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.

PCDD/F emissions from heavy duty vehicle diesel engines.

The currently available information on PCDD/F emissions from diesel vehicles is briefly surveyed. Considerable uncertainty is identified concerning the emissions from heavy duty diesel trucks which have been measured only twice so far. These measurements led to emission factors differing by a factor of 200; similar discrepancy was also revealed by measurements of ambient air in traffic tunnels.

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.

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.

Disulfide analysis reveals a role for macrophage migration inhibitory factor (MIF) as thiol-protein oxidoreductase.

The molecular mechanism of action of macrophage migration inhibitory factor (MIF), a cytokine with a critical role in the immune and inflammatory response, has not yet been identified. Here we report that MIF can function as an enzyme exhibiting thiol-protein oxidoreductase activity. Using a decapeptide fragment of MIF (MF1) spanning the conserved cysteine sequence motif Cys57-Ala-Leu-Cys60 (CALC), Cys-->Ser mutants (C57S MIF, C60S MIF, and C57S/C60S MIF) of human MIF (wtMIF), and alkylated wtMIF, we show that this activity is mediated by the CALC region and is important for the macrophage-activating properties of MIF.

The Th1/Th2 paradigm and experimental murine leishmaniasis.

In recent years, the Th1/Th2 concept has become of prime importance in the understanding of heterogeneous responses of the immune system and implications thereof for infectious and autoimmune diseases. Originally established on the basis of different cytokines produced by T cell clones, it is now known that the Th1/Th2 concept really defines totally different immune pathways that affect most if not all cells of the immune system. Murine experimental leishmaniasis was the first model to confirm the relevance of the Th1/Th2 concept in vivo.

Molecular characterization and functional analysis of murine interleukin 4 receptor allotypes.

The murine interleukin 4 receptor (IL-4R) exists as a transmembrane protein transducing pleiotropic IL-4 functions, or as soluble (s)IL-4-binding molecule with potent immunoregulatory effects. In this study we identified and characterized a murine IL-4R allotype. Sequence analysis of the IL-4R cDNA of BALB/c mice revealed 18 base substitutions leading to three extracellular and five cytoplasmic amino acid changes when compared with the published IL-4R sequence of C57BL/6 mice.

Structure activity studies of the cytokine macrophage migration inhibitory factor (MIF) reveal a critical role for its carboxy terminus.

Carboxy-truncated mutants of human MIF (MIF(1-104) and MIF(1-109)) were used in structure activity studies. CD spectroscopy revealed an overall structural similarity between the mutants and MIF. Denaturant-induced unfolding demonstrated that the C-terminus contributed significantly to the conformational stability of MIF.

A dominant mechanism coordinately suppresses the expression of Th2 lymphokines.

This study, we present evidence for a negatively acting control mechanism that coordinately suppresses the synthesis of the Th2 lymphokines IL-4, IL-5 and IL-6. This control mechanism operates in the murine thymoma cell line BW 5147. When cells of this line were fused to four independently established, well-defined Th2 cell clones, all resulting 74 lymphokine-secreting hybridomas secreted IL-2 which was not secreted by any of the parental Th2 cell clones.

Intracellular murine IFN-gamma mediates virus resistance, expression of oligoadenylate synthetase, and activation of STAT transcription factors.

IFN-gamma is a pleiotropic cytokine that plays a major role in anti-infectious immune responses. The physiologic effects of IFN-gamma are thought to be mediated by the binding of extracellular IFN-gamma to its receptor at the cell surface, thereby triggering an intracellular signaling cascade. In this work, we present evidence for a completely intracellular mechanism for IFN-gamma to induce virus protection.

Two distinct stimulus-dependent pathways lead to production of soluble murine interleukin-4 receptor.

The IL-4R exists in two forms, either membrane bound or as a soluble (s) molecule. Since the sIL-4R binds to its ligand with high affinity, thereby acting as an immunoregulatory molecule, we were interested in the processes leading to its release. First, the release of sIL-4R in the model of murine leishmaniosis was analyzed.

Invasion, control and persistence of Leishmania parasites.

Significant advances in research on the immunopathogenesis of leishmaniasis include the discovery of novel putative evasion and survival strategies of Leishmania parasites, a more detailed understanding of the function and regulation of interleukin-12, definition of molecules involved in cognate interaction between macrophages and T cells and new ideas concerning the mechanisms of host resistance and susceptibility. The use of transgenic mice for (re)probing certain immunological aspects of leishmaniasis has yielded not only predictable and confirmatory but also unexpected and pioneering results which require critical appreciation.

Expression and co-cytokine function of murine thioredoxin/adult T cell leukaemia-derived factor (ADF).

Human ADF (adult T cell leukaemia-derived factor), an isoform of thioredoxin, promotes proliferation of certain human lymphoid cell lines and is involved in many thiol-dependent reducing reactions. To study functional aspects of the murine homologue, we established inducible overexpression of murine ADF in E. coli and a purification method which led to an apparently homogeneous 14 kDa protein.

Stable transfection of cloned murine T helper cells.

Here we describe a protocol for the stable transfection of murine T helper (Th) cells and long term culture of the resulting transfectants. The electroporation protocol was established for the murine Th2 clone L1/1 by testing different parameters determining the electric field (capacitance, voltage, single or twin pulse) as well as the activation status of the cells. The transfected T cells were genetically altered by stable integration of the neomycin resistance gene, encoded in the vector pM5neo, into the genome.

Effect of IL-7 treatment on Leishmania major-infected BALB.Xid mice: enhanced lymphopoiesis with sustained lack of B1 cells and clinical aggravation of disease.

