Manfred P. Dierich

Neopterin as a marker for activated cell-mediated immunity: Application in HIV infection

Abstract The production of neopterin is closely correlated with activation of cell-mediated immunity. The increased neopterin concentrations appear to be produced by human macrophages specifically stimulated with gamma-interferon. As Helmut Wachter and colleagues review here, neopterin studies reveal that preactivation of cell-mediated immunity is associated with poor prognosis in cancer patients. In addition, in human immunodeficiency virus (HIV) infection, neopterin levels increase in parallel with progressive disease, are inversely correlated with CD4 + /CD8 + T-cell subset ratios and are of predictive significance.

Enhancement of sheep red blood cell human lymphocyte rosette formation by the sulfhydryl compound 2-amino ethylisothiouronium bromide.

Abstract In vitro treatment of sheep red blood cells with the sulfhydryl compound 2-aminoethylisothiouronium bromide (AET) under suitable experimental conditions greatly enhances their ability to form rosettes with human lymphoid cells. The effect of AET depends on the concentration, pH of the solution and time of incubation. AET treated sheep red blood cells lose their ability to form rosettes after varying time periods depending on the temperature of storage and on the concentration of the AET solution. Treatment of human peripheral lymphocytes with AET abolishes their capability to form rosettes with both normal and AET treated sheep red blood cells. Lymphocytes from some patients with chronic lymphocytic leukemia react differently with AET-treated sheep red blood cells than with normal sheep red blood cells.

Epidemiology and outcome of infections due to Aspergillus terreus: 10-year single centre experience.

Aspergillus terreus, a less common pathogen, appears to be an emerging cause of infection at our institution, the Medical University Hospital of Innsbruck. Thus the epidemiology and outcome of A. terreus infections over the past 10 years was assessed. We analysed 67 cases of proven invasive aspergillosis (IA) according to the European Organisation for Research and Treatment of Cancer/Mycoses Study Group criteria, investigated antifungal susceptibility of amphotericin B (AMB), voriconazole and caspofungin and performed molecular typing of A. terreus. Patients with proven IA caused by A. terreus (n = 32) and non‐A. terreus (n = 35) were evaluated. The two groups were comparable in terms of age, gender, underlying disease, antifungal prophylaxis and duration of neutropenia (P > 0·05). Leukaemia was the most common underlying malignancy. Fungal dissemination occurred in 63% of the patients. Aspergillus terreus infections were associated with a lower response rate to AMB therapy (20%), compared with 47% for patients with non‐A. terreus infections (P < 0·05). In vitro, A. terreus was found to be resistant to AMB and molecular typing discriminated between patients isolates, showing a high strain diversity with 26 distinct types (I‐XXVI) identified by combination of three primers. Aspergillus terreus infections displayed evidence of AMB resistance in vitro and in vivo and were associated with a high rate of dissemination and poor outcome; A. terreus causes systemic infections of endemic character in Tyrol, Austria. The onset of A. terreus infection depends not on the degree of immunosuppression but on environmental Aspergillus spp. exposure.

Uncaria tomentosa (Willd.) DC.—Ethnomedicinal use and new pharmacological, toxicological and botanical results

The medicinal system of the Asháninka Indians in Perú is portrayed. Three categories of medical disorders and healers are recognized. A human is viewed to consist of a physical and a spiritual being who communicate with each other by means of a regulating element. The significance of Uncaria tomentosa (Willd.) DC. (Rubiaceae), locally known as unã de gato, in traditional medicine is emphasized by its exclusive use by priests to influence this regulation. Pharmacological and toxicological results obtained with extracts or isolated compounds are summarized. Pentacyclic oxindole alkaloids stimulate endothelial cells in vitro to produce a lymphocyte-proliferation-regulating factor. Tetracyclic oxindole alkaloids act as antagonists. A significant normalization of lymphocyte percentage was observed in vivo although total leucocyte numbers did not change.

