novel IL-1alpha-mediated autocrine pathway of Th2 cell proliferation.

Previous studies have shown that triggering of Th2 cells via the TCR is sufficient for production of IL-4 but not for proliferation of these cells. Proliferation of Th2 cells occurs only in the additional presence of a costimulatory signal delivered by IL-1. For the majority of Th2 cell clones, this type of proliferation was found to be independent of IL-4. Here, we further investigated the mechanism of IL-4-independent proliferation. We demonstrate that, after costimulation via TCR and IL-1R, but not via either receptor alone, Th2 cells are triggered to produce cell-associated IL-1alpha, as detected at the level of function, protein, and mRNA expression. In the presence of the TCR signal, autocrine IL-1alpha is then able to costimulate IL-4-independent proliferation of Th2 cells and to further enhance its own production. Thus, our results point to a novel, IL-4-independent, self-amplifying autocrine pathway of Th2 cell proliferation that requires a signal via the TCR and a costimulatory signal via IL-1R. This pathway may explain frustrating results in experimental models that attempted to treat established Th2-mediated diseases in vivo with IL-4-neutralizing agents alone.

Animals↗