The Xid immunodeficiency was characterized by a total lack of B1 cells and reduced numbers and functions of B2 cells. In BALB.Xid mice, this defect results in an reduced susceptibility against infections with parasites such as Trypanosoma cruzi and Leishmania major.

Fibroblasts as efficient antigen-presenting cells in lymphoid organs.

Only so-called "professional" antigen-presenting cells (APCs) of hematopoietic origin are believed capable of inducing T lymphocyte responses. However, fibroblasts transfected with viral proteins directly induced antiviral cytotoxic T lymphocyte responses in vivo, without involvement of host APCs. Fibroblasts induced T cells only in the milieu of lymphoid organs.

Neutralization of endogenous IL-6 suppresses induction of IL-1 receptor antagonist.

IL-1 is a potent cytokine that promotes host defense and inflammation. These processes may be modulated by an IL-1 receptor antagonist (IL-1Ra) that binds to and blocks IL-1 receptors. The objective of this study was to define the cellular origin and regulation of IL-1Ra production during bacterial infection.

Stimulation of B-cell lymphopoiesis by interleukin-7 leads to aggravation of murine leishmaniasis.

The effect of recombinant interleukin-7 (IL-7) on the clinical course of murine leishmaniasis and the development of the accompanying immune response was investigated. Previously, IL-7 has been shown to possess stimulatory capacity for different cell types of the immune and haematopoietic system critically involved in the defence against Leishmania major (L. major), such as macrophages which are activated for the elimination of the parasite by IL-7.

Soluble IL-4 receptor, potential for therapeutic and prophylactic intervention.

Many bacterial, protozoal and viral infections trigger a cell-mediated immune response. Of special importance for the clinical outcome of disease, however, is the relative predominance of T helper (Th) cell populations (Th1 and Th2) secreting different patterns of lymphokines. Preferential development of one Th subset occurs apparent at the early stages of an infection, suggesting that the mechanisms driving the immune response in one direction or the other operate soon after exposure to the antigen.

Interleukin-10 inhibits antimicrobial activity against Leishmania major in murine macrophages.

The stimulation of macrophages is of importance to the defense against intracellularly replicating microorganisms such as Leishmania. In this study the direct effect of recombinant interleukin-10 (IL-10) on the leishmanicidal effector functions of murine peritoneal or bone marrow derived macrophages was investigated. IL-10 almost completely inhibited the killing of intracellular leishmania at concentrations above 10 ng/ml.

Recombinant soluble interleukin-4 (IL-4) receptor acts as an antagonist of IL-4 in murine cutaneous Leishmaniasis.

This study was performed to evaluate the soluble interleukin-4 receptor (sIL-4R) as a potential antagonist of interleukin-4 (IL-4) in an infectious disease. It is shown that antigen-triggered proliferation and cytokine secretion of Leishmania major-specific, cloned Th2 cells in vitro can be inhibited dose dependently by recombinant murine, but not control human, sIL-4R. In vivo, we found that endogenous synthesis of IL-4 mRNA is upregulated during the first week of infection, while an increase of IL-4R mRNA occurred later after infection of BALB/c mice with L.

Homodimeric murine interleukin-3 agonists indicate that ligand dimerization is important for high-affinity receptor complex formation.

Homodimeric murine interleukin 3 (mIL-3) agonists were generated by intermolecular cystine-bonding. Steady-state binding assays and association kinetics performed at 4 degrees C using these agonists revealed specific binding to both the high- and low-affinity receptor. DSS-mediated crosslinking studies performed at 4 degrees C with agonist concentrations compatible with high-affinity receptor complex formation allowed to detect protein complexes of the alpha chain, the beta chain(s) and the high-affinity receptor complex migrating with apparent molecular weights of 90 kDa, 140 kDa, and above 180 kDa, respectively.

Differential regulation of IL-9-expression after infection with Leishmania major in susceptible and resistant mice.

IL-9 is a pleiotropic lymphokine, one of its activities being the growth stimulation of certain CD4+ T lymphocytes. In murine cutaneous leishmaniasis, depending on the genetic background of the host mouse strain, vigorous proliferation of either mainly Th1 in resistant C57BL/6 mice or Th2-type CD4+ T cells in susceptible BALB/c mice occurs after infection with Leishmania major (L. major).

Cytokines in leishmaniasis: a complex network of stimulatory and inhibitory interactions.

The work of immunologists, cell biologists and parasitologists in the field of leishmaniasis has not only provided important insights into the immunopathogenesis of this disease, but also yielded fundamental contributions to our understanding of basic immunological phenomena and of host-parasite interactions. The ability of recombinant interferon-gamma to induce the microbicidal activity of phagocytes and the opposite effect of inhibitory cytokines was first demonstrated with Leishmania-infected macrophages. The selective development of protective and disease-mediating CD4+ T lymphocytes as well as their differential influence on the course of the disease has been long investigated in the murine Leishmania major model and now represents one of the best examples for the in vivo induction of type 1 versus type 2 T helper lymphocytes.

Interleukin-7 enhances antimicrobial activity against Leishmania major in murine macrophages.

Recently, it has been shown that interleukin-7 (IL-7) is able to induce secretion of cytokines and tumoricidal activity by human monocytes. This study shows that treatment of murine macrophages infected with Leishmania major with IL-7 without any other stimulus reduced the percentage of infected cells, as well as the parasite burden per cell, in a dose-dependent manner to a limited degree (45% reduction of the number of amastigotes per 100 macrophages). Simultaneous treatment of macrophages with gamma interferon and IL-7 led to nearly complete (> 99%) elimination of amastigotes.