Human macrophages degrade tryptophan upon induction by interferon-gamma

Human peripheral blood mononuclear cells, monocytes-macrophages and T-cells were stimulated with human recombinant interferon-gamma, interferon-alpha and phytohemagglutinin. The culture supernatants were analyzed for tryptophan, kynurenine, 3-hydroxyanthranilic acid, anthranilic acid and neopterin by high performance liquid chromatography. Tryptophan was decreased and the four other compounds were increased in supernatants of peripheral blood mononuclear cells activated by interferon-gamma (250 U/ml), interferon-alpha (10.000 U/ml) and phytohemagglutinin (1 microgram/ml). After splitting of peripheral blood mononuclear cells by adherence, the monocytes and macrophages but not the T-cells degraded tryptophan upon stimulation by interferon-gamma in a dose dependent manner. Supernatants of phytohemagglutinin stimulated but not of resting T-cells were found to induce tryptophan degradation by macrophages, the active principle being neutralized by an antiserum for interferon-gamma. Thus phytohemagglutinin acts by activating T-cells to release interferon-gamma which in turn induces macrophages to degrade tryptophan. In all experiments the appearance of neopterin in the culture media was correlated to the observed tryptophan degradation.

Neopterin modulates toxicity mediated by reactive oxygen and chloride species

Neopterin, a pyrazino‐pyrimidine derivative, is synthesized in excess by human monocytes/macrophages upon stimulation with interferon‐γ, a cytokine derived from activated T‐cells. Neopterin is furthermore produced constitutively. A relatively constant ratio between neopterin and its reduced form, 7,8‐dihydroneopterin, has been described in human serum. In the study presented here we tested the ability of neopterin and its reduced form to modulate the effects of cytotoxic substances like hydrogen peroxide or hypochlorous acid and N‐chloramine derivatives. We show that 7,8‐dihydroneopterin potently reduces biological and chemical effects of these substances independently from the pH value. In contrast, at slightly alkaline pH (pH 7.5) neopterin enhances hydrogen peroxide and chloramine‐T activity. This is demonstrated by increase of signal intensity in a luminol assay and also by enhancement of toxicity towards bacteria. Thus, the macrophage derived substance neopterin is able both to enhance and to reduce cytotoxicity in dependence of pH value and its oxidation state, and it may have a pivotal role in modulation of macrophage mediated effector mechanism.

Shiga Toxin Activates Complement and Binds Factor H: Evidence for an Active Role of Complement in Hemolytic Uremic Syndrome

Infections with enterohemorrhagic Escherichia coli (EHEC) are a major cause of hemolytic uremic syndrome (HUS). Shiga toxins (Stxs), especially Stx2, are believed to represent major virulence factors of EHEC, contributing to HUS pathogenesis. Beside EHEC-associated HUS, there are hereditary atypical forms of HUS, which are mostly caused by mutations of complement regulators. The aim of the present study was to investigate whether or not complement is also involved in the pathogenesis of EHEC-induced typical HUS, by being activated either directly or indirectly by involvement of its inhibitors. Purified Stx2 markedly activated complement via the alternative pathway and was found to bind to factor H (FH), however, only when it was active. No apparent cleavage or destruction of FH was visible, and cofactor activity in fluid phase was unaffected, but clearly delayed for surface-attached FH, where it is essential for host cell protection. Binding studies using FH constructs revealed that Stx2 binds to short consensus repeats (SCRs) 6–8 and SCRs18–20, but not to SCRs16–17, i.e., to regions involved in the surface recognition function of FH. In conclusion, complement, and in particular FH, not only plays an important role in atypical HUS, but most probably also in EHEC-induced HUS.

Acquisition of host cell-surface-derived molecules by HIV-1

Objective: To determine the acquisition of host cell‐membrane‐derived molecules by HIV‐1 during the budding process, and to investigate whether the uptake of these molecules is cell‐type‐specific and selective. Design: Virions, propagated by four different cell types were analysed for the presence of adhesion molecules, glycosylphosphatidylinositol (GPI)‐anchored proteins and various cell‐surface markers. The pattern was compared with the phenotype of the HIV‐1‐infected cell. Methods: For phenotypic analysis of virions a two‐step assay was used. Int the first step, virus was captured with monoclonal antibodies (in some cases polyclonal sera) against different cell‐membrane proteins. In a second step, the presence of virus was measured by determining the concentration of the virus‐specific p24 core antigen. The expression of surface molecules on uninfected and HIV‐1IIIB‐infected cells was analysed by FACS. Results: Depending on the cell type used for virus propagation, different cell‐membrane molecules were found on the virus surface reflecting the corresponding cell type. The uptake of these molecules was selective to a certain degree. No CD4 and CD87 molecules were detectable on HIV‐1, although both molecules were present on uninfected and HIV‐1‐infected cells. CR3 and CDw108 could not be seen on uninfected cells, but were detectable on infected cells and virions. Conclusions: During the budding process HIV‐1 acquires a variety of cell‐type‐specific cell‐surface molecules. Certain cell‐membrane molecules become upregu‐lated during HIV‐1‐infection and are then found on virions, whereas other molecules remain on the cell surface and do not become incorporated.

Strain-Specific Differences in the Amount of Shiga Toxin Released from Enterohemorrhagic Escherichia coli O157 following Exposure to Subinhibitory Concentrations of Antimicrobial Agents

Abstract There is no consensus regarding the benefit versus harm of antibiotic therapy for treatment of disease due to enterohemorrhagic Escherichia coli O157. The effects in vitro of subinhibitory concentrations of 13 antimicrobial agents on the release of Shiga toxin (Stx) by three different Escherichia coli O157 strains expressing Stx 1 or Stx 2 either alone or in combination were investigated. The Stx-induced cell death of Vero cells was determined using a colorimetric assay based on the measurement of lactate dehydrogenase (LDH) released into the supernatant from the cytosol of damaged cells. Growth of all O157 strains in broth cultures containing subinhibitory concentrations of cotrimoxazole, trimethoprim, azithromycin, or gentamicin was accompanied by a marked increase in the release of Stx. Exposure to cefixime, ceftriaxone, or erythromycin caused a marked increase in the release of Stx by the O157 strain producing Stx 2 alone, but decreased toxin production was observed with the Stx 1 producer and the strain producing Stx 1 and Stx 2. Exposure to ampicillin caused increased Stx release in the Stx 2-producing strain but had no effect on Stx production in the other two test isolates. Exposure to penicillin G, streptomycin, ciprofloxacin, fosfomycin, or sulfamethoxazole caused an increase in toxin production in two of the three test strains in each case, while decreases were observed for the other isolates. The response of Escherichia coli O157 isolates to subinhibitory concentrations of antibiotics seems to be highly dependent on the nature of the strain involved.

Immune activation and the anaemia associated with chronic inflammatory disorders

Chronic inflammatory disorders are associated with an increased risk of patients developing anaemia. There is some evidence that cytokines released during cell‐mediated immune responses are capable of inhibiting bone marrow haematopoiesis. In vitro, interferon gamma and tumournecrosis factor alpha inhibit growth of erythroid precursor cells. The mode of action of these cytokines is probably associated with their antiproliferative capacity. Decrease of serum iron and increase of storage iron in patients appears to be a consequence of the defense strategy of macrophages during long‐lasting inflammatory disorders. Decreased serum iron correlates to decreased haemoglobin concentrations. In view of this, the development of anaemia seems likely to result from the altered iron metabolism induced by stimulated macrophages. Low haemoglobin levels and associated hypoxia up‐regulate the release of erythropoietin, which can explain why increased circulating erythropoietin is usually found in patients with anaemia.